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US 5,962,238 AGranted
Method and apparatus for cell counting and cell classification
AssigneeBiometric Imaging, Inc.
InventorsNing L. Sizto, Louis J. Dietz
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AssigneeBiometric Imaging, Inc.
InventorsNing L. Sizto, Louis J. Dietz
Estimating the term…
A method and an apparatus for analyzing a material within a container, such as blood within a capillary in a volumetric cytometry system provides for detecting the edges of the container, counting the cells within the container, characterizing the cells within the container, and evaluating channels of data which contain information relevant to more than one of the detectable characteristics of the cells. A scanner scans a container of material including certain cells. Sampling circuitry is coupled to the scanner to generate scanned images of the material in the container. Two or more scanned images are generated based on fluorescence data from dyes that have overlapping spectra. The two scanned images are processed using a linear regression analysis among corresponding pixels in the scanned images near certain cells to characterize relative contents of two fluorescing dyes in a target cell. Target cells are identified from the scanned images using processing resources which identify a peak sample within a neighborhood, and compare the amplitude of the peak with the amplitude of pixels on the perimeter of the neighborhood. Upon identifying a target cell in this manner, data from the plurality of scanned images corresponding to the identified cell are saved for further analysis.
A method of detecting peak events indicative of target components in a fluid sample contained in a sample holder, the method comprising:
A method of analyzing a sample of cells in a sample holder, the sample of cells having detectable fluorescent characteristics, the method comprising:
A method of analyzing a capillary containing a sample of cells having detectable fluorescent characteristics, the method comprising:
A method of analyzing a sample containing fluorescent labeled target cells in a sample holder that contains a fluorescent background which exhibits background characteristics, the method comprising the steps of:
Dependencies are inferred from the claim text.
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Type of objects: microscopic objects, e.g. biological cells or cellular parts
Optical investigation techniques, e.g. flow cytometry: with spatial resolution of the texture or inner structure of the particle
Chemistry: molecular biology and microbiology: high energy substrates, e.g. fluorescent, chemiluminescent, radioactive
Chemistry: molecular biology and microbiology: simultaneous determination of more than one analyte
Term ended 17 Feb 2013 (estimated)
USPTO file history isn’t available for this patent, so only indexed dates are shown.
1. Field of the Invention
The present invention relates generally to the processing of volumetric capillary cytometry data for the purpose of counting and characterizing cells or cell constituents in a volume of material.
2. Description of Related Art
FIG. 1 is a plan view of a scanner apparatus with a data processing system according to the present invention.
FIG. 2 is a schematic block diagram of the data processing system used in combination with the system of FIG. 1.
FIG. 3 is a schematic diagram illustrating the scanning process used in the scanner of FIG. 1.
A detailed description of preferred embodiments of the present invention is provided with, respect to the figures. FIGS. 1 and 2 illustrate a hardware environment for the present invention. FIGS. 3-13 provide an explanation of the processing resources used to count and characterize target cells or cell constituents according to the present invention.
As can be seen in FIG. 1, an apparatus for volumetric capillary cytometry is provided. The machine is designed to process material within a capillary 10, which has a known volume. In the preferred system, the capillary may be rectangular in cross-section, having a width of about 0.4 millimeters to 1.5 millimeters, a length of about 40 millimeters, and a depth of about 25 to 225 microns, and in one embodiment, 100 microns. This capillary is suitable for detecting or characterizing a variety of cells or cell constituents. In one embodiment, it is used for the characterization of a CD3/CD4 assay, in which concentration of CD3 and CD4 antibodies are to be determined. In this type of assay, there are typically three populations present: CD3 positive cells stained with a Cy5 labelled antibody only; CD4 cells stained with both a Cy5 labelled CD3 antibody and a CyFr labelled CD4 antibody. Monocytes are stained with the CyFr labelled CD4 antibody only. It is found that the cells of interest in this assay are about 10 microns in diameter and are well classified using a capillary of the dimensions outlined above. It will be appreciated that the present invention is not limited to the characterization of a CD3/CD4 assay.
According to one method useful with the invention, a biological fluid, such as whole uncoagulated blood, can be reacted with an excess amount of a binding agent that contains a fluorophore excitable at a given wavelength. The fluorescently-labeled binding agent is selected to react with binding sites present within the sample. For example, a fluorescently-labeled antibody directed to the CD4 cell surface marker present on some leukocyte blood subclasses is reacted with a sample of whole blood. The labeled binding agents and the binding sites, i.e. labeled anti-CD4 antibodies and the surfaces of CD4-bearing leukocytes in the example, form fluorescent complexes that will emit a signal when used with the apparatus of the present invention.