United States Patent ( 10 ) Patent No . : US 7479 , 375 B 2
(54) METHOD FOR CLASSIFYINGA 5,372,946 A 12/1994 Cusak et al. MCROORGANISM IN A BIOLOGICAL 5,389,547 A 2f1995 Tanaka et al. SAMPLE 5,401,647 A 3, 1995 Tanaka et al. 5,474,984 A 12/1995 Tanaka et al. (75) Inventors: Norman R. Wainwright, Falmouth, MA 5,504,011 A 4, 1996 Gavin et al. 5,518,006 A 5/1996 Mawhirt et al. (US); Dana M. Nutter, Charleston, SC 5.534.226 A 7/1996 Gavin etal (US) 5,550,030 A 8/1996 Tanaka et al. 5,574,023 A 11/1996 Shibata et al. (73) Assignee: Charles River Laboratories, Inc., 5,591.403 A 1/1997 Gavin et al. Wilmington, MA (US) 5,591,628 A 1/1997 Baek et al. 5,605,806 A 2/1997 Tanaka et al. (*) Notice: Subject to any disclaimer, the term of this 5,637,474 A 6/1997 Takaoka et al. patent is extended or adjusted under 35 D380,555 S 7/1997 Kurosaki et al. U.S.C. 154(b) by 264 days. 5,681,710 A 10/1997 Tanaka et al. 5,695,948 A 12/1997 Tanaka et al. (21) Appl. No.: 11/331,600 5,702,882 A 12/1997 Tamura et al. D390,661 S 2/1998 Foggia (22) Filed: Jan. 13, 2006 (Continued)
A new kinetic single-stage Limulus amoebocyte lysate method for the detection of endotoxin in water and plasma.
A convenient kinetic chromogenic Limulus amoebocyte lysate (LAL) method, based on components originally intended for a two-stage end-point method, has been developed. The components (LAL and chromogenic substrate) can be pooled and subsequently used in a single-stage kinetic procedure adapted to microplates. With the help of a kinetic software, it is possible to measure over the three log range 0.005-12 EU/ml in one test run--a range considerably wider than the range of the end-point procedure. A good linearity of the log-log standard curve, reflected by coefficients of regression between 0.997 and 1.0, is shown as well as a high-resolution (> 700 s) between the negative control and the lowest standard point. Moreover a good precision (C.V. < 10%) is obtained in both water and plasma, showing the usefulness of the method in different applications. Finally, a strong correlation (r = 0.95-0.99) to other LAL methods is demonstrated.
Evaluation of antioxidants by a rapid polarographic method
A simple, rapid, precise, and reproducible method and an adaptable apparatus for polarographic measurement of lipid peroxidation and antioxidant activity is described. This method measures antioxidant activity within a few min time as compared to the hours or days needed for several widely used methods. Peroxidation rates were measured for different lipids as a function of amount of lipid, and amount of biocatalyst. The method is applicable to the detection and measurement of the antioxidant effects exhibited by weak antioxidants and has been satisfactorily used for the measurement of the antioxidant activity of different concentrations of powerful antioxidants.