Patent Description:
Conventional tissue samples, such as those processed and studied in a biobank or pathology department, or other similar facility, are often embedded into paraffin wax blocks for later sectioning or slicing. The paraffin block supports the tissue sample such that the tissue sample may be thinly sliced. Such slicing is generally performed with a microtome or other similar tissue slicing instruments. The paraffin embedded tissue blocks are sliced (sectioned) with each slice of the tissue placed onto respective slides. Each sample slice is processed and the final stained tissue sample slice covered and then stored. Each of the stored tissue slides may also be individually identified for later retrieval. <CIT> shows a method of capturing tissue sample images having the features of the preamble of claim <NUM>.

The present invention provides a method for capturing images of tissue samples as claimed in claim <NUM>. Embodiments of the present invention provide an apparatus and methods for imaging tissue biopsy cassettes, microscope slides, and other tissue containers as well as the specimen tissue therein contained (also known herein as tissue/block/sample containers).

An apparatus for barcode tracking, imaging, and analyzing images of tissue samples is described below. The apparatus includes an imager, controller, an actuated lens, a range finder, a sample holder, a lighting system, and a processor. The imager is configured to capture images within a selectable field of view. A sample/block/tissue container is positionable within the field of view. The imager is configured to capture images of a plurality of sample/tissue containers. The sample holder is configured to provide adjustable support for each type of sample container as well as providing an optical path for imaging barcodes not in a direct field of view, as well as providing an encoding system to identify the sample container type at the time of image capture. The lighting system is configured to illuminate the field of view. The processor is configured to receive a first plurality of captured images of sample/tissue containers. The processor is configured to analyze the first plurality of captured images and to locate and read the barcode for the purpose of providing a base filing identifier for automatic archiving of the images. Additional images of the first plurality of captured images are used for documentation purposes and are analyzed by the processor to determine whether there is tissue missing from any one of the first plurality of captured images.

In a further aspect of the present invention, a method for capturing images of tissue samples includes positioning tissue sample containers within a selectable field of view. The tissue sample containers positioned within the field of view are illuminated. Images of the tissue sample containers positioned within the field of view are captured. A first plurality of captured images of tissue sample containers are arranged as a progressive series of paired images. The method further includes storing the first plurality of captured images in an archive.

In another aspect of the present invention, a method for capturing images of tissue samples includes providing an imager system for capturing the images that includes a light source for lighting a field of view, a processor for processing the captured images, and a range finder for determining a focus of the imager. The method further includes determining a focus for the imager for a cut surface of a sample/block. The focus will vary depending on an amount of cutting performed prior to each imaging. The range finder is calibrated to focal positions of an actuated lens of the imager. The light source and the imager are positioned with respect to the cut surface at an angle of <NUM>-<NUM> degrees from perpendicular, such that the reflecting light angles off the sample/block to enhance the cut surface of the sample/block. The imager and the light source are positioned relative to each other at a sample angle. A first image is captured and processed with the processor to indicate where boundaries of the cut tissue exist in the first image. The light source is polarized such that light reflecting off the cut surface is eliminated, such that subsurface (uncut tissue) is observable. A second image is captured with the polarized light. The first and second images are compared by the processor. Such comparison includes overlaying one of the first and second images upon the other of the first and second images, such that those parts of the sample tissue are identified that are below the surface and therefore not cut and not transferred to a slide for examination. Lastly the method includes providing adjustable viewing controls to improve an observer's ability to distinguish those parts of the sample tissue that are below the surface of the sample/block.

In an aspect of the present invention, the range finder is a pair of ultrasonic time-of-flight (TOF) sensors.

In a further aspect of the present invention, the imaging system further includes a polarized light source and a shallow pocket configured to support standard, <NUM> x <NUM> microscopy slides and <NUM> x <NUM> microscopy slides on a white background for aiding in capturing images of barcodes to provide a base filing identifier, as well as capturing images suitable for processing to indicate where cut tissue boundaries exist.

In another aspect of the present invention, the processor is configured to compare a first image of a paraffin wax embedded tissue sample block to a second image of a tissue sample slide. The first image comprises an image of tissue present in the second image. The processor is further configured to determine whether any of the tissue present in the first image is missing from the second image.

Thus, tissue sample slides and paraffin wax embedded tissue sample blocks may be imaged and the resulting images archived (e.g., stored in a database in a memory). These archived images may then be indexed by patient/case such that a series of images related to a particular patient/case may be retrieved and analyzed at a later date. Such analysis may include image analysis of a progressive series of "final" slide images and block images to determine if there is any tissue missing from the final slide images.

These and other objects, advantages, purposes and features of the present invention will become apparent upon review of the following specification in conjunction with the drawings.

Referring to the drawings and the illustrative embodiments depicted therein, an imagery management system and methods are provided for archiving images of paraffin embedded tissue blocks (hereinafter "blocks") into the electronic files of a laboratory information system ("LIS") that are related to the tissues. The electronic files may also be related or linked to a patient from which the tissue sample was excised. One purpose of the imaging is to record a "pattern" of the cut tissue as well as the subsurface tissue present in a particular block for later comparison to images taken of the final stained and cover slipped slides (e.g., the slides may be imaged using whole slide imaging (WSI) to ascertain whether any tissue (i) was not cut that should have been cut, or (ii) was lost during tissue/cassette and slide processing (note that such tissue may also be "lost" from image acquisition and processing). Optionally, the imagery management system and methods may incorporate automated image recognition algorithms to determine if any tissue is missing from a final slide and provide a user with an alert to inform the user that a particular slide is missing tissue relative to the tissue block (i.e., the original tissue block images). Additionally, the imagery management system and methods may provide a data management system for acquired images that may be used for recording and reporting mismatches. Therefore, as discussed herein, the exemplary imagery management system includes an imaging apparatus, an image acquisition and archiving module, and an image analysis module.

<FIG> illustrates an exemplary imagery management system <NUM>, which includes an analyzer/image processor <NUM> and an imaging apparatus <NUM>. As illustrated in <FIG>, the analyzer/image processor <NUM> includes an image acquisition and archiving module and an image analysis module, both of which may be implemented as separate hardware modules of a multi-core graphics processing unit (GPU) <NUM> or as software modules implemented by a multi-core micro processing unit (CPU) <NUM>. <FIG> also illustrates the imagery management system <NUM> communicatively coupled to a server <NUM> that provides the storage and retrieval of archived imagery, such as a database <NUM> stored in a memory <NUM>. The imaging apparatus <NUM> may be implemented as either a stand-alone instrument <NUM> (see <FIG>) or the imager <NUM> may be incorporated as an accessory of a tissue slicing instrument <NUM> (e.g., a microtome or other similar instruments). The imaging apparatus <NUM> captures images of tissue samples <NUM> such as paraffin wax embedded tissue samples <NUM>, <NUM> (see <FIG> and <FIG>) or slides <NUM>, <NUM> carrying tissue sample slices (see <FIG> and <FIG>).

Optionally, and as illustrated in <FIG>, the imagery management system <NUM> includes external data ports <NUM> (such as USB ports) arranged on a housing to facilitate accessory connections (e.g., mouse, keyboard, barcode scanner, and thumb drive access). An external LAN port <NUM> or other suitable data connection may also be provided to allow for connection to an institution's network. An external power switch may be arranged for powering ON/OFF the lighting system <NUM>, <NUM>. Similarly, a main power switch for the analyzer/image processor <NUM> and imaging apparatus <NUM> may also be arranged on the housing of the imagery management system <NUM>.

The image acquisition and archiving module of the analyzer/image processor <NUM> provides a user interface (displayed on a display screen <NUM> of the imagery management system <NUM>) for control of the imaging apparatus <NUM>. The image acquisition and archiving module may also provide a user with access to a hospital or similar institute's information technology (IT) infrastructure and electronic archives. The image acquisition and archiving module may also read an identifier barcode on images of cassettes <NUM>, <NUM> (configured to hold paraffin wax blocks <NUM>, <NUM>) and tissue sample carrying microscopy slides <NUM>, <NUM> to allow for the automatic archiving of the associated images (see <FIG>). The image analysis module of the analyzer/image processor <NUM> analyzes the images of the blocks and the slide images (e.g., WSI) and provides a PASS/FAIL output to the user indicating if any tissue is missing from an image of a finalized slide <NUM>, <NUM>. Additionally, the image analysis module will archive the imagery analysis results into the specific patient/case electronic files stored in the LIS.

<FIG> illustrates the imaging apparatus <NUM> implemented as an exemplary stand-alone unit. The imaging apparatus <NUM> includes a computer-controlled imager <NUM> (as illustrated in <FIG>, the imaging apparatus <NUM> is communicatively coupled to the analyzer/image processor <NUM>, which, as noted above, controls the operation of the imaging apparatus <NUM>). Optionally, the imaging apparatus <NUM> is coupled to the analyzer/image processor <NUM> via a computer interface (e.g., USB <NUM>). The imager <NUM> includes a lens system <NUM> that is adjustable for a desired focus and/or iris setting. Optionally, the lens system <NUM> may be a manual lens with manually selectable focus, shutter speed, and iris (aperture) controls. In a further optional embodiment, the lens system <NUM> has automatic settings that are adjustable via the analyzer/image processor.

As illustrated in <FIG>, the imager <NUM> may be communicatively coupled to a lighting system <NUM> that provides specialized lighting settings to assist in effectively capturing suitable images of the tissue samples in the cassettes and on the slides. Optionally, the lighting system <NUM> may be computer controlled via the analyzer/image processor <NUM>. The lighting system <NUM> may also include polarizers that are used to illuminate a field-of-view with polarized light. Illuminating the field of view with polarized light can reduce glints or glares in the field of view of the imager <NUM>, thereby improving the imager's ability to obtain clear images of tissue samples, such as portions of tissue below a cut surface of the wax block. The field of view of the imager <NUM> is adjustable to cover the entire microscopy slide (WSI) <NUM>, <NUM> (carrying a tissue sample slice) or an entire paraffin wax embedded tissue sample (that is, a tissue sample <NUM> embedded in a paraffin wax block <NUM>, <NUM> that is carried by a cassette <NUM>, <NUM>). Optionally, there may be a plurality of computer-controlled lighting sources configured to enhance different aspects of the sample tissue containers. Note, that for the sake of clarity, the tissue samples <NUM> illustrated in <FIG>, <FIG> are illustrated without the cassettes <NUM>, <NUM> and the associated paraffin wax blocks <NUM>, <NUM> into which the samples <NUM> are embedded.

The imager <NUM>, lens system <NUM>, and lighting system <NUM> are arranged within and/or supported by a housing <NUM>. Optionally, the imager <NUM> and its associated lenses <NUM> may be sealed within the housing <NUM> with a window and access cover to keep dust, debris, and fluids from contaminating the imager <NUM> and its associated optical system (<NUM>). A ventilation port in the housing <NUM> may also be provided to facilitate fume abatement when used in the presence of typical pathology fixatives. The housing <NUM> will also allow for associated cabling to be unobstructed.

The imaging apparatus <NUM> also includes a positioning system <NUM> that provides for a quick change between imaging paraffin blocks <NUM> of different sizes (and thicknesses) and between slides <NUM>, <NUM> of different sizes, to ensure proper centering and focus of the items to be imaged. As illustrated in <FIG>, the positioning system <NUM> is mechanically coupled to the imager <NUM> via the housing <NUM>. Optionally, the positioning system <NUM> may be computer controlled via the image acquisition and archiving module of the analyzer/image processor <NUM>. The imager <NUM> may also be configured to function as a barcode reader for reading the barcodes on the cassettes and slides holding tissue samples <NUM>.

The imaging apparatus <NUM> includes a sample holder <NUM> with multiple configurable "positions. " A first position for the sample holder <NUM> is a "slide" position. When the sample holder <NUM> is in the slide position, the sample holder <NUM> ensures that a slide-mounted tissue sample <NUM> is held in a proper position for centering and focal distance for the imager <NUM>. The sample holder <NUM> may be configurable to accept and/or accommodate "whole mount" cassettes and "whole mount" slides. The sample holder <NUM> may be oriented into a "block" position. When the sample holder <NUM> is in the block position, the sample holder <NUM> ensures that a sample block <NUM> is held in a proper position for centering and focal distance for the imager <NUM>.

The paraffin wax blocks <NUM>, <NUM> (carried by cassettes <NUM>, <NUM>, respectively) and microscopy slides <NUM>, <NUM> (each carrying tissue samples <NUM>) are imaged with the imager <NUM> and its associated lens arrangement <NUM> selected and oriented to provide a field-of-view sufficient to image the entire tissue sample, with an optical diffraction blur circle equal to or less than the camera pixel size, and lockable adjustments on the lens system <NUM> of the imager <NUM> to allow imager settings for iris, focus, and polarization orientation. The exemplary fragments of tissue can be very small and will need to be visualized. It is also noted that embodiments of the present invention are directed to locating discrete sample pieces that were supplied in a sample container and then transferred into a cassette and then transferred into a paraffin wax substrate or embedding. In other words, exemplary embodiments are not directed to visualizing the details of the tissue sample, they are directed to visualizing whether or not the tissue is even present. Therefore, a typical core needle may be used as a guide to determine a smallest "practical or typical" tissue sample size to be tracked. For example, the smallest expected tissue sample size may be specified as ¼ of the diameter of a <NUM> gauge needle, or ½ x <NUM> = <NUM>. In other words, the number of pixels per inch depends upon the smallest "practical/typical" piece of tissue to be tracked. A minimum resolution may therefore be based upon a Nyquist sampling criteria of ><NUM> pixels per axial direction or ><NUM> pixels for <NUM> axis area detection resulting in a pixel pitch of <NUM>. In practicality, a safety factor of greater than twice per axial direction <NUM> x <NUM> = <NUM> or <NUM> pixels per <NUM> axis area. The largest tissue/block/sample containers to image will be "whole mount" slides with an area up to <NUM> x <NUM>. Combined with the pixel pitch (<NUM>) results in a minimum camera resolution of <NUM>,<NUM> x <NUM>,<NUM>.

The above described core or biopsy needles may typically range from <NUM> gauge to <NUM> gauge. Furthermore, the tissue samples acquired by such needles may break apart. When a tissue sample from a core/biopsy needle breaks apart, there could be ten or more smaller fragments within a single paraffin wax block that will need to be imaged. In addition, imagery management systems and methods of the present invention may be used to image blocks with tissue microarrays (TMAs), where the core of each tissue in a TMA can be as small as <NUM> in diameter, with a resulting required minimum resolution adjusted accordingly. In one embodiment, the imager <NUM> is a monochromatic imager. Other embodiments are also possible, such as color imagers and the like.

<FIG> illustrates an imager <NUM> installed as an accessory tool of a tissue slicing instrument <NUM>. The imager <NUM>, lens system <NUM>, and lighting system <NUM> of <FIG> may be the same as the imager <NUM>, lens system <NUM>, and lighting system <NUM> of <FIG>. The imager <NUM>, when installed as the accessory attachment, is positionable via a positioning system <NUM> to be properly oriented for capturing images of paraffin wax embedded tissue samples <NUM> placed/mounted onto a working surface <NUM> of the tissue slicing instrument <NUM>. The tissue slicing instrument <NUM> has an adjustable slicing depth, such that a thickness of a slice may be selected. For example, a slice from a side of a tissue sample <NUM> may be typically <NUM>-<NUM>, but may range to <NUM> to <NUM>. When properly positioned, the imager <NUM> captures images of the tissue samples <NUM> during the sectioning or slicing process. As illustrated in <FIG>, the imager <NUM> is positioned to capture an image of an outer surface of a tissue sample <NUM> before that outer surface of the tissue sample <NUM> is sliced away from the tissue sample <NUM> by the tissue slicing instrument <NUM>. That image of the tissue sample <NUM> may then be compared to a later image of that tissue sample slice after it has been treated and coverslipped. The comparison is performed to determine whether or not any tissue is missing from the later image. The comparison may also be performed to determine whether or not the tissue has become misaligned on the slide, or to detect other anomalies.

The image acquisition and archiving module of the analyzer/image processor <NUM> provides the following major functions: a live image preview function, an image acquisition function, a block finished/end cut image acquisition function, an image review function, and a pathologist review function. The live image preview function provides for a live image to be displayed on the display screen <NUM> that is visible to the user. Using the live image displayed on the display screen <NUM>, the user is then able to setup the imager <NUM>, <NUM> and the lighting <NUM>, <NUM>. The display screen <NUM> may optionally be a touch responsive display. Other embodiments are also available, e.g., a detachable display screen <NUM> or cabling to a remote display screen <NUM>.

The image acquisition functionality of the analyzer/image processor <NUM> may be triggered to capture an image of each slide <NUM>, <NUM> that has a tissue sample and a barcode label on it. As discussed herein, the imager <NUM> may optionally include a barcode reader. Similarly, a tissue sample block <NUM> may also be interrogated or scanned for a barcode. Once the associated barcode is read, its data may be parsed into appropriate field variables of a patient/case electronic record. Such records may be stored in a database <NUM> in either local memory <NUM> or a remote memory <NUM> of a server <NUM>. Using the field variables, the captured image may be stored in the database <NUM> for the associated case/patient with field metadata indicating case/patient number, slide or tissue block number, and any other necessary fields.

The block finished/end cut image acquisition function of the analyzer/image processor <NUM> provides for the capturing of a first image of the paraffin block (and embedded tissue sample) <NUM> before any sections (slices) have been taken. A subsequent final image (after any sections/slices have been removed from the paraffin block <NUM>) is also taken of the finished/end cut condition of the paraffin block <NUM> before the paraffin block <NUM> is filed away. Similar to the functionality discussed above with respect to processing slide mounted tissue samples <NUM>, using the field variables from side barcodes previously captured, each image of a tissue sample <NUM> embedded in a wax block <NUM>, <NUM> is stored in the database <NUM> for the case/patient with field metadata.

<FIG> illustrates an alternative imaging apparatus <NUM>, which includes a computer-controlled imager <NUM>. As illustrated in <FIG>, the imaging apparatus <NUM> is communicatively coupled to the analyzer/image processor <NUM>, which as noted above, controls the operation of the imaging apparatus <NUM>. Optionally, the imaging apparatus <NUM> is coupled to the analyzer/image processor <NUM> via a computer interface (e.g., USB <NUM>). The imager <NUM> includes a lens system <NUM> that is adjustable for a desired focus and/or iris setting. Optionally, the lens system <NUM> may be a manual lens with manually actuated focus, shutter speed, and iris (aperture) controls. In a further optional embodiment, the lens system <NUM> has automatic settings that are actuated via the analyzer/image processor <NUM>. In one embodiment, the imager <NUM> is a monochromatic imager. Other embodiments are also possible, such as color imagers and the like.

As illustrated in <FIG>, the imaging apparatus <NUM> also includes a range finder <NUM> for determining a range between the imager <NUM> and a surface of a paraffin wax block <NUM>, <NUM>. In one embodiment, the range finder <NUM> is a pair of ultrasonic, time-of-flight (TOF) sensors. A first ultrasonic sensor is used to read a standard reference distance in order to provide a temperature, pressure, and humidity compensation factor. A second ultrasonic sensor measures the focal distance of interest, and uses the compensation factor from the first sensor to ensure that the focal distance is measured accurately in the present environmental conditions. As discussed herein, the range can vary depending on the type of cassette (<NUM>, <NUM>) used, as well as varying according to the number of slices that have been sliced (sectioned) from the tissue embedded paraffin wax block. The range between the cut surface of the paraffin wax block <NUM>, <NUM> and the range finder <NUM> will increase as the sectioning process continues. Once the range finder <NUM> determines a distance to the surface of the paraffin wax embedded tissue sample, a focal length is determined such that the lens system <NUM> adjusts to focus on the surface of the paraffin wax embedded tissue sample. Optionally, the imaging apparatus <NUM> determines the updated focal length and adjusts the focus of the lens system <NUM> in an automated fashion. In an alternative embodiment, using the updated focal length information, the lens system <NUM> is manually adjusted by an operator to bring the surface of the paraffin wax embedded tissue sample into focus.

The imaging apparatus <NUM> also includes a reflected light panel <NUM> and an oblique polarized light panel <NUM> (and associated polarizer <NUM>). Controlled by the imaging apparatus <NUM>, the light panel <NUM>, the polarized light panel <NUM>, and polarizer <NUM> provide specialized lighting settings to assist in effectively capturing suitable images of the tissue samples in the cassettes <NUM>, <NUM> and on the slides <NUM>, <NUM>. Optionally, the lighting system (<NUM>, <NUM>) may be computer controlled via the analyzer/image processor <NUM>. As discussed herein, illuminating the field of view with polarized light (via the polarized light panel <NUM> and polarizer <NUM>) can reduce glints or glares in the field of view of the imager <NUM>, thereby improving the imager's ability to obtain clear images of tissue samples <NUM>, such as portions of tissue (202b) below a cut surface of the wax block <NUM>, <NUM>. The field of view of the imager <NUM> is adjustable to cover the entire microscopy slide <NUM>, <NUM> or an entire paraffin wax block <NUM>, <NUM>, each wax block including an embedded sample <NUM>. As also illustrated in <FIG>, the lighting system <NUM> and the imager/lens system (<NUM>, <NUM>) are positioned with respect to the cut surface of the wax block <NUM>, <NUM> at an angle of about <NUM>-<NUM> degrees from perpendicular, such that the reflecting light (from the lighting system <NUM>) angles off the wax block <NUM>, <NUM> and enhances the cut surface of the wax block. The effect of angled light reflecting off the cut surface of the wax block <NUM>, <NUM> is discussed in detail below. As also illustrated in <FIG>, the lighting system <NUM> and the imager/lens system (<NUM>, <NUM>) are positioned relative to each other at a suitable angle so that the cut surface of the wax block is highly reflective of the light, which is directed from the wax block to the lens, and creates significant contrast between the cut surface of the wax block and the cut surface of the tissue in the wax block.

The imager <NUM> and lens system <NUM> may be arranged within or supported by a housing <NUM>. Optionally, the imager <NUM> and its associated lenses <NUM> may be sealed within the housing <NUM> with a window and access cover to keep dust, debris, and fluids from contaminating the imager <NUM> and its associated optical system <NUM>. A ventilation port in the housing <NUM> may also be provided to facilitate fume abatement when used in the presence of typical pathology fixatives. The housing <NUM> will also allow for associated cabling to be unobstructed.

The imaging apparatus <NUM> also includes a positioning system <NUM> that provides for a quick change between imaging paraffin blocks <NUM>, <NUM> of different sizes (and thicknesses) and between slides <NUM>, <NUM> of different sizes, to ensure proper centering and focus of the items to be imaged. The positioning system <NUM> is mechanically coupled to the imager <NUM> via the housing <NUM>. Optionally, the positioning system <NUM> may be computer-controlled via the image acquisition and archiving module of the analyzer/image processor <NUM>. As discussed below, the imager <NUM> may also be configured to function as a barcode reader for reading the barcodes on cassettes <NUM>, <NUM> and slides <NUM>, <NUM> holding tissue samples <NUM>.

The imaging apparatus <NUM> includes a sample holder <NUM> with multiple configurable "positions. " As illustrated in <FIG>, the sample holder <NUM> is equipped with a plurality of feet <NUM> that include a plurality of detents for position stops, as well as a plurality of Hall Effect sensors <NUM> for position encoding. A first position for the sample holder <NUM> is a "slide" position. When the sample holder <NUM> is in the slide position, the sample holder <NUM> ensures that a slide-mounted tissue sample <NUM> is held in a proper position for centering and focal distance for the imager <NUM>. The focal length may be confirmed with the range finder <NUM>. The sample holder <NUM> may be configurable to accept and/or accommodate "whole mount" cassettes and "whole mount" slides. Next, the sample holder <NUM> may be oriented into a "block" position. When the sample holder <NUM> is in the block position, the sample holder <NUM> ensures that a sample <NUM> embedded in a wax block <NUM>, <NUM> and carried on a cassette <NUM>, <NUM> (see <FIG> and <FIG>), is held in a proper position for centering and focal distance for the imager <NUM>. Optionally, the block position may be divided into a plurality of sub-positions, such as a "small cassette" (<NUM>) position and a "large cassette" (<NUM>) position.

<FIG> illustrates the imager <NUM> used to read a barcode <NUM> printed on a cassette <NUM>, <NUM> without having to remove the cassette <NUM>, <NUM> from the sample holder <NUM>. The cassettes <NUM>, <NUM> illustrated in <FIG> and <FIG> have barcodes <NUM> that are facing downward at a <NUM>-degree angle. As illustrated in <FIG>, the sample holder <NUM> includes a cavity <NUM> configured for retaining a cassette <NUM>, <NUM> positioned on the sample holder <NUM>. The sample holder's cavity <NUM> includes a mirrored surface <NUM> (or a mirror <NUM> positioned against that surface of the cavity <NUM>) that is positioned to allow the imager <NUM>'s field of view to view the barcode <NUM>. As illustrated in <FIG>, the field of view of the imager <NUM> is reversed by the mirror such that the barcode <NUM> is imaged right side up without need for removing the cassette <NUM>, <NUM> from the sample holder cavity <NUM>.

<FIG> illustrates a method for image enhancement for clearly identifying the cut tissue boundaries of a tissue sample <NUM> embedded in a wax block <NUM>, <NUM> as it is sliced (sectioned). As discussed herein, multiple tissue samples <NUM> can be embedded in a single wax block <NUM>, <NUM>, and can be embedded at different depths. Furthermore, even if only a single sample <NUM> was embedded into a wax block <NUM>, <NUM>, it is possible for the tissue sample <NUM> to be improperly or unusually placed in the wax block <NUM>, <NUM>, e.g., at an odd or incorrect angle. Therefore, whenever tissue-embedded wax blocks are sectioned, there is a concern as to whether all of a tissue sample has been completely sectioned or whether there is more of the tissue sample <NUM> deeper into the wax block <NUM>, <NUM>. Such determinations are complicated by the fact that the wax is slightly translucent and it is not always clear what portions of the sample <NUM> are below the cut surface of the wax block <NUM>, <NUM>.

To address these issues, an image enhancement process is provided that combines a first image of the tissue sample embedded wax block <NUM>, <NUM> illuminated with polarized light, with a second image of the tissue sample embedded wax block <NUM>, <NUM> illuminated with non-polarized light. When the polarized light source (<NUM>) and polarizer <NUM> are used, the captured image contains details of a tissue sample that is deep within the wax block <NUM>, <NUM> because the surface reflections found in traditional images of the wax block are absent. When the second image is captured, the glancing, non-polarized light allows for the relatively dull surface of the tissue sample <NUM> to be easily visualized against the relatively shiny surface of the wax block <NUM>, <NUM>. Thus, while the first image captures details deep within the wax block <NUM>, <NUM>, the second image captures surface details, such as an outline of the tissue sample <NUM> at the cut surface of the wax block <NUM>, <NUM>. Note that the resulting outline of the tissue sample <NUM>, <NUM> is of a cut or sectioned surface of the wax block <NUM>, <NUM>. Thus, when the first or second image is overlaid upon the other image, a practitioner or an image analysis program is able to view both the below-surface portions (202b) of the embedded tissue sample <NUM> (from the first image) and the outline (202a) of the tissue sample <NUM> present at the cut surface of the wax block <NUM>, <NUM> (from the second image) (see <FIG>). Optionally, an intensity of the polarization or edge detection may be adjusted to change both the image overlay and the edge detection. That is, by adjusting the image processing, the practitioner or image analysis software is able to focus on a desired feature of the wax block <NUM>, <NUM> (e.g., seeing more deeply into the wax block <NUM>, <NUM> and/or more clearly identifying an outline of the sample <NUM> at the cut surface of the wax block <NUM>, <NUM>.

The enhancement process begins in step <NUM> of <FIG>, where a first image is captured of a cassette <NUM>, <NUM> holding a wax block <NUM>, <NUM> with an embedded tissue sample <NUM>. As discussed herein, the embedded tissue sample <NUM> is located within the imager's field of view when the cassette <NUM>, <NUM> is positioned in the cavity <NUM> of the sample holder <NUM>. This first image is captured with polarized light (utilizing the oblique polarized light panel <NUM> and the polarizer <NUM>). With the first image captured with polarized light, the normal reflections and glare have been greatly reduced and sub-surface details (202b) below the cut surface of the wax block <NUM>, <NUM> will be visible. This is illustrated in <FIG>, where details of the tissue sample <NUM> are visible (cut tissue 202a, and uncut tissue 202b). As discussed herein, the cut tissue 202a is found within that portion of the tissue <NUM> that has been sliced (sectioned), while the un-cut tissue 202b is that portion of the tissue sample <NUM> that is below the surface of the wax block <NUM>, <NUM> and is yet un-cut. <FIG> illustrates that while using polarized light allows for subsurface details of the tissue sample <NUM> to be visualized, those portions of the tissue sample <NUM> that are cut tissue (202a) are hard to identify with this single image.

In step <NUM> of <FIG>, a second image is captured of the cassette <NUM>, <NUM> of step <NUM> (still holding the same wax block <NUM>, <NUM> with the embedded tissue sample <NUM>). This second image is captured with non-polarized light (utilizing the reflected light panel <NUM>). With the second image captured with angled, non-polarized light, glare and reflections are purposefully produced upon the waxy surface of the wax block <NUM>, <NUM>, while low reflections are found in other portions of the second image that are areas with cut tissue 202a, which are particularly dull. This results in significant contrast between the waxy portions of the wax block <NUM>, <NUM> that do not contain cut tissue 202a, and those portions of the surface of the wax block <NUM>, <NUM> that do contain the cut tissue portions 202a. This is illustrated in <FIG>, where details of the cut tissue portions 202a are more easily visualized as compared to in <FIG>.

In step <NUM> of <FIG>, an image of the barcode <NUM> is captured. As illustrated in <FIG>, the field of view of the imager <NUM> is shifted to find the reflection of the barcode <NUM> in the mirror <NUM>. Because of the changing field of view, the imager <NUM> will refocus to find the barcode <NUM>.

In step <NUM> of <FIG>, the second image is used as an overlay atop the first image. In the alternative, the first time is used as an overlay atop the second image. Such an overlay image is illustrated in <FIG>. In <FIG>, the overlay image (created from the combination of the first and second images) is further processed. The image processing may include: <NUM>) conversion of the images to monochrome, <NUM>) histogram adjustment for contrast enhancement to aid in identifying the edges of the cut tissue 202a, <NUM>) actual edge detection, <NUM>) palletization of the color channel to enhance viewing, and then <NUM>) presenting the image as an overlay with adjustable transparency and intensity controls. The enhanced tissue boundary images may then be used for analysis of the plurality of images taken from cut wax blocks, raw slides, and fully processed slides to identify any loss of tissue between paired images of the same tissue sample (comparing an original image of a wax block-embedded tissue sample to an image of a slide mounted tissue sample slice).

The image processing is further illustrated in <FIG>. In <FIG>, the overlay image of <FIG> has been converted into a monochromatic (black & white) image. In <FIG>, the black and white image of <FIG> is further converted into a "negative" image to aid in visualization of the tissue sample boundaries. Meanwhile, <FIG> and <FIG> illustrate the boundaries of the cut tissue 202a and the boundaries of the un-cut tissue 202b (below the surface of the wax). Boundaries of the cut tissue 202a are illustrated in <FIG>, while boundaries of the un-cut tissue 202b are illustrated in <FIG>. These boundaries may be adjusted through image processing to make either or both of the boundaries more visible or to provide desired information related to the cut and un-cut tissue portions 202a, 202b.

The image acquisition functionality of the analyzer/image processor <NUM> also provides for an image view mode that is displayed on the display screen <NUM>. The image view mode allows the user to inspect captured images and either accept or reject the image they are currently viewing. If rejected, the image acquisition functionality will prompt the user to retake the image (or images).

The image review function may be used to call up previously stored images (from the database <NUM>) from a selected case/patient entry to review the called-up images. The image review function allows those images to be measured and annotated, as well as exported.

The pathologist review function of the analyzer/image processor <NUM> allows for a pathologist, or other professional, to sign out a case for review. The pathologist or other professional may then review any matching tissue sample embedded wax blocks <NUM>, <NUM> or slides <NUM>, <NUM> (associated with the patient/case) by looking them up from within a case archive or from within the LIS.

The image analysis module of the analyzer/image processor <NUM> may be included as part of an image acquisition workstation or as a standalone image review system. As part of an imaging workstation, the image analysis module will also support a live mode preview of the tissue slides <NUM>, <NUM>. The image analysis module will also support image capture, barcode reading and parsing from the captured image data. The image analysis module will also associate the body of the slide <NUM>, <NUM> to a barcode read. The archived slide images (including the start and finished/end images) may be retrieved from the electronic medical records (stored in the database <NUM>) for use by the image analysis module of the analyzer/image processor <NUM>. The image analysis module may be used to determine if there is tissue missing from the corresponding scanned slides <NUM>, <NUM>. The image analysis may include the use of tissue centroid positions, tissue area comparisons, and previous and next slide progressions to determine whether centroid and area progressions are within reason or are questionable and require user review. As used herein, centroid or area "progression" refers to a change of location of the centroid or area of a tissue sample as determined from one image to the next. Lastly, as a part of an image acquisition work station, the image analysis module includes interfaces for: flagging slides <NUM>, <NUM> for user review, archiving the images and results to the LIS for the case, and reporting to review completed cases via associated images, data, and reports.

As part of a standalone image review system, the image analysis module of the analyzer/image processor <NUM> will support the retrieval of slide images (including the start and finished/end images) from the LIS. With the retrieval of the slide images, as discussed above, image analysis may be performed to determine if there is tissue missing from the slide images. The image analysis may include the use of tissue centroid positions, tissue area comparisons, and previous and next slide progressions to determine whether centroid and area progressions are within reason or are questionable and require user review. Similar to the imaging workstation, when the image analysis module is included as part of a standalone image review system, the image analysis module includes interfaces for: flagging slides <NUM>, <NUM> for user review, archiving the images and results to the LIS for the case, and reporting to review completed cases via associated images, data, and reports.

<FIG> illustrates the steps to a method for image analysis used to determine if there is any tissue missing from slide images. In step <NUM>, images are retrieved from an electronic file archive (e.g., the database <NUM> either stored locally or remotely in a server <NUM> that is part of a LIS. The images associated with a particular case/patient may be selected from a case index. The selected images may include images of tissue slices on microscopy slides <NUM>, <NUM>, and tissue samples <NUM> embedded in paraffin wax blocks <NUM>, <NUM>. In step <NUM>, a series of slide images are compared in a progression of images. For example, a first image of a cut surface of a paraffin wax embedded tissue sample <NUM>, along with a progressive series of images of the surface of the paraffin wax embedded tissue sample <NUM> during sectioning, are compared to a series of "final" slide images that include the surface of the paraffin wax embedded tissue sample <NUM> after it has been processed and mounted in a slide <NUM>, <NUM>. In other words, an image of each tissue sample surface before sectioning is compared to a slide image of that tissue sample surface after sectioning and final processing. By comparing "before" and "after" images, in step <NUM> it can be determined whether there is any tissue missing from the after or final slide images, or whether there are any other anomalies. Steps <NUM>, <NUM>, and <NUM> are performed in the GPU <NUM> or alternatively in the CPU <NUM>.

The imagery management system <NUM> may also include provisions to facilitate HIPAA compliant data transmission and encryption and backup protocol features to ensure data security. Such steps are related to the encryption of transmitted data as well as a buffered hierarchical database structure that will store data locally for use during LAN outages.

When the imagery management system <NUM> locates data targeted for the workstation that it is on, it sends a notification to the user's own desktop computer or other personal computing device that new data is available. The user then selects the data, copies the data to temporary memory ("clipboard"), such as memory <NUM>, and then can transfer ("paste") the data into the laboratory information system tracking input field found in the server's memory <NUM>. This can save the user time and reduce the occurrence of error related to missed containers.

A tissue chain of custody module may be configured to work in conjunction with the scanned data stored in the database <NUM> and will provide several quality assurance (QA) evaluations. In an exemplary manual evaluation mode, slide images are captured from a slide index, the barcode is read (on the image), and then the corresponding images for the slide just after tissue cutting and the cut block are retrieved for comparison to a final stained slide image for the operator to evaluate if any tissue has been lost or significantly altered during processing. The operator is also able to determine if any tissue was not cut at the current cut level (that is, was displaced deeper in the paraffin). In an automated evaluation mode, the just-cut slide image will be compared to the final stained slide image. Image analysis will be used to determine if any tissue is missing in the final slide. The automated evaluation mode will also flag any slides that exhibit differences for the operator to review. Images of the cut block will also be provided to the operator such that the current slide may be evaluated for cut depth issues.

Thus, microscopy slides <NUM>, <NUM> and paraffin wax embedded tissue samples <NUM> (e.g., tissue samples <NUM> embedded into wax blocks <NUM>, <NUM> that are mounted on cassettes <NUM>, <NUM>, respectively) may be imaged and the resulting images archived (e.g., stored in a database <NUM> in a memory <NUM>). These archived images may then be indexed by patient/case such that a series of images related to a particular patient/case may be retrieved and analyzed at a later date. Such analysis may include image analysis of a progressive series of "final" slide images and block images to determine if there is any tissue missing from the final slide images. In other words, an image of each tissue sample surface before sectioning is compared to a slide image of that tissue sample surface after sectioning and final processing (e.g., staining). By comparing "before" and "after" images, it can be determined whether there is any tissue missing, altered, or displaced from the after or final slide images.

Claim 1:
A method for capturing images of tissue samples, the method comprising:
positioning a tissue sample within a selectable field of view, wherein the tissue sample (<NUM>) is embedded in a substrate block (<NUM>,<NUM>);
illuminating the substrate block (<NUM>,<NUM>) embedded tissue sample (<NUM>) with a polarized light;
capturing a first image of the substrate block embedded tissue sample (<NUM>) from light reflected by the tissue sample (<NUM>) positioned within the field of view;
illuminating the substrate block embedded tissue sample (<NUM>) with a non-polarized light;
capturing a second image of the substrate block embedded tissue sample (<NUM>) from light reflected by the tissue sample (<NUM>) positioned within the field of view; characterized by
processing the first and second images to create an overlay image comprising the second image overlaying the first image;
identifying cut tissue (202a) in the substrate block embedded tissue sample (<NUM>), wherein the cut tissue is tissue present at a cut surface of the substrate block after slicing;
identifying uncut tissue (202b) in the substrate block embedded tissue sample (<NUM>), wherein the uncut tissue is tissue present below the cut surface of the substrate block after slicing;
identifying a boundary of the cut tissue with respect to the uncut tissue; and
storing the overlay image in an archive.