[ 0032 ] fig3 is a block diagram of a system according to the invention for acquiring a virtual microscope slide , that includes a z - axis image dimension across the entire virtual slide . the system includes a microscope subsystem 15 with a digitally controlled stage platform 28 for supporting the glass microscope slide 21 . the digital stage platform 28 can operate over a large number of increments to position the stage in the horizontal x and y plane with high precision . a glass microscope slide or other substrate 21 is placed on the stage 28 . the system also includes a controlling computer system 32 with a keyboard 37 , a mouse 38 with a mouse wheel control 39 , and a display monitor 40 . the controlling computer system keyboard and mouse are used via the automatic focus step size setup window 12 to input the automatic focus step size parameters and the z stack step size setup window 13 is used to input the z stack step size parameters . [ 0033 ] fig3 a shows the focus setup step size 55 input control . also in fig3 a are shown the associated setup controls for the frequency of focus 56 , a threshold for controlling whether automatic focus should be performed on a specific field of view , and a control 58 to manually increment the z - axis dimension to move the microscope stage 28 up or down vertically in incremental units . as may be appreciated by the foregoing description , and the following descriptions , and as is well known in the art , the control of focus at high magnification and small depth of field is complicated and involves many variables . it is also time consuming if performed on every specimen image field of view in constructing or capturing a virtual microscope slide image data set . therefore , in the preferred embodiment and in the subsequently described alternative embodiments there are additional control and setup parameters to overcome major variables and to enable a faster overall virtual slide scan capture time . some of these are seen in fig3 a . for example , the frequency of focus 56 parameter allows automatic focus to occur on every adjacent field of view if it is set to 1 , or on every other field if it is set to 2 , etc . in the following detailed description the assumption is that the frequency of focus is set to 1 . however , if it is set to a higher number the reference tiles of field of views not focused take the default focus contrast value of the last previous focused image tile . in the alternate embodiments of the invention , the reference tile position is sometimes obtained in a different fashion as described for those embodiments . there is a significant speed of scan advantage related to not focusing on every field of view . however , on many specimens the disadvantage is that the resulting scan may not have an optimum depth of field position for the reference image . it may also be appreciated that one of the z stack images may then offer a more optimum image for the final remote viewer of the images . also , often there is not enough image structure in a field of view to obtain an automatic focus , for example in the case of a substantially blank or empty field of view . in that case the control 57 allows for a focus contrast threshold value input parameter that can be checked to allow skipping such fields . also , in that instance the default reference z - axis position for the next image requiring focus is the last previous focused image tile . [ 0034 ] fig3 b shows the z stack step size 60 input control . it may be appreciated that there are a multitude of factors that would require this parameter to be changed for a specific specimen . however , the most important of these is usually the magnification of the specific microscope objective lens being used , since each lens has a different depth of field specification , in combination with the type of specimen and estimated thickness of the specimen preparation . also , shown in fig3 b is a checkbox control 59 to either enable or disable the z stack image save for a specific virtual microscope data capture scan . according to the teachings of the aforesaid patent the computer controlled microscope is moved to start a scan of the entire specimen object 31 using the stage controller 14 to move the precision stage 28 to a new objective lens 10 field of view to acquire an initial image at that position and compute a focus contrast parameter on that image . according to the present invention the relative z - axis reference position for the first new field of view image tile is obtained by an iterative automatic focus procedure . the controlling computer system 32 sends the microscope subsystem 15 z - axis control signals to change the z - axis position control to move the stage incrementally to go up 4 times in the automatic focus step size and then to go down 4 times in automatic focus step size . at each incremental change in the z - axis position the image acquisition electronics 17 are controlled to acquire an image . a focus contrast parameter is computed on each image . the automatic focus z - axis position is then determined by choosing the z - axis position associated with the largest value of the focus parameter from the initial reference image and the set of 8 images . if the largest value is at one end of the sequence , i . e the 4 th image down or the 4t image up from the reference image , that image becomes the reference image , and the procedure is recursively repeated until the largest focus contrast value is found in the middle range of the sequence of images , i . e . not at either end image . this becomes the relative z - axis reference position for the new field of view image . as explained more fully in the following , the image tile associated with this relative z - axis position is then stored in the virtual slide data structure . in the preferred embodiment of the invention the controlling computer system is operated under a windows operating system ( microsoft corporation , redmond , wash ., usa ). referring to fig3 a , the virtual microscope slide data structure is stored as a windows operating system file folder where each tiled image is a . jpg image file with an incremental image name automatically assigned by the controlling computer systems software program . the . jpg image names are numbered so that the first acquired image is called da0 . jpg , the second da1 . jpg , the third da3 , etc . in fig3 c there is depicted a virtual slide data folder 42 with portions of the data structure 43 , 44 , 45 , 46 , and 47 also depicted . the set of 9 image tiles 43 , 45 and 46 named da98 are associated with a specific x , y specimen image plane position , and an adjoining set 44 and 48 , named da99 are associated with another abutted specific x , y specimen image plane position . the two reference tiles are depicted as 43 and 44 for the data structure at the two x , y locations . these tiles are in the automatic focus determined z - axis position , and the recorded . jpg image contains the depth of field image structure associated with that z - axis position and the field of view at the respective x , y location . during the system program operation to produce a virtual microscope slide , the controlling computer system also creates an additional text information file of the windows operating system format . ini . as depicted in fig3 c , this file 47 is named finalscan . ini . among other things this file contains a list of names corresponding to each reference tile in the virtual microscope data structure . for each reference tile in the list , that tiles x , y , and z digital location is tabulated . as taught in the aforementioned patent this information is then used by the virtual internet server and virtual microscope visual display programs to abut and reconstruct the various tiled images to allow a remote viewer to view contiguous regions of virtual slide images . it may be appreciated that the components of the data structure shown by example in fig3 c may be stored in a database or any other form allowing rapid sequential access to the reference image and the full z stack components . a novel aspect of this data structure is the close association of these image components . this greatly facilitates client server interactions in remote internet viewing . since the this subsequent viewing is through the limited x , y planar view of an image display device , only a few reference tiles ( and in certain limited situations only one reference tile ) may be in the available user view for a focus request to the server . as will be appreciated in the following description , this type of data structure facilitates rapid transmission of z - axis image content to the client computer . some internet server computers facilitate serving very large images requiring zooming , by using a pyramid data structure where different levels of image zoom are pre - constructed from the original image and kept in memory or virtual memory at one time . this requires a very large amount of memory when considering the requirement of keeping multiple planes of such image structures , such as shown conceptually in fig9 . the data structure of this invention is much more efficient when used specifically for virtual microscope slides with special viewing programs , since it in essence is pre - constructed to serve reference and z stack images rapidly from memory or digital disk storage in these small reference and z stack image units . after capturing a relative tile for the z - axis position at a given x , y specimen plane position the system of the invention proceeds to use the z - axis step size and execute the z stack save procedure . to accomplish this , the controlling computer system 32 is directed to control the z - axis positioning control 16 of the microscope subsystem 15 first to move down the z - axis in incremental z - axis step sizes , and at each step to acquire an image tile . these image tiles 45 are stored in the data structure depicted in fig3 by example for the data - structure set da98 . secondly , the controlling computer system 32 is directed to control the z - axis positioning control 16 of the microscope subsystem 15 to move up the z - axis in incremental z - axis step sizes from the reference z - axis position , and at each step to acquire an image tile . these image tiles 46 are stored in the data structure depicted in fig3 by example for the data structure set da98 . the same series of events described above for the data structure capture of the tile set da98 is repeated for all field of views associated with the capture of the virtual microscope slide . for example , in fig3 as the data structure set da99 , and so forth . the result of the above described preferred embodiment of the system of the invention is in effect to first factor out , or neutralize , the z - axis irregularities in optimum focus position over the x , y surface of the slide for the initial relative captured image tile , and then , secondly to create a set of cohesive z - axis dimensioned captured image planes , where each plane relates to a different , real , physical depth of field position in the specimen . the first relative z - axis positioning has brought into parallel positioning capture , the optimum depth of field portions of each specimen , and the z stack capture has resulted in image planes above and below that . this image sequence sampling can be reconstructed from the data structure storage elements 43 thru 43 , 44 , 44 , 45 , 46 , and 48 when used with the x , y location information stored in data structure element 47 . this reconstruction is depicted in an idealized fashion as shown in fig4 in cross section and in fig9 in perspective schematic , as an aid in visualizing the resulting complete virtual microscope slide data structure . as described below , in fact only a small portion of this is seen by the remote viewer at one time , because of the limitations of the image display of commonly used computer screens . however , it is all available for viewing by scrolling and requesting additional image tiles from a virtual microscope slide server . it will be appreciated by those familiar with the art that the above preferred description of the embodiment of the invention may be modified in other ways to enable the creation of a virtual microscope slide with z - axis image dimension information . in this regard , an alternative method of practicing the invention is described . this method is more applicable for specimen objects that don &# 39 ; t cover a large area , or in those instances where the stage platform 28 and microscope slide 21 are positioned to present the specimen 31 in a reasonably flat plane , or where a lower power objective is used that has a larger depth of field . for a given level of magnification ( such as 10 × for example ) the microscope objective 10 with associated video camera is adjusted up or down , or as in the preferred embodiment , the stage is adjusted up or down , either adjustment to bring into view an initial reference image into the focal plane depth of field of the microscope objective 10 and used to create magnified images of the specimen 31 for a given x , y position in the specimen plane . a first series of planar abutted image tiles are obtained as described in the preferred embodiment as the reference tile set , and stored in the data structure previously described , and as shown by example in fig3 c , wherein the example reference tiles 43 and 44 are depicted . the reference tiles z - axis position in this embodiment are computed using the results of a prior setup procedure where the z - axis positions at three separate places on a specimen are determined and a mathematical z - axis plane is determined across the x , y plane of the specimen by computations involving fitting a plane in the z - axis by using three x , y points with known z - axis values . in this case during the scanning process this computed position is used instead of the iterative , recursive , automated focus procedure described in the first preferred method . this results in a faster scan and image capture process . by way of illustration the capture of the complete set of tiles in this plane may be visualized in cross section as the depth of field 41 in fig4 . this scan captures the upper surface of much of the specimen 31 . then , in accordance with this embodiment , the stage z - axis position is changed according to one z stack increment and another series of images are captured and stored in the data structure shown in fig3 . for example , if the first series of images used the focal distance corresponding depth of field represented by reference numeral 41 , then by decreasing the stage z - axis position relative to the microscope objective , the next series of images would be represented by reference numeral 42 . conversely , by increasing the stage z - axis position relative to the microscope objective the next series of images would be represented by reference numeral 44 . subsequent series of images can likewise be captured by positioning different z - axis planes in the depth of field region of the objective . in the embodiment depicted in fig4 in addition to the original image series captured 41 , two other series represented by reference numerals 42 and 43 and two additional series represented by reference numerals 44 and 45 are also captured . by capturing and storing these additional images from different regions of the specimen , a virtual focusing capability can be realized as described below in more detail . it may also be realized that this method of scanning may be more suited to a type of triple pixel line sensor described above as a 3 by 2098 sensor . sometimes this is referred to as a single line sensor . in this case since small discrete individual tiles are not available , the prediction of a reference plane by computation , or simple assumption of a completely flat and parallel x , y - plane may be preferred . this type of scan results in saving images of longer strip tiles , with a width of 2098 pixels inside the field of view of the microscope objective 10 in one direction , but the saved images extending beyond the field of view by continuous scanning and storing in the other direction . the abutted 2098 pixel wide strip tiles taken together side by side form a virtual microscope slide . also as illustrated in fig4 there are two focal depth of fields above and two focal depth of fields below , both provided with respect to the initial reference setting . in a given application , it may be appropriate to provide only a single additional set of images using only one slightly different focal depth of field ( or focal plane ) above and below . for most purposes , however , images captured at a plurality of differing focal planes are appropriate . in the preferred embodiment , four focal planes above and four focal planes below are used in addition to the original reference focal plane to provide a total of nine sets of focal plane images . wherein each set of focal plane images corresponds to a given focal distance from the reference setting and all of the sets share the same level of magnification . by providing this many sets both above and below the reference focal plane , relatively smooth and detailed virtual focusing can be realized that well mimics the look and feel of focusing with an actual microscope within a useful range of focusing . as described in the above , the various depths of field substantially abut one another . in an alternative embodiment , and as illustrated in fig5 a given depth of field 51 for a given series of images can partially overlap with another depth of field 52 for a different series of images . or , if desired and as illustrated in fig6 different depths of field 61 and 62 as corresponding to different image series can neither overlap nor abut one another . instead , a small gap can exist between the two fields . in general , adjusting the focal distances such that the fields are substantially adjacent one another with little or no overlap probably represents an optimum configuration , but the other alternatives may be useful for some purposes depending upon user requirements . with reference to fig7 an initial focal plane 71 ( as initially determined or predetermined manually or automatically ) having a corresponding depth of field 41 a may be appropriately used when imaging a particular section of a specimen ( not shown ) that is within the field of view when the microscope is located in a first position 10 a . that is , when the microscope is so positioned , this initial focal distance 71 represents an optimum focus by whatever standard the user applies . in accordance with the various embodiments above , one or more additional images are also taken of this same field of view with slightly different focal distances . at another portion of the slide , however , when the microscope is positioned at a second position 10 b , it may be that a different initial focal plane 72 will yield an optimum focus when using the same standard as was applied earlier . this different initial focal distance 72 will have a corresponding depth of field 41 b that is substantially identical in size to the depth of field 41 a for the first position &# 39 ; s initial focal distance 71 but that is positioned a different distance from the slide 21 . this is often the case when imaging tissue microarray ( tma ) cores as described in u . s . pat . no . 6 , 466 , 690 b2 ( entitled method and apparatus for processing an image of a tissue sample microarray ). there the image capture is from a great many different objects , tma cores , arranged over essentially the entire surface of the glass microscope slide . therefore , while the resulting images still comprise a abutted composite representations of the object , they refer to different reference image focal planes . and , according to this embodiment , regardless of differences as may exist between the initial focal reference focal plane from object to object , each resulting image will nevertheless have an identical number of z stack focal planes available for fine focusing by a user . as discussed above , virtual microscope slides , whether created from many small tiles as in the preferred embodiment , or whether created in strips of line segments , and whether they are stored in a tiled data structure or whether they are stored as one large reconstructed image in memory , such as one focal plane from the set of 5 focal planes 91 in fig9 cannot usually be viewed in their entirety at the original captured resolution because of the finite size and pixel dimensions of a remote viewers computer display screen . as depicted in fig8 one prior art approach that is useful in this regard utilizes a plurality of individual images 83 , referred to as tiles , to form a larger composite image of the slide 81 and the specimen 82 . u . s . pat . no . 6 , 396 , 941 b1 ( entitled method and apparatus for internet , intranet and local viewing of virtual microscope slides ), the contents of which are incorporated herein by this reference , teaches the internet or intranet display of virtual microscope slides . as taught therein , a virtual microscope slide typically comprises a digitized magnified view of part or all of a microscope slide and an object ( such as a biological specimen ) disposed thereon . the aforementioned patent also teaches various internet server and thin client , and other java applet and activex viewer methods enabling the reconstruction of the virtual microscope image content for a remote viewer . it will be appreciated that the viewing of a single focal plane depth of view is accomplished whether the image is stored as a tiled database structure or as a complete single image plane in computer core memory . in the preferred embodiment of this invention however , when the remote viewer sends a request to the server for a reference image tile focus for a defined region of interest , the server also sends the associated z stack images all in sequence for that region of interest . the associated z stack images are cached by the local computer so that a smooth and rapid local viewing can simulate the analog optical focusing operation of a real microscope . referring now to fig1 , in one embodiment , a user can employ a standard computing platform to interface to the virtual slide server and data storage facility that retains the virtual microscope slide information as described above for a given specimen . a standard client / server model works well to facilitate such a relationship , but other data transfer mechanisms can be used as well as appropriate to a given application . the relevant process begins with a user platform retrieving 101 a desired image at a particular magnification × ( such as , for example , 40 ×). as described in the aforementioned patent , all images for the object need not be immediately retrieved and made available locally . to minimize network transactions , in fact , only the data required to display a single field of view need to be immediately retrieved and displayed . in the system and method of the current invention and the various embodiments above , each field of view has a corresponding plurality of images with each image representing a different focal plane . therefore , when retrieving and displaying the first image , one of these images must be selected first . in the preferred embodiment the selection is the reference image set corresponding to those tiles that will fill the view window of the remote viewers image display screen . also , the associated z stack images for each reference tile are transmitted and cached in the local computer . in one embodiment , where the initial automatically determined optimum focal plane image is flanked on each side by four different focal plane images , the initial image itself can be automatically selected for initial retrieval 101 and display 102 . the process then monitors 103 for instructions from the user to modify the focus . when no such instruction appears , the process continues 104 in accordance with whatever other functions are supported ( for example , input from the user indicating a desire to scroll the image in a particular direction can be received and used to cause retrieval and display of corresponding images ). when a focus modification instruction is received , however , the process retrieves 105 the image from the local memory cache for that field of view that corresponds to the instruction and displays 106 it . pursuant to one embodiment , the user can be limited to moving the focus in a step by step process with a mouse wheel 93 or keyboard 37 up or down arrow keys , such that each increment causes retrieval and display of the next adjacent image in the z - axis dimension . in the preferred embodiment the user , or remote viewer , can move about and focus on the virtual microscope slide with a wheeled mouse control , essentially as one moves about and focuses with a physical microscope and slide . with this capability , a wide variety of specimens can be readily viewed with good results . not only can the resultant virtual microscope slides be used for educational and training purposes , but also for both qualitative and quantitative analysis purposes in support of various diagnostic processes . with reference to fig1 , pursuant to one optional embodiment , when a user seeks to modify 103 the focus as described , the process can determine 111 whether a focusing limit has been reached . for example , if the user platform has already retrieved and displayed the image that was captured using the focal plane at the furthest z - axis dimension from the reference tile and the user is now instructing the platform to focus on an even further distance , the present display can be maintained 112 . optionally , a text message or other indicator can be provided 113 to the user to alert the user that the focus limit has been reached . in another embodiment , a visual indicator can be provided to the user to indicate a present focusing position within a range of focusing possibilities , such that the user can ascertain for themselves such a condition . those skilled in the art will recognize that a wide variety of modifications , alterations , and combinations can be made with respect to the above described embodiments without departing from the spirit and scope of the invention , and that such modifications , alterations , and combinations are to be viewed as being within the ambit of the inventive concept . it is intended in the appended claims to cover all those changes and modifications which fall within the true spirit and scope of the present invention .