Manufacturers' support policies.

Manufacturers' support policies. Choosing an effective plan for supporting a medical device is critical to its safe use, cost-effectiveness, and longevity. Hospitals can choose from a variety of support providers, including manufacturers, third-party service vendors, or hospital clinical engineering (CE) departments. However, if the hospital plans to use a third-party service vendor or its own CE department to provide support, the manufacturer's cooperation or assistance will still be needed to implement the support plan effectively. Over the years, ECRI has received many comments from hospitals about the way in which manufacturers respond to their equipment support needs. We have learned that some manufacturers are not willing to assist third-party service vendors or in-house service programs or do not always deliver the support they promise. Also, hospitals do not always consider their support needs before purchase, when they have the most leverage to negotiate flexible support arrangements. To help foster better equipment support and customer satisfaction, we polled manufacturers that have participated in recent Health Devices Evaluations to obtain detailed information about their policies toward manufacturers' contract, third-party, and in-house support. Ready access to this information will help hospitals evaluate whether manufacturers' support policies will meet their needs, and it will allow them to minimize problems by working with the manufacturer to negotiate optimal support arrangements during the purchase process. In this article, we briefly discuss the factors to consider when evaluating support alternatives and manufacturers' support policies. We also present the questions posed to each manufacturer on our Manufacturers' Support Policies Questionnaire, along with a summary of the responses that we received for each question.(ABSTRACT TRUNCATED AT 250 WORDS)


IV free-flow--still a cause for alarm.

IV free-flow--still a cause for alarm. A nurse's aide, in transferring a mother in labor to the delivery room, turned off the infusion pump delivering Pitocin, a drug administered intravenously to accelerate contractions. The aide removed the infusion set from the pump without first closing the manual clamp on the line. A free-flow infusion occurred, and the mother received nearly 35 times the prescribed amount of drug. The infant suffered organ damage and pneumonia and died four days later. Free-flow infusions can have tragic consequences when a potent drug is involved. Although other causes of overinfusion and free-flow exist, such incidents are typically associated with removing a disposable intravenous (IV) infusion set from an infusion device without first closing the manual clamp. We first raised this issue in our 1982 Evaluation "Infusion Controllers" and have emphatically and repeatedly addressed it in Health Devices and other ECRI publications. Yet, hospitals continue to report free-flow infusions, a problem that can be addressed by both hospitals and device manufacturers. In this article, we describe the causes of free-flow--both user error and device design; report numerous incidents, some resulting in death; and provide recommendations for reducing the likelihood that such problems will continue to occur.


A bifunctional HBED-derivative for labeling of antibodies with 67Ga, 111In and 59Fe. Comparative biodistribution with 111In-DPTA and 131I-labeled antibodies in mice bearing antibody internalizing and non-internalizing tumors. To investigate whether bifunctional ligands containing chelating structures other than EDTA and DTPA and metallic radiotracers other than 111In will reduce the non-specific radioactivity uptake in the liver during immunoscintigraphy, we synthetized an isothiocyanato-substituted phenolic polyaminocarboxylic acid (HBED-CI) for labeling of MAbs with 67Ga, 111In and 59Fe. Biodistribution of HBED-CI-labeled MAbs was compared to that of 131I and 111In-DTPA labeled MAbs in nude mice bearing tumors, which differ with regard to intracellular internalization and catabolism of the corresponding MAb-antigen complex. In the liver a continuous radioactivity excretion for 67Ga-HBED-CI-labeled MAbs was observed with kinetics that parallel 131I clearance after administration of 131I-MAbs, while 111In-HBED-CI-labeling led to a constant 111In liver level quite similar to that of 111In-DTPA-MAbs. In tumors, 67Ga-HBED-CI-MAb uptake again paralleled that of 131I-MAbs, showing continuous accumulation in tumor tissues when internalization of the MAb-antigen complex was not involved. A much lower uptake, which peaked between 24 and 48 h, was found in the case of MAb-antigen internalization. 111In of 111In-HBED-CI- and 111In-DTPA-labeled MAbs continuously accumulated in both types of tumors. Compared with 111In-DTPA-MAbs, an improvement in tumor-to-liver ratios, due to the reduced liver radioactivity associated with 67Ga-HBED-CI-labeled MAbs, could only be obtained with non-internalizing tumors. The time course of radioactivity distribution in the liver and in MAb-internalizing tumors after administration of 67Ga-HBED-CI-, 111In-HBED-CI- and 111In-DTPA-labeled MAbs further indicates a dominating influence of the metallic radiotracer rather than the ligand on retention or excretion of radioactivity in MAb-catabolizing tissues.


Attachment of 99mTc to lipid vesicles containing the lipophilic chelate dipalmitoylphosphatidylethanolamine-DTTA. The binding of 99mTc to negatively-charged and neutral unilamellar lipid vesicles was investigated in the absence and presence of the ligand diethylenetriaminepentaacetic acid (DTPA) covalently attached to the headgroup of phosphatidylethanolamine at the surface of the membrane. Even in the absence of DTPA on the membrane surface, 99mTc reduced by Sn bound to the membrane surface but rapidly dissociated from the vesicles in the presence of plasma in vitro. When DTPA was present on the membrane surface, dissociation of 99mTc from the vesicle surface in plasma was much reduced. The dissociation of 99mTc from the surface of negatively-charged vesicles was less than for neutral vesicles in the absence of ligand but was markedly greater than for vesicles containing the ligand DTPA, suggesting that the binding of 99mTc to vesicles with surface-attached DTPA could not be explained solely on the basis of the negative charge provided by the DTPA. In vitro experiments using 14C-labeled lipids as well as in vivo imaging studies indicated that dissociation of 99mTc from the surface of the vesicle did not arise predominantly because of lipid exchange with plasma components or due to cleavage of Tc-DTPA from the vesicle surface. For vesicles with surface-attached DTPA, 99mTc dissociation from the vesicle surface in plasma was further reduced by addition of the antioxidant ascorbate.


Evaluation of S-[11C]citalopram as a radioligand for in vivo labelling of 5-hydroxytryptamine uptake sites. The biologically active S-enantiomer of [N-methyl-11C]citalopram was evaluated as a radioligand for in vivo labelling of the 5-hydroxytryptamine uptake site in brain, using ex vivo tissue counting in rats and positron emission tomography in man. In rats, the maximal signal for total versus non-specific binding was approx. 2 at 60-120 min after radioligand injection. Subsequent studies in man failed to identify a specific signal over a 90 min scanning period, due to prolonged retention of non-specific label.


Ligands and tracers for PET studies of the 5-HT system--current status. The status of the radiochemical development and biological evaluation of radioligands and tracers for PET studies of the serotonergic system is reviewed, indicating those agents with present value and those with future potential. Practical recommendations are given for the preparation of two useful radioligands for PET studies of central 5-HT2 receptors, namely [18F]setoperone and [18F]altanserin. Though, it has not proved possible to recommend tracers or radioligands for the study of other aspects of serotonergic system, prospects for future radiochemical development are indicated, especially for developing radioligands for the 5-HT re-uptake site, and for the 5-HT1 and 5-HT3 receptors.


(S)- and (R)-[11C]nicotine and the metabolite (R/S)-[11C]cotinine. Preparation, metabolite studies and in vivo distribution in the human brain using PET. In order to investigate [11C]nicotine binding and metabolism in the living human brain by PET, routine protocols were developed for the preparation and purification of (S)- and (R)-[11C]nicotine and the metabolite (R/S)-[11C]cotinine. (S)- and (R)-[11C]nicotine were prepared by N-methylation with [11C]methyl iodide of the appropriate secondary amine, which was liberated in situ by 2,2,6,6,-tetramethylpiperidine (TMP) from its corresponding biscamsylate-salt. (R/S)-[11C]Cotinine was prepared by N-methylation of the amide precursor using tetrabutylammonium hydroxide as a phase transfer catalyst. Straight-phase semipreparative HPLC was in all purifications found to be superior to reversed-phase since the contamination by the norcompounds was eliminated. Reaction in acetonitrile for both (S)- and (R)-[11C]nicotine (5 min, 130 degrees C) and (R/S)-[11C]cotinine (1 min, 80 degrees C) with subsequent straight-phase HPLC purification resulted in 35-45% radiochemical yield (from EOB and decay-corrected) with a total synthesis time of 30-35 min, a specific radioactivity of 1000-1500 Ci/mmol (37-55 GBq/mumol, EOS) and a radiochemical purity > 99%. The uptake and distribution of these tracers in the human brain was studied in healthy volunteers by PET. The metabolite (R/S)-[11C]cotinine did not cross the blood-brain barrier to any significant degree. The amount of the total radioactivity representing (S)-[11C]nicotine measured in plasma by HPLC was 75% at 4 min and 25% at 50 min.


Photoreactive 111In-cyclodextrin inclusion complex: a new heterobifunctional reagent for antibody labeling. The compound of interest, N-5-azido-2-nitrobenzoylaminomethyl-111In-acetylacetone-alpha-cycl odextrin (CD) (V) was synthesized by the selective tosylation of alpha-CD to form 6-tosyl-6-deoxy-CD, which was then reacted with NaN3 to form 6-azido-6-deoxy-CD (II). This was followed by catalytic hydrogenation to yield III. Compound III and 111In-acetylacetone were mixed to form an inclusion complex, which was then reacted with N-5-azido-2-nitrobenzoyloxysuccinimide to yield compound V. Anti-melanoma MAbTP41.2 was added to compound V, followed by immediate photoreactivation labeling by u.v. light at 320 nm. The final product VI was purified from a Sephadex G-50 column. 111In-DTPA-MAbTP41.2 was also prepared as a control. Immunoreactivity via the cell-binding assay of VI was 87%, compared with 57% by the BADTPA method. Biodistribution in non-tumor rats yielded a liver concentration in %ID/g of 3.5, 1.7 and 1.0 for compound VI, compared to the 5.5, 5.2 and 3.1 for the BADTPA compound, at 4, 24 and 48 h post-injection, respectively.


[The entrapment syndrome of the popliteal artery: diagnosis, therapy, results]. The popliteal artery entrapment syndrome is caused by an anatomical anomaly in the popliteal fossa. Mostly the popliteal artery is compressed by a faulty insertion of the medial head of the gastrocnemius muscle. The main symptom is an intermittent claudication at an early age. We review our experience with four typical cases. All patients were examined because of a unilateral sudden claudication in the calf at ages between 23 and 43 years. In two patients we could find angiographically a typical stenosis, in two others a complete occlusion of the popliteal artery. Additionally a female patient showed a 6 x 8 cm poststenotic aneurysm in the popliteal fossa. Therapeutically we restored the flow of the artery on two patients by dividing the compressing gastrocnemius muscle. As a late complication two years after the first operation one of these patients had an occlusion of popliteal artery which was bypassed by a popliteo-crural vein graft. On two patients we primary inserted a vein graft. All patients were checked 6 months till 5 years after the first operation by duplex scanning, by measurement of pressure at the ankle and by pulse volume recording at the big toe. All our postoperative examinations demonstrate patency of the repairs and the patients have very few problems.


[A new intraluminal endarterectomy instrument]. In 40 human cadaver arteries endarterectomy was performed using a new intravascular device. The system and technique are described. According to our preliminary laboratory experiences the new device proofed to be save and effective. It will allow us to approach more complex lesions not ideal for balloon angioplasty. Clinical investigations will be performed in the near future.


[Axillo-femoral bypass: indications and results]. Axillofemoral bypass grafting is an accepted method for the treatment of vascular insufficiency of the lower limb. It is especially useful in high risk patients whose conditions preclude aortofemoral bypass grafting. The goal of our study is to evaluate whether the benefit of this operation outweigh the potential risks in the age group "sixty-plus". We performed 23 axillofemoral bypass grafting procedures (axillofemoral in 20, axillobifemoral in 3 cases) mainly be mean of 8 mm Dacrom prosthesis. The 30-day operative mortality rate was 8.7% and the cumulative 5-year patient survival rate was 52.2%. Overall the 1- and 5-year life table patency rate were 80% and 64%, respectively. The cumulative limb salvage was 90% and 72% at 1 and 5 years. We obtained an average reduction in Fontaine classification of 2 stages in the first year and 1.3 stages after 5 years. The comparison of the cumulative survival rate with the limb salvage rate indicates that advanced age should not be considered a contraindication to performing an axillofemoral bypass graft. The reduction in Fontaine stage means a remarkable improvement in quality of life.


[Anomalies of immunoglobulin synthesis in vitro in surgical patients]. It is known that blood mononuclear cells from surgical patients produce large amounts of IgG in vitro, and that this synthesis is not increased by pokeweed mitogen stimulation. The objective of the present study was to determine if this abnormal pattern of IgG synthesis extended to other immunoglobulin classes. Patients were stratified according to delayed type hypersensitivity as reactive (induration > 5 mm to > one skin test antigen), or anergic (no response). Healthy laboratory personnel were studied as controls. Mononuclear cells were cultured for 10 days, without or with pokeweed mitogen. IgG, IgM, and IgA in supernatant was measured using a competitive solid phase radioimmunoassay. Unstimulated IgG and IgA synthesis was increased in surgical patients, especially in those with reduced delayed type hypersensitivity responses. IgM synthesis was normal or low. With mitogen stimulation, normal increases in IgG synthesis were seen in control and reactive subjects. In anergic patients, IgG was not increased by pokeweed mitogen. For IgM, mitogen stimulation increased synthesis but to a lesser degree in patients than in controls. For IgA, synthesis was increased by mitogen stimulation in all groups. In conclusion, surgical patients show abnormalities in the pattern of spontaneous and mitogen stimulated immunoglobulin synthesis. The pattern varies for each isotype. B cell activation is present in surgical patients, especially those with reduced delayed type hypersensitivity. This suggests either recent antigenic stimulation, or non-antigen specific dysregulation of the B cell system in surgical patients.


[Eosinophilic gastroenteritis: two case reports]. Eosinophilic gastroenteritis is a rare lesion that may present a diagnostic problem to the surgeon. It may be confused with a malignant tumor because it often presents with intestinal obstruction. We treated two cases at the Hôpital de zone de Morges. In one case the lesion was in the gastric antrum and in the other the ascending colon. Only 27 cases of colonic eosinophilic gastritis are described in the medical literature.


[Meckel's diverticulum: surgical complications]. A 20-year (1971-1990) retrospective follow-up of all patients with a Meckel diverticulum (MD) out of the Kantonsspital and St. Claraspital Basel is reported. There were 57 men and 34 women with a mean age of 46.7 years (14-89). 60.4% of the cases were occasional findings, 39.6% were symptomatic MD. During the same period of time 5000 appendectomies were performed (ratio diverticulectomy to appendectomy was 1:50). 38.5% of the resections of MD were done when an acute appendicitis was supposed. In one third of the cases the MD was responsible for the symptomatology. Histological findings were 13% unspecific inflammation, 10% perforation, 9.8% ectopic tissue, 6.5% bleeding and 4.3% tumors. 8 out of 36 patients with pathological findings showed postoperative complications (21.6%), the mortality rate was 8.1%. In 55% resected MD without any additional histological findings only one complication (postoperative ileus) (1.9%) was found; mortality was zero per cent. The analysis of these data shows that complications of MD does not only appear in young patients. Complications occur more often in patients with MD with additional ectopy. We recommend the resection of an incidentally discovered Meckel diverticulum.


[Massive hemorrhage from presacral veins during resection of the rectum]. During the last 53 abdominoperineal amputations of the rectum for carcinoma, we had to control 5 massive hemorrhages. 4 of which were due to a lesion of the presacral venous plexus and caused a mean blood-loss of 4750 ml. During a "normal" Mile's operation our 49 other patients lost an average of 1750 ml (median 1200 ml) of blood. The negative effects of this type of complication and the following need for massive transfusion is not only local, circulatory and pulmonary, but also due to the immunodepressive effects of massive transfusion. We describe the different methods of control of these hemorrhages and insist on the advantage of using sterilised metallic thumbtacks, a method which was described 1984 by doctor Wang Qinyao of Shanghai.


[Severe necrotizing fasciitis].

[Severe necrotizing fasciitis]. The pathophysiology of necrotizing fasciitis remains unclear in patients with no apparent immunologic disorders. Between 1987 and 1990 we treated six patients with necrotizing fascitis and septic-toxic multiple organ failure, three patients survived. The mean age was 38 years (25-62). In all patients the primary bacteriological examination revealed streptococcus. Between the first symptoms and an adequate therapy were 4 days in surviving patients and 7 days in patients who died. Four patients showed spread of the gangrene into the adjacent tissue: muscles (n = 3), bowel (n = 2), mediastinum (n = 1). Adequate débridement was not possible or not performed in patients with spread into the abdominal cavity or the mediastinum. These patients did not survive. The duration of intensive care treatment in surviving patients were 14 to 78 days. We conclude that survival of patients with severe necrotizing fasciitis is influenced by the delay before adequate treatment, the localisation of the gangrene and intensive care facilities.


Detection of aneuploid subpopulations in colon cancer biopsies by cultivation in vitro prior to flow cytometric analysis. Flow cytometry offers a relatively simple method to determine the DNA content of colorectal cancers. However, the technique can not discover minor changes in karyotype which may characterize near-diploid tumors, nor will it detect very small populations of aneuploid cells. In an attempt to improve the detection of such cells, we incubated biopsy material prior to analysis. This method revealed an aneuploid subpopulation in one of two biopsies studied. This simple protocol may ultimately lead to a more accurate assessment of the ploidy in colon cancer.


[Callus distraction in traumatology: concepts and indications]. The understanding of the conditions under which bone and soft tissue can be regenerated by distraction has opened a new era in the field of limb reconstruction. Success in lengthening for posttraumatic limb length discrepancy has led to different applications of the distraction techniques. We present an overview of the clinical new possibilities, the strategy, the techniques and the complications using the distraction technique in the trauma patient. The optimal biologic method (limb distraction after resection or callus distraction by a transport technique) and the optimal fixation devices depend on the indication (local aspect, polytrauma, patient, motivation).


[Definitive surgery in complicated gastroduodenal ulcers]. Indications for surgical therapy in uncomplicated peptic ulcer disease have decreased considerably since the introduction of H2-receptor blocking drugs and more recently omeprazole. On the other side, the number of acute complications such as perforation or hemorrhage has remained nearly constant. The recent literature seems to indicate that the pattern of patients presenting with complications has changed and that the number of acute ulcers has increased. In a review of 283 patients, we found 150 perforated ulcers (PU) and 133 bleeding ulcers (BU). Almost all the patients with PU and 70% of the patients with BU have been treated operatively. The mortality is 14.3% and 12.5%, respectively. The vast majority of our patients have chronic ulcers, and only 7% have acute or subacute lesions confirmed by histologic examination. Based on our experience and the literature, we propose a therapeutic algorythm for these two conditions.


[Technique and organization of heart removal from the multi-organ donor]. The orthotopic heart transplantation is an accepted treatment for terminal cardiac disease. The technique of heart procurement and preservation is explained and the primary graft function in 108 subsequent heart transplantations is assessed. The mean ischemia time is 41 +/- 10 min in local, 98 +/- 19 min in distant (< 100 km) and 114 +/- 16 min in distant (> 100 km) organ procurement. Our method of preservation consists of cold cardioplegic arrest with potassium (30 mEq/L) cardioplegic solution. The incidence of the indication for high dose katecholamine-treatment after surgery and the maximal creatininekinase levels rose with ischemia time. All hearts recovered within a few days and the stay in the intensive care unit was not prolonged. We conclude that the heart preservation with cold cardioplegic arrest results in a good primary graft function. It is important to keep the ischemia time as short as possible.


[Bursitis iliopectinea--a rare differential diagnosis of painful inguinal swelling]. The syndrome of iliopectineal bursitis is an important differential diagnosis of a painful swelling in the inguinal region. Symptoms are local swelling and pain, radiation of pain along the femoral nerve, troubles of arterial or venous circulation and even dysuria or dysmenorrhoea in case the bursa penetrates into the pelvis. The diagnosis is established by conventional radiology (signs of osteoarthritis as one of the pathogenetic reasons), ultrasound (liquid mass lateral to the femoral vessels), punction (clear fluid, eventually synovial cells), contrast injection (dimension of the bursa and communication to the hip joint) and computed tomography (dimension, relation to hip and vessels). The treatment is the excision of the bursa, if conservative therapy is not successful. The excision requires tedious, cautious dissection because of the important structures adjacent and a possible communication to the hip joint must be eradicated.


[Experimental atrophy/hypertrophy complex of the liver after portal and/or biliary ligation in the rat]. Patients with lobar or segmental, benign or malignant strictures of the biliary tree (with or without impairment of the portal blood flow) show a considerable atrophy of the involved area of the liver with a compensatory hypertrophy/hyperplasia of the remaining non affected liver. To investigate the importance of the biliary and portal obstruction for the development of this process, we used a rat model, with selective biliary and/or portal ligation of the anterior liver lobes (two thirds of the liver mass). Weight measurements, morphometry, functional scintigraphy (Hepatoiodida-scan) and blood enzyme analyses were done immediately postoperatively, at 30 hours and 4, 8 and 28 days after the operation. The major findings were: 1. 28 days after biliary and/or portal ligation there was no difference between the body weight of the four groups, all ligated animals having compensated for an initial greater percentage body-weight loss. 2. Total liver weight remained constant, while atrophy and hypertrophy/hyperplasia occurred although a progressive derangement of liver morphology was observed during that time. 3. A severe atrophy-hypertrophy-complex (AHC) developed after selective portal ligation, which in our experiment did not appear after selective biliary ligation. 4. Morphometrical changes after selective biliary ligation were reversible, whereas in liver lobes with selective portal ligation a progressive parenchymal destruction and involution with subsequent impairment of hepatic function of the concerned lobe was observed.


[Acute abdomen in abdominal wall hemorrhage in anticoagulation]. 10 patients with a history of systemic anticoagulation presented with an acute abdomen which proved to be due to a rectus sheath hematoma. Four patients gave a history of antecedent trauma. The prothrombin time was prolonged beyond the therapeutic range in 8 patients and therapeutic in two. The diagnosis of rectus sheath hematoma was made by non-invasive imaging (sonography [n = 8], CT-scan) in 9 patients and by operative exploration in one. 8 patients were treated non-operatively and one patient required a delayed evacuation of the infected hematoma. One patient died from myocardial infarction. We conclude that a careful history, physical examination and prothrombin time, in combination with radiologic evaluation (sonography, CT-scan) can confirm the suspected diagnosis of rectus sheath hematoma. It is important to think of this specific hemorrhagic complication induced by anticoagulation to prevent unnecessary surgery.


[Necrosis of the terminal ileum after drug poisoning. Case report]. In an attempt to commit suicide, a 32-year-old women swallowed a vast amount of psychiatric drugs, i.e. tranquilizers, amphetamines, hynotic and antidepressant agents. By intensive care, using high doses of catecholamines and appropriate antidota, satisfactory circulation and oxygenation could be maintained. 3 days after admission a peritonitis became apparent. A 50 cm long section of the distal ileum was found to be completely necrotic and had to be resected. However, circulation of the correspondent mesenterium was not disturbed at all. A drug-induced non-occlusive intestinal ischemia was postulated to be the pathophysiological mechanism of intestinal necrosis. Non-occlusive intestinal ischemia is rare; it has been reported in young adults intoxicated by cocaine or phenobarbital, in children with high overdosage of iron compounds, in elderly individuals suffering from low-output congestive heart disease and in patients treated with digitalis drugs, with or with or without overdosage.


[Gardner syndrome: possibility of early detection of carriers by examination of the ocular fundus?]. Through indirect ophthalmoscopic examination it was possible to analyse three families originated from the same Swiss alpine valley (Poschiavo) and carrying Gardner syndrome (SdG) to check if their members with SdG were presenting hypertrophic changes of the retinal epithelium which is supposed to be an early symptom of that syndrome. This lesion was found to be present only in two out of 11 patients (18%) with at least one symptom of SdG. Because of its lacking sensitivity, ophthalmoscopic examination is not able to substitute colonoscopy as a valid screening method to find out which members of a SdG-family are at risk to develop colonic poliposis and cancer.


[E. Delorme correction of rectal prolapse]. Rectal prolapse is a distressing condition. Among the multiple corrections described, the perineal approach has generally been abandoned. However, Delorme's procedure is a good, simple and efficient intervention with a low morbidity for certain elderly selected patients. The purpose of this paper is to recall the details of this technique and review its results.


[Radiation-sparing intramedullary nailing with a radiolucent drilling system]. The indications for intramedullary fixation of tibial and femoral shaft fractures markedly increased by the introduction of interlocking techniques. Distal interlocking however remains a major problem although a variety of different aiming devices have been proposed. To reduce the radiation exposure both of the surgeon and the patient the AO has produced a radiolucent drive which allows easy aiming and drilling with a minimum of X-ray exposure.


[Technique of intraoperative laparoscopic cholangiography]. Since laparoscopic cholecystectomy has been established at our institution, the routine use of intraoperative cholangiography was abandoned but preoperative intravenous cholangiography and sonography are mandatory. However, these investigations are not always conclusive and in some cases not applicable. We have therefore started to use selective intraoperative laparoscopic cholangiography for laparoscopic cholecystectomy. Our technique and the materials required for this peroperative investigation are described in the present article.


[General aspects of kidney removal from donors with cardiovascular arrest]. The increasing list of patients waiting for an organ transplantation shows clearly the shortage of donor organs at disposal. This shortage is particularly high in the field of kidney allotransplantation. As the nephrectomy in non-heart beating (NHB) donor is technically practicable in most of our hospitals, the aim of this contribution is to pass on further informations about the nephrectomy to those surgeons who are interested in the procedure. The first part of the paper explains the different facts of the end stage renal disease in Switzerland and of transplantation activity. We focus then on the conditions and preparations of the nephrectomy in NHB donor, the exact technique of the operation and the organ perfusion. As it is shown by different studies, the long-term results of NHB donor kidneys are good. The nephrectomy in NHB donor is an effective method to solve kidney shortage.


HB Al-Ain Abu Dhabi [alpha 18(A16)Gly----Asp]: a new hemoglobin variant discovered in an Emiratee family. During a routine program of hemoglobin screening performed in the United Arab Emirates, we observed an electrophoretically fast-moving variant in a 9-month-old girl and in several members of her family. The structural determination, performed by reversed phase high performance liquid chromatography and amino acid sequencing, revealed a new variant that we named Hb Al-Ain Abu Dhabi [alpha 18(A16) Gly----Asp]. Its functional properties were normal.


Rapid molecular diagnosis of hemoglobin variants by RT-PCR of reticulocyte mRNA and direct sequencing. We have developed a rapid and simple approach for the molecular characterization of hemoglobin variants by a one-step reverse transcription-polymerase chain reaction of reticulocyte mRNA and direct sequencing of the product. This method can selectively amplify the alpha 1- or alpha 2-globin gene or the beta-globin gene transcript. The amino acid substitution of Hb G-Taichung is due to a G----C mutation at codon 74 of the alpha 1-globin gene, that of Hb J-Meinung to a G----A substitution at codon 56 of the beta-globin gene, and that of Hb Kaohsiung (or New York) to a T----A substitution at codon 113 of the beta-globin gene. The amplified segment encompassed the sequence from upstream of the initial codon behind the Cap site to downstream of the terminal codon before the polyadenylation addition signal. Hence, all hemoglobin variants should be able to be characterized by this approach.


Estimation of changes in genetic parameters in selected lines of mice using REML with an animal model. 1. Lean mass. Analysis was undertaken using Restricted Maximum Likelihood (REML) with an animal model of the results of selection for 20 generations for predicted lean mass in 10-week-old male mice. There were three replicates, each comprising high, low and unselected control lines. The overall estimates of heritability (h2) and common environmental correlations (c2) from results of the first seven generations were 0.51 +/- 0.03 and 0.21 +/- 0.01, respectively. Analyses of data from different lines and different numbers of generations were undertaken but with all pedigrees and data included, which enabled inferences to be drawn on changes in variance that were not due simply to inbreeding or short-term effects of selection. Estimates of h2 were lower in selected lines than the control, increasingly so in later generations, indicating departure from the infinitesimal model assumption of unlinked additive genes each of very small effect. In addition, values of c2 became higher in high than in control or low selected lines.


Estimation of changes in genetic parameters in selected lines of mice using REML with an animal model. 2. Body weight, body composition and litter size. Restricted Maximum Likelihood (REML) with an animal model was used to estimate genetic parameters of body weight, body consumption and litter size of lines of mice selected for 20 generations on an index of lean mass at 10 weeks in males, highly correlated with body weight, and for a further 18 generations on body weight at 10 weeks in males and females. Univariate and multivariate estimates of heritability were about 0.5 and those of common environment correlations were about 0.25 for both body weight and composition. Body weight and fat pad weight had genetic and phenotypic correlations of about 0.5. The heritability estimate of litter size was about 0.15 from univariate analysis, rather lower from multivariate, and the estimate of its genetic correlation with body weight was about 0.25. There were reductions in heritability of both body weight and litter size in later generations, even though full pedigrees were fitted and inferences made to the base population, but a plateau in response to selection for increased body weight could not be explained by a complete attenuation of genetic variance.


Genetic determinism of the cellular immune reaction in Drosophila melanogaster. Larvae of Drosophila melanogaster produce a haemocytic reaction against eggs of the parasitoid Leptopilina boulardi, which leads to the formation of a multicellular capsule surrounding the foreign object. Melanization resulting from the conversion of phenol to o-quinones frequently accompanies the cellular reaction. Although various cytological and biochemical aspects of this reaction have been investigated, very little is known about genetic determinism of the insect immune response. The heredity of the capacity to encapsulate was analysed by comparing 16 reciprocal crosses made using inbred resistant and susceptible parental strains. We conclude that differences in the encapsulation capacity of D. melanogaster are inherited autosomally, with the reactive phenotype showing complete dominance over the non-reactive one. There were neither sex-linked nor maternal effects. The results of all crosses suggest a single major segregating locus with two alleles and complete dominance of the resistant allele, with cytoplasmic factors and minor modifying genes acting on the major locus.


Genetic basis for female receptivity in Drosophila melanogaster: a diallel study. Receptivity of sexually mature, virgin females, measured as the time to copulation, has been investigated. Two independent 5 x 5 diallel crosses were carried out, each one with a different male tester genotype. Interactions between male and female genotypes were detected because some lines appeared as dominant in one diallel and recessive in the other. The overall picture emerging from the analyses shows a genetic system for female receptivity characterized by additive genes, with dominance for high receptivity in one of the diallel crosses. This remarkable additive component does not support the action of natural selection favouring females with extreme values of receptivity, rather, the lack of intrasexual female competition for mates is suggested.


Loss of a paternal chromosome causes developmental anomalies among Drosophila hybrids. Hybrids between Drosophila virilis and D. lummei suffer from developmental anomalies. Previous reports also suggest that these hybrids lose the D. lummei sixth chromosome early in development. Genetic and cytological data presented here confirm the loss of the microchromosome from both the soma and the germ-line of these hybrids and provide strong evidence that this loss causes the hybrid developmental anomalies.


vestigial suppressor genes and resistance to aminopterin in Drosophila melanogaster. We have shown that the vestigial (vg) mutant of D. melanogaster has a perturbed nucleotide metabolism compared to various wild-type strains. The mutant is particularly spontaneously resistant to aminopterin. The resistance seems to correlate with an increase in dihydrofolate reductase (DHFR) activity and quantity. The DHRF is a target enzyme of aminopterin. Our results suggest that the vg+ gene could be a regulatory gene acting on the DHFR gene. The wing mutant phenotype being due to a decrease in the thymidylate pool (dTMP) (Silber et al., 1989). In order to understand better the action of the mutant gene on nucleotide metabolism, we have induced suppressor genes of the mutant phenotype by mutagenesis with ethyl methanesulfonate (EMS) and bromouridine (BUR). The suppressor strains obtained display a phenotype intermediate between wild-type and vg phenotype. The action of three independent suppressor genes on eight parameters of nucleotide metabolism is reported here [three enzyme activities, resistance to aminopterin and to fluorodeoxyuridine (FUdR), auxotrophy test and the ability to use exogenous thymidine and uridine]. In comparison to the original vg strain, major changes for the parameters tested are observed. The most striking effects are obtained with the vgBUR27 strain, which is highly sensitive to aminopterin and to fluorodeoxyuridine and didplas the highest thymidine kinase (TK) and DHFR activities within the strains tested. The potential actions of suppressor genes on the vg mutant are discussed.


Therapeutic options for localized carcinoma of the prostate: the role of external beam radiation therapy. During the last three decades the use of ionizing radiation, both external beam radiotherapy and interstitial radionuclide implant (brachytherapy), has greatly increased for the treatment of cancer of the prostate. The increased use of radiation therapy is in part due to the technological advance of high-energy megavoltage units as well as the steadily improving long-term results of radiotherapy. Because of the comparable tumor control rates, patients with early stage prostate cancer have several therapeutic options for curative treatment including radical prostatectomy, external beam radiation therapy, and interstitial brachytherapy. The role of external beam radiotherapy is discussed in terms of the primary management of early and locally advanced prostate cancers as well as the treatment of residual or recurrent disease after prostatectomy. A new approach combines antimitotic chemotherapeutic agents and radiation therapy for the treatment of locally advanced cancers of the prostate.


Radical surgery in the treatment of localized carcinoma of the prostate. New methods of early detection combined with recent advances in surgical techniques have resulted in more patients undergoing radical surgery for treatment of localized carcinoma of the prostate. Over 350 radical prostatectomies have been performed by our group since January 1987. We review the role of radical prostatectomy in the treatment of prostate cancer and our experience with 100 patients undergoing radical retropubic prostatectomy since the advent of nerve-sparing techniques to preserve potency.


Management of impotence after treatment of carcinoma of the prostate. Impotence commonly occurs as a result of treatment of carcinoma of the prostate. We review the etiology, evaluation, and treatment options available for these patients as well as our experience with fifty prostate cancer patients who underwent placement of penile prostheses. Several excellent alternatives are available for patients with impotence resulting from treatment of carcinoma of the prostate.


Maximal androgen ablation: a review.

Maximal androgen ablation: a review. Primary management of advanced (stage D) adenocarcinoma of the prostate is androgen ablation. Since this principle was discovered in the early 1940s, therapeutic alternatives and "progress" have centered around different ways to obtain castrate levels of androgens. The role of adrenal androgens in supporting prostate or prostatic cancer growth has been debated for decades and until recently was believed to be minimal. In the 1980s the concept of maximum androgen suppression, involving both the testes and adrenal glands, was reintroduced with some investigators claiming exceptional results. We review studies that have examined this concept, with emphasis on the largest trial which was carried out by the National Cancer Institute.


Megakaryoblastic termination of myeloproliferative disorders. Megakaryoblastic termination of myeloproliferative disorders is rare. The morphology of megakaryoblastic transformation can be subtle and is often mistaken for myeloid or lymphoid proliferations. Previously reported observations suggest a relatively poor prognosis for this category of patients, making precise diagnosis imperative. A multifaceted approach using morphology, ultrastructure, cytochemistry, and immunological membrane analysis may be helpful. We present two cases of myeloproliferative disorder with aggressive megakaryoblastic phases (myelofibrosis with agnogenic myeloid metaplasia and chronic myeloid leukemia with blast crisis). The clinical course is described and the results of the morphological, cytochemical, ultrastructural, and cytogenetic studies of both cases are presented. In addition, immunochemical studies (flow cytometry) and platelet function studies (aggregation, beta-thromboglobulin, and platelet factor IV release) were done for one of these patients.


Cerebral oxygen extraction during severe viral encephalitis. Viral encephalitis can cause devastating neurologic injury. Little is known about cerebral hemodynamics and metabolism in this condition. We report two patients with severe encephalitis, one proven and the other suspected to be due to herpes simplex, in whom the global cerebral oxygen extraction ratio (OER) and carbon dioxide (CO2) responsiveness was assessed. OER was low in both patients throughout the acute period. CO2 responsiveness was present initially in both and disappeared later in the more severely affected child. These cases demonstrate that cerebral hyperemia occurs in severe viral encephalitis and that hyperventilation can effectively reduce the intracranial pressure.


Federal, state, and local partnerships in providing primary care: one urban health department's endeavor with a state university medical center. We describe how the federal government, the City of Chicago, and the State of Illinois worked together to increase the availability and accessibility of health care services on Chicago's underserved west side by reopening a bankrupt, federally-funded community health center. The federal government made the building available to the City which then contracted with a state university medical center to be the provider of services. This partnering has allowed the Chicago Department of Health to offer services in a previously underserved area. The University has gained an opportunity for community-based primary care teaching, as well as community relations. Patients have increased access to a wide variety of specialty and inpatient care. If public health providers are to be successful in this financial climate, they must look to new partners and new ways of delivering services to increase availability of services at a time when they are greatly needed.


The emerging role of HTLV-I/II and HIV-1 among intravenous drug users in Detroit. During 1987-1988, a seroprevalence study of the human immunodeficiency virus (HIV-1) and the human T-cell lymphoma/leukemia virus (HTLV-I/II) was performed among Detroit intravenous drug users unaffiliated with substance abuse programs. Seroprevalence data along with patient demographic information were compared to a similar study performed in 1985-1986. In the earlier study, 12 (12.5%) of 96 individuals tested positive for HIV-1. Of the 74 available negative samples retested in 1987-1988 for retroviruses, 7 (9.5%) tested positive for HTLV-I/II. Thus, the overall retroviral (HIV-1, HTLV-I/II) seropositive rate for 1985-1986 was 22%. In 1987-1988, 11 (15.7%) of 70 individuals tested positive for HIV-1 and 7 (10%) tested positive for HTLV-I/II. Concomitant infection with both viruses was found in 2 (2.9%) of the 70 individuals. Thus, retrovirus seroprevalence in 1987-1988 was 22.9%. In 1987-1988, significant differences between the retroviral-positive group and the retroviral-negative group consisted of intravenous drug use greater than 16 years (P = 0.059) for an odds ratio of 3.80 (CI 1.12-12.89) and sex with female prostitutes (P = 0.029) for an odds ratio of 5.38 (CI 1.38-20.95).


Primary papillary serous carcinoma of the peritoneum: a case of complete remission of bulky peritoneal disease after chemotherapy. Peritoneal papillary serous carcinoma is a rare tumor that involves the surface of the pelvic and/or abdominal peritoneum. Long-term survival among patients with this tumor has been rare. Most patients with this cancer have been treated with debulking surgery and postoperative chemotherapy. A case of incompletely resected peritoneal papillary serous carcinoma with a complete response to cisplatin-based chemotherapy is reported. Subsequent laparotomy revealed no residual tumor. This case suggests that primary chemotherapy may be successful in treating unresectable primary papillary serous tumors of the peritoneum.


Interpretation of the pattern in rate ratios across strata. The pattern in the ratio of disease rates over strata is a summary statistic used to describe the changing risk of disease in one group relative to another. While patterns of the ratios of disease rates over strata appear to correspond to specific changes in disease rates, plots of the disease rates over strata seem to contradict the information yielded by the ratios. For example, if disease rates from populations A and B have identical rates of decline (parallel lines), the difference in the rates (A-B) at each strata remains constant, while the ratio of the rates (A/B) increases over strata. Through simple algebraic manipulation, one can show that the pattern of the rate ratio is a function of the rate difference relative to the endemic disease rate. Thus, rather than describing the behavior of the disease rates, ratio patterns reflect the importance of exposure relative to the disease rate in the unexposed population.


Presentation and evaluation of patients with epidermoid head and neck cancers. Epidermoid cancer of the head and neck occurs most frequently in individuals with predisposing factors for neoplastic development. Primarily these include exposure to smoking and alcohol, although other predisposing factors may be present. Site-specific symptoms along with a history of predisposing environmental exposure can direct physicians to suspect head and neck cancer. Endoscopy and ancillary testing are performed for tumor staging and subsequent treatment planning.


The why and wherefore of fructosamine.

The why and wherefore of fructosamine. GH and FA are useful monitors in the care of diabetic patients. For most situations, GH is the preferred test and should be routinely monitored. FA should be reserved for exceptional situations in which blood glucose control over one to two weeks must be assessed or in patients with a hemoglobinopathy. Patients with diabetes should be advised of their present GH level and the preferred goal.


Public health care delivery in five U.S. municipalities: lessons and implications. Increasing pressures on private and public hospitals have necessitated a reassessment of urban health care delivery. Patients left unserved by stressed private hospitals have placed a greater burden on public institutions, which themselves are often old, underfunded, and in danger of closure. As policy analysts consider remedies, primary care in community-based settings has reemerged as an important component of planning. We present results of a comparative analysis of five public health care delivery systems (Boston, Dallas, Denver, Milwaukee, and Seattle), reflecting their economic, political, and cultural dynamics. Although significant differences in the relative centralization of care and reliance on community-based clinics are evident, the five cities discussed have incorporated an increased emphasis on preventive and primary care. The diversity among the systems is highlighted; adaptability is apparently a vital component in designing a public health care system appropriate to the needs of particular communities. Implications for Chicago and other cities are discussed.


Training the urban health care provider: one department's first steps. As our country faces a national crisis in health care, few have outlined plans to improve the shortage of primary care physicians. This is especially critical in urban areas where sociocultural impacts on health are large. The Department of Family Practice and Community Medicine at the University of Texas School of Medicine in Houston has begun development of a division of Urban Family Medicine to address the special training needs of the urban family practitioner. Subdivisions that have been formed focus on undergraduate curriculum, graduate educational strategies, service, and research and policy to further develop the training model.


Community health centers: a resource for service and training. The American Medical Student Association (AMSA) Foundation is assisting the U.S. Public Health Service in increasing the number of primary care physicians trained and committed to practice in medically underserved areas. In collaboration with the American Academy of Family Physicians, the Ambulatory Pediatrics Association, and the Society of General Internal Medicine, AMSA conducted an assessment of federally-funded residency programs to identify and describe their affiliations with federally-funded community and migrant health centers (C/MHCs). Of the 260 programs assessed and the 147 responses, 125 offer community-based training. Of these, 73 offer training in primary care centers and 39 offer training in federally-funded C/MHCs. Residents training in the C/MHCs have positive experiences in both personal and professional development and are frequently hired by the health centers upon graduation. Benefits realized by the affiliations include a community orientation for the residents and enhancement of service and education missions for the collaborating institutions.


Health care consortia: a mechanism for increasing access for the medically indigent. In response to poor coordination among health and social service providers, health care consortia have emerged in many areas of the United States. Consortia link multiple providers in a common structure to create comprehensive systems of care. They can be formally structured or informal combinations of providers that engage in coordination but otherwise do not comprise an independent organization. The functions most common among all types of consortia are shared services and service coordination; however, a number of consortia also operate outreach/education programs. Consortia represent an innovative response to the need both for vertical integration--case management of all levels of care--and horizontal integration to prevent duplication among primary care providers. We outline the history of consortia in which federally-funded community health centers have participated. We also suggest an analytical framework for the various types of consortia; discuss lessons learned about building and maintaining consortia; and provide preliminary outcome data.


The structure and function of urban pharmacies: visits to community pharmacies in inner-city Chicago. Visits were made to 21 pharmacies in two poor neighborhoods on the west side of Chicago and interviews conducted with pharmacists-in-charge. The objective of the study was to provide a comprehensive description of the function, capabilities, and problems of urban pharmacy. We present results on the structure and function of these inner-city pharmacies. The pharmacies fit one of three structural forms: chain, independent, or medical center. The majority of respondents viewed the function of the inner-city pharmacy as patient-centered but also identified several barriers to effective patient communication. The results suggest that inner-city physicians and pharmacists should communicate with patients more often and in ways that patients understand. Also, Medicaid and other drug insurance programs should develop patient information networks and coverage packages intended to maximize patient health status.


Moving from health care research to action. Although the United States spends more on health care than any country in the world, access to that care is becoming increasingly difficult. The National Cancer Institute and other federal agencies are sponsoring innovative research for delivering effective medical services, particularly to underserved populations. Models of successful collaboration between private and public sectors concerned with health care can be adapted and implemented at the national, state, and local levels. However, other measures are needed to ensure access to adequate health care for all Americans. Minimal but effective regulations are needed to ensure quality control, reduce duplication of services, and minimize cost increases. Public and private sectors also need to consider ways to extend adequate health insurance coverage to all Americans and to provide compensation for preventive services.


The epidemiology of prostate cancer in black men. Data on the epidemiology of prostate cancer from the 1930s to the present document a dramatic racial difference in incidence, survival, and mortality rates in American men. American black men have the highest incidence and mortality rates of prostate cancer in the world. Survival data have been related to access to medical care, genetic and environmental factors, and cultural differences, including diet and social habits. Most reports present conflicting data with no clear positive correlations, and conclusions are often speculative. Better controlled, prospective studies of epidemiologic variables and a comprehensive genetic evaluation of black families with prostate cancer are needed to better understand the racial disparity affecting American black men and the biology of this disease in all men.


Health Care Linkage Project: improving access to care. The primary objective of the Health Care Linkage Project, funded by a grant from the Chicago Community Trust, is to develop, implement, and evaluate a primary health care linkage network within the city of Chicago that creates formalized linkages between community health centers, the Chicago Department of Health clinics, and hospitals. Six linkage networks are currently operational, with an additional two sites phased in during 1991. The success of the pilot project has been demonstrated by hundreds of patients receiving primary care and ancillary services on a more timely basis, by greater coordination between the public and private sector, by cost-savings to both patients and providers through reducing inappropriate use of services, and by a variety of spin-off projects which have improved the quality and accessibility of services. A second important objective is the development of a Health Care Linkage Manual that describes the practical experience and lessons gained from the linkages, the status of comparable arrangements in other U.S. cities, replicability of the linkage models, and recommendations for policy changes which will make linkages more effective.


An update on the role of ploidy in prostate carcinoma. One of the characteristic features of prostate carcinoma is its marked variation in biologic behavior. DNA quantitation has been studied in prostate carcinoma using a variety of techniques. Evaluation of tumor ploidy suggests that this may be the best predictor of the biologic behavior of prostate cancer in individual patients. This comprehensive review addresses the current studies, stage by stage, to clarify the clinical significance of these findings.


Usage of penicillinase as an enzyme marker in enzyme linked immunosorbent assay for the detection of IgG antibodies to Toxoplasma gondii. An Enzyme linked immunosorbent assay system has been developed using penicillinase as an enzyme marker for the detection of IgG antibodies to Toxoplasma gondii. This system is simple (end result assessed visually) reliable (the results are matching with other available commercial ELISA based kit), sensitive (detection of antitoxoplasma IgG antibodies to a level of 15 IU/ml) and reproducible (4.3 & 6.7% intra and inter coefficient of variation respectively). Experimental results with human serum samples using our system has shown consistent results and were in total agreement with other commercially available diagnostic kits. It is therefore suggested that ELISA system using penicillinase can be a simple, sensitive and reliable method for the diagnosis of toxoplasmosis in human.


Detection of toxoplasma IgM antibodies by ELISA method: a comparative study of different enzymes as markers. Detection of toxoplasma IgM antibodies by employing enzyme linked immunosorbent assay (ELISA) technique was developed using different enzymes viz, horseradish peroxidase (HRP) (EC. 1.11.17), urease (EC. 3.5.15) and penicillinase (EC 3.5.2.6) as markers. Of these enzymes, HRP is light sensitive and needs dark chamber, also inactivated by preservative sodium azide. Similarly urease test system is extremely pH sensitive and demands special care during ELISA technique. Whereas penicillinase showed certain distinct advantages viz. stable at room temperature, high specific activity and economical. In the present studies it was observed that the sensitivity of penicillinase is similar to HRP and urease, marker enzymes used in commercially available diagnostic kit. The prominent feature of detection of toxoplasma IgM antibodies involving these three enzymes are: a) Shorter incubation time (About 2.5 hours) b) No false positive reaction. Moreover, these enzyme conjugates were prepared from F (ab')2 fragments of antitoxoplasma rabbit serum to elicit specific interaction with IgM antibodies only, avoiding cross interaction with other non-specific proteins like compliment systems and rheumatoid factor.


Localization of laminin in nephritic glomeruli as revealed by a quick-freezing and deep-etching method with immunohistochemistry. The three-dimensional localization of laminin in rat glomeruli at the chronic phase of Masugi nephritis was investigated by a quick-freezing and deep-etching method combined with immunohistochemistry. Light-microscopically, laminin was localized in increased mesangial matrix and thickened glomerular basement membrane. The quick-freezing and deep-etching method revealed that the increased mesangial matrix, which was newly formed in axial portions and areas of mesangial interposition, was composed of fine fibrillar networks. They were revealed with the 3,3'-diaminobenzidine tetrahydrochloride (DAB) reaction products of peroxidase-labelled secondary antibody following anti-laminin antibody. However, these reaction products were not uniformly distributed in the newly formed matrix. Although the fibrils organizing lamina densa were also immunostained with anti-laminin antibody, the fibrils connected to mesangial cells, podocytes and endothelial cells had smaller amounts of DAB reaction products for laminin. These results indicate that one of the components of fibrils in the mesangial matrix and lamina densa is laminin, which is heterogeneously distributed in the newly formed matrix.


Histochemical study of glycoconjugates in the nasal mucosa of the rat and guinea pig. A histochemical study was carried out on the glycoconjugates of the nasal mucosa of rat and guinea pig using conventional techniques and peroxidase-labelled lectins. Both the respiratory mucosa and neuroepithelium were studied. Sulphate and sialic acid groups were found in the mucous layer of the neuroepithelia, Bowman's glands and goblet cells. In contrast, the nasal glands did not possess these groups, and only a few showed neutral mucins. Carbohydrate residues were more numerous in the acini of the Jacobson glands. Thus, the nasal glands in the rat and guinea pig are probably of a serous type because of the scarcity of carbohydrate residues.


Electron microscopic immunocytochemical localization of proline-rich proteins in normal mouse parotid salivary glands. Rabbit polyclonal antibodies against isoproterenol-induced mouse proline-rich proteins (PRPs) were used to localize PRPs in the parotid salivary glands of normal adult BALB/c mice. The antibodies recognized both acidic-type and basic-type PRPs. Immunoblotting experiments revealed that the glands contained an acidic-type and a basic-type PRP. Parotid gland tissue was fixed with Karnosky's fixative and embedded in Lowicryl resin at low temperature. PRPs were localized at the electron microscope level using an indirect post-embedding staining technique with protein A-gold. The secretion granules of the acinar cells were strongly labelled. Pre-absorption of the antibody with purified acidic-type and basic-type PRPs indicated that the basic-type PRP is mainly located at the periphery of the granules but that the acidic-type PRP is more evenly distributed within the granules. Pre-absorption of the antibody with alpha-amylase did not affect the staining pattern, suggesting minimal cross-reactivity. PRPs were also detected within the rough endoplasmic reticulum and the Golgi apparatus of acinar cells, within the granules of the proacinar cells and in the lumena of the ducts, but not within the intercalated or striated duct cell granules.


Dynamic nature and function of epidermal Langerhans cells in vivo and in vitro: a review, with emphasis on human Langerhans cells. Epidermal Langerhans cells (LC) are Birbeck granule-containing bone-marrow-derived cells, which are located mainly in the suprabasal layer of the epidermis. They can be readily identified by their strong expression of CD1a and MHC class II molecules. In addition to these 'classical' properties, an extensive phenotypic profile of normal human LC, summarized in this review, is now available. The powerful capacity of LC to activate T lymphocytes is clearly documented and, to date, LC are recognized as the prominent antigen-presenting cells of the skin immune system. They are generally believed to pick up antigens encountered in the epidermis and to migrate subsequently from the epidermis to the skin-draining lymph nodes. Upon arrival in the paracortex of lymph nodes, the antigen-laden LC transform into interdigitating cells and they present antigen to naive T lymphocytes in a MHC class II-restricted fashion; this results in the generation of antigen-specific immune responses. It has also been demonstrated that transformation of LC into interdigitating cells occurs when LC are cultured in vitro. Both in vivo and in vitro studies have indicated that properties of LC, such as phenotype, morphology and the stimulatory potential to activate T lymphocytes, are dependent on the local microenvironment in which the LC reside. The essential role of LC in the induction of contact allergic skin reactions and skin transplant rejection is well established.


Immunoelectron microscopic studies of glycosaminoglycans in the metaphyseal bone trabeculae of growing rats. The types and distribution of glycosaminoglycans (GAGs) were studied immunocytochemically in osteoid, mineralized bone matrix, and cartilage matrix of growing rat metaphyseal bone after aldehyde fixation and EDTA demineralization, using four monoclonal antibodies (mAbs 1-B-5, 2-B-6, 3-B-3 and 5-D-4). These mAbs specifically recognize epitopes in non-sulphated chondroitin (C0-S); chondroitin 4-sulphate (C4-S) and dermatan sulphate (DS); chondroitin 6-sulphate (C6-S) and C0-S; and keratan sulphate (KS) respectively. In osteoid, all mAbs except 1-B-5 weakly stained matrix material on and between collagen fibrils, and moderately stained organic material corresponding to bone nodules, which are known sites of mineralization. However, the staining of osteoid abruptly decreased at the mineralization front; weak staining was confined mostly to the organic material of bone nodules in mineralized bone matrix, with very weak or no staining of the rest of the bone matrix. This staining progressively decreased toward the mineralized cartilage matrix and became negative. The mineralized cartilage matrix and lamina limitans reacted strongly with all mAbs except 5-D-4. These results indicate that osteoid contains sulphated proteoglycans containing C4-S and/or DS, C6-S and KS, and subsequent bone matrix mineralization appears to require accumulation of these macromolecules within bone nodules and eventual loss of these substances for complete mineralization, whereas proteoglycans containing C0-S, C4-S and/or DS, and C6-S still exist in mineralized cartilage matrix and lamina limitans.


Prostaglandin E2 localization in the rat ileum. The application of anti-prostaglandin E2 immunoglobulin to plastic-embedded thin sections of the rat ileum has permitted the localization of prostaglandin E2 in this tissue. In agreement with the published data (Chock & Schmauder-Chock (1988), Schmauder-Chock & Chock (1989)), the results also suggest the presence of an arachidonic acid cascade in the granules of various secretory cells of the gut. Since antibody labelling was found within the secretory granules of connective tissue mast cells, goblet cells, and Paneth cells, the presence of the arachidonic acid cascade in these granules is implied. The appearance of prostaglandin E2 over the non-cellular internal elastic lamina of arterioles suggests that it may have been secreted along with the elastin. The even distribution of prostaglandin throughout the cytoplasm of the erythrocyte is consistent with the concept that this cell scavenges the eicosanoid from the circulation. These data further link the secretory granule to the production of eicosanoids and therefore illustrate the potential sources of prostaglandins in the rat ileum.


Calbindin D-28k-immunoreactivity in rat muscle spindles during postnatal maturation and after denervation. Calbindin D-28k-immunoreactivity has been demonstrated in some of the intrafusal muscle fibres and in the capsule of adult rat muscle spindles. In this study, the immunocytochemical localization of calbindin D-28k in the muscle spindles of triceps surae muscle was studied during postnatal maturation and after denervation. In young rats calbindin D-28k-immunoreactivity was seen in a few intrafusal fibres, first at the age of 4 days. At the 7th day, three calbindin D-28k-immunoreactive fibres and one unlabelled fibre were seen in most muscle spindles, as in adult rats. The spindle capsule and perineurial sheath of nerves were first seen to exhibit calbindin D-28k immunoreactivity at the age of 14 days, and thereafter the localization of calbinding D-28k-like immunoreactivity was similar to that in adult rats. After denervation, calbindin D-28k-immunoreactivity remained in intrafusal muscle fibres and the spindle capsule for a long period. After two months of denervation, calbindin D-28k immunoreactivity could still be seen in the spindle capsule, but the intrafusal fibres were not labelled. The innervation is known to have trophic effects on the intrafusal fibres. The present findings suggest that the expression of calbindin D-28k-immunoreactivity in maturating muscle spindles may be induced by the developing innervation. The decrease of calbindin D-28k-immunoreactivity in intrafusal fibres after denervation may be due to the loss of trophic factors released by the nerves.


A chromogranin peptide is co-stored with insulin in the human pancreatic islet B-cell granules. The immunoreaction of a rabbit chromogranin A and B antiserum was studied in normal human pancreatic islets. By examination of consecutive light microscopical sections, it was revealed that, at high antiserum concentrations (1:2000 or less), the whole islet area was heavily labelled, although the peripheral glucagon (A)-cells were the most intense in their immunoreaction. At low antiserum concentrations (1:4000 or more) the A-cells still showed the same intense labelling reaction, but the central B-cells were weakly labelled. Electron microscopically, reactivity towards the chromogranin A and B antiserum and the monoclonal insulin antibodies was present in the same central electron-dense core of the B-cell secretory granules, as demonstrated after application of the immunogold technique at different antibody dilutions. In the A-cells, the chromogranin immunoreactivity was concentrated at the peripheral mantle of the secretory granules. The D-cell granules showed a weak immunolabelling. Examination of human islets with the monoclonal chromogranin A antibody LK2H10 revealed immunogold labelling only in the peripheral mantle of the A-cell granules, while the B-cell granules were unlabelled. The present results show that a chromogranin peptide is co-stored with insulin the in normal human B-cell secretory granules. Although the exact composition of this B-cell chromogranin is unknown, it is not identical to that of the chromogranin A present in the A-cell granules.


Cellular heterogeneity in human transitional cell carcinoma: an analysis of optical properties and lectin binding. Flow cytometry was used to measure the binding of a panel of ten fluorescein isothiocyanate(FITC)-conjugated lectins to fifteen samples of normal and neoplastic human urothelium. Concurrent measurement of light scattering and fluorescence permitted the quantification of lectin binding to cellular subpopulations defined by their light-scattering properties. In normal urothelium, we previously demonstrated levels of lectin binding to the cellular subpopulations derived from the superficial and intermediate cell layers which were higher than levels which bound to the subpopulation derived from the basal cell layer (Ward et al., 1987). This difference was most marked with Maclura pomifera agglutinin (MPA), Ricinus communis agglutinin (RCA) and Ulex europeus agglutinin (UEA). We now report a similar correlation between the degree of differentiation of a cellular subpopulation and the level of lectin binding in human transitional cell carcinomas (TCCs). Morphological differentiation in human TCCs is accompanied by alterations in cell-surface carbohydrates which are similar to those which accompany cellular differentiation in the corresponding normal tissue. No systematic difference in lectin binding was observed between the corresponding subpopulations of normal and neoplastic urothelial cells.


Characterization of a human cervical carcinoma-associated antigen by lectin binding and immuno-electron microscopy. The specific binding and nature of the epitope recognized by monoclonal antibody (Mab) 1H10, which binds an antigen expressed on human cervical tumors, was characterized by enzyme digestion, lectin competition assay and immuno-electron microscopy. Membrane homogenates of CaSki cervical carcinoma cells were digested with various enzymes, then analysed by SDS-PAGE and immunoblotting. Cells grown on coverslips were treated with various enzymes and in situ binding of Mab 1H10 to cells was analysed by electron microscopy. The ability of lectin-conjugates to block Mab 1H10 binding to CaSki cells was also examined. Treatment of samples with sodium periodate abrogated antigen recognition by Mab 1H10. Neuraminidase and hyaluronidase digestion decreased but did not eliminate Mab 1H10 binding to cells in situ. Chondroitinase ABC digestion, in contrast, removed Mab 1H10 binding sites both in vitro and in situ. Trypsin and chymotrypsin digestion of cell membrane homogenates decreased the molecular weight of the Mab 1H10 antigen but did not decrease the binding intensity. Wheat germ agglutinin (WGA) strongly bound to CaSki cells and partially blocked Mab 1H10 binding, indicating that the antigen contains N-acetyl-galactosamine residues at or near the epitope recognized by Mab 1H10. Ricinus communis agglutinin (RCA) exhibited a similar binding pattern to WGA. However, concanavalin A bound only weakly to CaSki cells and was ineffective at blocking Mab 1H10 binding. The tumor-associated antigen recognized by Mab 1H10 is concluded to be a chondroitin sulphate glycoprotein or proteoglycan rather than a mucopolysaccharide or lipoprotein.


Immunohistochemical, autoradiographic and electron microscopic studies on the transformation of fibroblasts into chondrocytes in the mouse subfascia induced by bone morphogenetic protein. Functional morphology on the transformation of fibroblasts into chondrocytes induced by bone morphogenetic protein (BMP) was studied by light and electron microscopy using 35S autoradiography and immunohistochemistry for S-100 protein and type-II collagen. A pellet containing BMP obtained from a murine osteosarcoma was transplanted into the mouse subfascia. By 3 days after implantation, many typical fibroblasts, which were free of the silver grains for 35S and devoid of both S-100 protein and type-II collagen, entered the pellet region. By 5 days, the fibroblasts in the pellet region became polygonal in shape, and cytoplasmic vesicles and vacuoles appeared, both containing a homogeneous substance of low electron density. At 5 days, autoradiography revealed many silver grains for 35S over the Golgi apparatus and vesicles and vacuoles of the cells in the pellet region as well as over the surrounding extracellular matrix. Moreover, the cells at 5 days displayed immunoreactivity to both proteins. The extracellular matrix around the cell began to show clear metachromasia and increased in amount with time. At 9 days all the cells in the pellet region were round or oval in shape and surrounded by an abundant cartilaginous matrix. The rough endoplasmic reticulum and Golgi apparatus were extremely well developed, and a large number of vacuoles and vesicles were seen in the cytoplasm. These cells showed intense immunoreactivity to both proteins, and strong accumulation of sulfur was visualized in the extracellular matrix by autoradiography. These results suggest that the fibroblasts in the pellet region change into chondroblasts by 5 days, and become typical chondrocytes by 9 days.


Lectin-binding carbohydrates in sexual differentiation of rat male and female gonads. Development and sexual differentiation of the mammalian gonad involve changes in the type and distribution of different proteins and glycoproteins in and around the epithelial gonadal cords, the future seminiferous tubules in the testis, and follicles in the ovary. To study changes in cellular carbohydrate-containing compounds in the sex-specific morphogenesis of rat gonads, sections from embryonic, fetal and early postnatal gonads were labelled with seven different fluorescein isothiocyanate (FITC)-conjugated plant lectins of various carbohydrate-binding specificities. Double labelling of laminin with tetramethylrhodamine isothiocyanate (TRITC)-conjugated antibodies was used to outline the epithelial tissues. From the results we conclude that the abundance and similar distribution of carbohydrates in the early gonads of both sexes supports their sexually indifferent nature. Furthermore, the basement membranes of the differentiating gonadal cords in both sexes have common features, which differ, however, from those of the other developing urogenital organs. Changes in carbohydrate composition appear with the sexual differentiation of the gonads; the similarity of the changes in lectin binding to the gonadal cords of embryonic and fetal male, and to postnatal female, suggests similar mechanisms of cell-cell interactions in both sexes although activated at different developmental stages. These carbohydrate specificities at the tissue level should be taken into account together with cell-type specific changes, e.g. in the formation of the zona pellucida, when the phenomenon of polymorphic expression of different compounds is integrated into theories of epithelial differentiation.


Neural regulation of differentiation of rat skeletal muscle cell types. Three monoclonal antibodies (LM5, F2 and F39) to the fast class of myosin heavy chain (MHC) were used to study the effect of denervation on the differentiation of muscle cell types in some rat skeletal muscles. Antibody LM5 in immunocytochemical investigations did not stain any myotubes during early fetal development but presumptive fast muscle cells started to stain during later fetal development. Unlike antibody LM5, antibodies F2 and F39 stained all myotubes during fetal development. The suppression of fast myosin heavy chains recognised in presumptive slow muscle cells was observed within 1-2 days after birth with antibody F39 but not until 10-14 days after birth with antibody F2. The emergence of subsets of fast muscle fibre types in rat extensor digitorum longus (EDL) and tibialis anteri (TA) detectable by F39 and F2 antibodies was not observed until 2-3 weeks after birth. Denervation of developing muscles led to marked changes in the expression of myosins identified by these antibodies.


Electron microscopic characterization of calcium-binding physodes in the green alga Mougeotia scalaris. Effect of the covalently cross-linking agents glutardialdehyde and osmium tetroxide, and of adsorption of the vital dye, neutral red, to the matrix of the calcium-binding "vesicles" from the green alga Mougeotia scalaris has been analysed in situ, both in terms of structural preservation and of the calcium-binding capacity of the vesicles. Upon cell fixation in glutardialdehyde without OsO4, the vesicles appear to dissolve, but upon simultaneous fixation in glutardialdehyde with OsO4 (1% w/v), the vesicles retain a globular form, are evenly stained by osmium and appear to be surrounded by a membrane-like structure. This structure was also observed around the vesicles in cells preincubated for 10 min in 0.1 mM neutral red and then fixed in glutardialdehyde/OsO4 for 1 h. More detailed information of the matrix structure is obtained when simultaneous fixation of the Mougeotia cells was shortened to 15 min: a membrane-like structure was no longer observed around the vesicles. After cell treatment in the presence of neutral red, no calcium at all was found inside the vesicles. A small amount of calcium remained, when cells were fixed simultaneously and extensively in the absence of neutral red. However, calcium was found, to a considerable extent, inside the vesicles after short simultaneous fixation of the cells in the absence of neutral red. Based on the ultrastructural and elemental features presented here, the calcium-binding vesicles in Mougeotia appear to represent a member of the large family of (calcium-binding) physodes in lower plants (CaBP).


Induction of G- and R-banding in human chromosomes by the demethylating agent S-adenosyl-L-homocysteine. In this report we describe the procedure of growing human lymphocytes with the demethylating agent S-adenosyl-L-homocysteine (SAH). After this treatment, which is not toxic for cell survival, both R- and G-banding were obtained by new experimental procedures: R-bands have been directly demonstrated with the GC-specific fluorochrome chromomycin A3 without the necessity of any AT-specific counterstaining agent; simultaneous G-banding and active nucleolar organizer regions have been obtained by silver impregnation of chromosomes and subsequent Giemsa staining. These results suggest a possible relationship between local differences in DNA methylation and the determination of the banded chromosome structure.


Molecular characterization of "inverted" pericentromeric heterochromatin of chromosome 3. Inversion of the pericentromeric region of human chromosome 3 [inv (3) (p11q11.2)] is a rare event. Initially, this inversion was identified with staining for Q-bands by fluorescence using quinacrine (QFQ) and later characterized with staining for C-bands by CBG technique. The molecular methods of fluorescence in situ hybridization (FISH) and AluI/Giemsa and TaqI/Giemsa techniques were utilized. The findings suggest that the variable band q11.2 on chromosome 3 contains alphoid DNA sequences, which appear to be similar to those identified by conventional methods in the centromeric region (band p11).


Ecdysteroid binding sites in gastrolith forming tissue and stomach during the molting cycle of crayfish Procambarus clarkii. The distribution of ecdysteroid binding sites in the stomach and gastrolith disc tissue of crayfish (Procambarus clarkii) was examined in relation to the molting stage by thaw-mount autoradiography. The radiolabeled hormone analogue ponasterone A (25-deoxy-20-hydroxyecdysone) was used. Ecdysteroid binding sites were demonstrated only in certain molting stages, the small gastrolith period and the aftermolt stage. In gastrolith epithelium, ponasterone A binding sites first appeared in the cytoplasm, and then in the nuclei and cytoplasm. In the stomach epithelium, many nuclear binding sites were detectable during the period of gastrolith secretion. These periodical changes in specific ponasterone A binding when correlated with the molting stages clearly show that ecdysteroids may function as an initiator for gastrolith formation and reabsorption. The findings also suggest that ecdysteroids control calcium transport in the stomach epithelium. The time-related and functional differences of cytoplasmic and nuclear concentration of ecdysteroid receptors indicate the presence of cytoplasmic and nuclear receptors associated with specific actions.


Direct detection of immunogold reactions by real-time video microscopy. Video-enhanced microscopy allows the detection and tracking of individual colloidal gold particles. The analysis of immunogold reactions can also be conducted as a function of time and thus allows the study of dynamic events in living cells. The direct visualization in real time is reported of the reaction of immunogold particles with a surface antigen. This time-resolved immunocytochemistry was achieved by continuous observation of living cells infected with a virus (respiratory syncytial virus) following their incubation with colloidal gold (30 nm) coated with antiviral antibodies. The progress of the immunoreaction was visualized as a sequential deposition of individual gold granules on the viral particles until saturation was reached after 60 min. Binding of colloidal gold was an irreversible event as no elution or dislocation of surface-bound granules took place. Comparative imaging of colloidal gold particles by electron microscopy and by video microscopy demonstrated that the video-imaged immunoreactions represented events involving single gold particles; their signal was sometimes clearly enhanced by secondary depositions taking place in close proximity, i.e. at a distance below the lateral resolution of the light microscope. Our experiments demonstrate that video-enhanced microscopy provides a powerful tool for studying antibody-antigen reactions with a high spatial and temporal resolution.


Nuclear matrix-bound replicational sites detected in situ by 5-bromodeoxyuridine. The nuclear matrix was prepared in situ from Swiss 3T3 cells, which were synchronized by contact inhibition and serum starvation and pulse-labelled for very short periods of time with 5-bromodeoxyuridine (5-BrdU). For the first time 5-BrdU has been employed to demonstrate the association of newly synthesized DNA with a nucleoskeleton. Immunofluorescence analysis using a monoclonal antibody to 5-BrdU revealed five different intranuclear staining patterns at different stages of the S phase. These patterns were observed also in intact cells and did not change during the matrix preparation steps which involve extraction with 2 M NaCl and DNase I digestion. Such an observation was also confirmed by spatial confocal microscopy studies. The intensity of fluorescence, which was evaluated by cytofluorometry, increased to reach a maximum during mid-S phase and then decreased. Because no significant difference was found in the time to label residual DNA of different 5-BrdU staining patterns, this strongly suggests that a different number of replicons is activated at different stages of the S phase. These results strengthen the hypothesis that eukaryotic DNA replication occurs in close association with an insoluble protein nuclear skeleton, which determines the three-dimensional spatial organization of chromosome duplication.


FMRFamide-immunoreactive structures in the brain of the brown hagfish, Paramyxine atami: relationship with neuropeptide Y-immunoreactive structures. Localization of the molluscan cardioexcitatory tetrapeptide FMRFamide (Phe-Met-Arg-Phe-NH2) in the brain and hypophysis of the brown hagfish, Paramyxine atami, was examined by immunohistochemistry specially regarding a possible relationship with neuropeptide Y (NPY). FMRFamide-immunoreactive fibers were demonstrated in many regions of the brain, with the highest density in the diencephalon. However, no immunoreactivity was found in the hypophysis. Labeled cells were chiefly located in the nucleus hypothalamicus of the diencephalon, although a few cells were recognized in the ventrolateral area of the caudal tegmentum. Examination of adjacent sections immunostained alternatively with anti-NPY antiserum and anti-FMRFamide antiserum showed overlapping of the distributional patterns of the immunoreactive structures in the brain. Moreover, the same cells in the nucleus hypothalamicus were immunostained with both antisera. Cross-blocking experiments showed that the FMRF-amide-immunoreactivity is abolished by preabsorption of the antiserum with homologous antigen, but not eliminated completely by pretreatment with appropriate antigens (NPY, avian pancreatic polypeptide and methionine-enkephalin-Arg-Phe). In contrast, the NPY-immunoreactivity was blocked by pretreatment of the antiserum with NPY, pancreatic polypeptide or FMRFamide, although no blocking by enkephalin was observed. Accordingly, the present study shows that, in the brown hagfish, FMRFamide-immunoreactive structures in the brain can be recognized by anti-NPY antiserum.


Sensitive mRNA detection using unfixed tissue: combined radioactive and non-radioactive in situ hybridization histochemistry. In the present study some experimental parameters for in situ hybridization histochemistry (ISHH) have been analysed using 35S-labelled and alkaline phosphatase-conjugated probes, in order to develop a reproducible double-labelling procedure. We have compared the total exclusion of tissue fixation with tissue sections fixed by immersion in formalin. In addition, the effect of dithiothreitol was assessed both when combining radiolabelled and non-radioactive probes on a single tissue section and when the probes were used separately. Hybridization of unfixed tissue resulted in stronger specific labelling and lower background both for radiolabelled and alkaline phosphatase-conjugated probes. No loss in tissue preservation was seen at the light microscopic level after hybridization of unfixed tissue. High concentrations (200 mM) of dithiothreitol strongly suppressed background when using 35S-labelled probes, whereas in the non-radioactive procedure, alkaline phosphatase labelling could only be achieved with very low dithiothreitol concentrations (less than 1 mM). This incompatibility led to a protocol using unfixed tissue sections and a sequential hybridization procedure, with the radiolabelled probe and high concentrations of dithiothreitol in the first step and the alkaline phosphatase-conjugated probe without dithiothreitol in the second step.


Initial reaction kinetics of succinate dehydrogenase in mouse liver studied with a real-time image analyser system. The initial reaction kinetics of succinate dehydrogenase in situ were investigated in sections of mouse unfixed liver using an ARGUS-100 image analyser system. The sections were incubated on substrate-containing agarose gel films. Images of a section, illuminated with monochromatic light (584 nm), were captured with the image analyser in real time at intervals of 10 s during the incubation. The absorbances of selected hepatocytes in the successive images were determined as a function of time. In every cell, the absorbance increased nonlinearly after the first minute of incubation. The initial velocity of the dehydrogenase was calculated from the linear activities during the first 20 s of incubation. Hanes plots of the initial velocities and succinate concentration yielded the following mean kinetic constants. For periportal hepatocytes, the apparent Km = 1.2 +/- 0.8 mM and Vmax = 29 +/- 2 mumol hydrogen equivalents formed/cm3 hepatocyte cytoplasm per min. For pericentral hepatocytes, Km = 1.4 +/- 1.0 mM and Vmax = 21 +/- 2 mumol hydrogen equivalents/cm3 per min. The Km values are very similar to those determined previously from biochemical assays. These results, and the observed dependence of the initial velocity on the enzyme concentration, suggest that the technique reported here is valid for the histochemical assay of succinate dehydrogenase.


Distribution of hyaluronan and its CD44 receptor in the epithelia of human skin appendages. Biotinylated hyaluronan (HA) binding complex (HABC) from bovine articular cartilage proteoglycan was used as a histological probe to study the localization of HA in human skin. The distribution of HA was compared with its presumptive cell surface receptor, CD44, using monoclonal antibodies. In epidermis both HA and CD44 were found in the basal and spinous cell layers, but neither was present in the stratum granulosum and stratum corneum. In the keratinizing parts of hair follicles, i.e. in the outer and inner epidermal root sheath, pilosebaceous duct and the actual hair, HA and CD44 were found between the vital but not the terminally differentiated cells. In the sebaceous glands a small amount of HA was found around all cells, whereas CD44 was restricted to the basal cell layer. The secretory acini of the sweat glands stained intensively with anti-CD44 antibodies but only weakly with HABC. In the sweat gland, CD44 was localized on the basal and lateral surfaces of the clear cells, whereas the dark cells and the myoepithelial cells were negative. Both the lower and upper layers of the sweat gland ducts showed a faint but constant staining for CD44 and only minor amounts of HA. While in the keratinizing skin epithelia both HA and its CD44 receptor showed an intense staining with a close co-distribution, in the sweat and sebaceous glands their distribution patterns were not similar. It is suggested that in epithelia with divergent differentiation programs the functions of CD44 and HA may be different.


Immunohistochemical localization of pancreatic spasmolytic polypeptide (PSP) in the pig. Pancreatic spasmolytic polypeptide (PSP) is a peptide that is isolated from the porcine pancreas and that affects intestinal motility and growth of intestinal tumour cells in vitro. The peptide was recently demonstrated to be present in large amounts in pancreatic juice. The cellular origin of the peptide, however, is largely unclarified and the localization was therefore studied of PSP in pigs using immunohistochemistry. Positive immunoreactions were seen in the pancreas, the stomach, the duodenum, the jejunum and the ileum. In the pancreas, the PSP immunoreaction was seen in all acinar cells; no immunoreaction was seen in the endocrine islets. In the stomach, it was localized to the mucous cells of the glands in the cardiac gland region, the corpus and the pylorus. In the duodenum a strong immunoreaction was present in Brunner's glands and in the cells of their excretory ducts. In the jejunum and ileum, PSP immunoreactivity was seen in some of the cells in the epithelium of the crypts of Lieberkühn. A peptide chromatographically identical to highly purified PSP was identified in pancreas and stomach extracts. Thus epithelial cells in all parts of the stomach and small intestine contribute to the supply of PSP to the gut lumen.


5'-nucleotidase in spinal meningeal compartments in the rat. An immuno and enzyme histochemical study. The distribution of 5'-nucleotidase (5'-Nu) is reported in spinal meninges of the rat on the basis of an immunohistochemical and enzyme histochemical investigation. Strong immunoreactivity was found in the arachnoid membrane and in the sheaths of the spinal roots as well as in septa subdividing the roots. Also the superficial layer of the ligamentum denticulatum showed enzyme staining. No immunoreactivity could be detected in the pia mater or along the spinal nerve roots outside the subarachnoid space. Within the arachnoid mater the immunoreactivity was concentrated in the basal zone of the arachnoid membrane, thus appearing as a narrow fluorescent band near the border of the dura. An accentuation of immunoreactivity could be observed in areas where small dural blood vessels approach the subarachnoid space. It is well known that adenine nucleotides released from neural and glial cells of the central nervous system finally reach the cerebrospinal fluid. We presume that 5'-Nu in the arachnoid membrane and spinal root sheaths is responsible for the conversion of adenine nucleotides into adenosine and that this conversion is associated with the reabsorption process of cerebrospinal fluid which most probably also takes place in spinal meninges. Adenosine, the product of 5'-nucleotidase, could play a role in the reabsorption process by its vasodilatatory effect on dural and epidural vessels.


Amalgamation of two endocytic probe techniques: fluoresceinated cationized ferritin can show up, sequentially, selected structures, first in living cells and then by electron microscopy. Fluorescein isothiocyanate-cationized ferritin (FITC-CF) has hitherto been used mainly to identify structures in living cells by light microscopy, by virtue of its fluorescent properties. We show here that this conjugate can be used, after immediate fixation of the same cell sample and preparation of thin sections, to recognise the same structures, by virtue of the ferritin's electron opacity. The conjugate should thus have a new use as a single-application, dual-purpose probe, e.g. in endocytic studies. The procedure may have advantages over similar dual-purpose probes in not requiring staining or special treatment.


Fibronectin and laminin in transverse tubules of cardiac myocytes studied by laser confocal microscopy and immunocytochemistry. The distribution of two non-collagenous glycoproteins of high molecular weight, fibronectin (FN) and laminin (LMN), was investigated in myocardial cells from the ventricle of rats, and from biopsies collected from the auricle of patients undergoing a coronary bypass operation. In order to elucidate the expression of FN and LMN across cells, non-invasive serial sectioning has been carried out by laser scanning confocal microscopy of frozen, immunostained tissue sections. In addition, immunoelectron microscopy was used to study the distribution of these antigens at higher magnifications. These studies show that FN is part of the basement membrane of the surface sarcolemma of both ventricular and atrial cells, in addition to being an abundant protein of the extracellular matrix (ECM). Along transverse tubular(TT)-membranes, FN was only detected in tubules exceeding 200 nm in diameter. Even here, the intensity of labelling varied greatly and was generally low. By contrast, a heavy investment of LMN was organized in the basal lamina along the surface sarcolemma and along ramifications of the entire TT-system in ventricular heart muscle cells. In this way, the network of TT-membrane systems of working heart muscle cells provides a supply of LMN to all depths of the myocardial fibre. In human atrial muscle cells, a regular TT-system appears to be absent. Instead occasional, deep sarcolemmal invaginations occur with diameters of 300-500 nm, the surfaces of which are also invested with LMN. The significance of the present findings has been discussed, with special reference to LMN as a possible component of a series of proteins involved in transmembrane communication between the ECM and the sarcoplasm.


Which fluorophore is brightest? A comparison of the staining obtained using fluorescein, tetramethylrhodamine, lissamine rhodamine, Texas red, and cyanine 3.18. There are several red-emitting fluorophores available for immunofluorescence studies, including tetramethylrhodamine, lissamine rhodamine, Texas Red, and cyanine 3.18; however, it is unclear which of these is best. The present study compared the brightness of these fluorophores to that of fluorescein. Staining was attempted using a primary antibody raised against serotonin and a secondary antibody that was conjugated with either fluorescein or one of the red fluorophores. The intensity of staining was determined densitometrically. It was found that a conjugate of cyanine 3.18 provided significantly brighter staining that conjugates of any of the other fluorophores, including fluorescein. It is concluded that cyanine 3.18 should be useful for multicolor fluorescence experiments and that it may be the brightest fluorophore available for single-color fluorescence immunocytochemistry.