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System for microvolume laser scanning cytometry

  • US 7,336,812 B2
  • Filed: 02/03/2004
  • Issued: 02/26/2008
  • Est. Priority Date: 07/21/1999
  • Status: Expired due to Fees
First Claim
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1. A method for analyzing a sample containing particles to detect and characterize target particles having a plurality of detectable characteristics in a fixed volume capillary that contains a fluorescent background and which exhibits background characteristics, the method comprising:

  • (a) scanning the fixed volume capillary containing the sample to generate a plurality of channels of data, wherein each channel of data comprises a distinct detectable characteristic and a distinct background characteristic;

    (b) sampling each of the channels of data to produce corresponding sets of pixel values;

    (c) generating sets of enhanced pixel values by independently modifying each set of pixel values to selectively enhance spatial features that are indicative of a target particle;

    (d) removing from one or more sets of enhanced pixel values the distinct background characteristic for the corresponding channel;

    (e) independently establishing threshold values for the detection of said particles for each set of enhanced pixel values;

    (f) independently identifying, in each set of enhanced pixel values, groups of above-threshold pixels located in patterns that are diagnostic of said particles;

    (g) independently identifying, for each group of above-threshold pixels located in a diagnostic pattern in a particular set of enhanced pixel values, the corresponding below-threshold or at-threshold pixels in the remaining sets of enhanced pixels in the remaining spectral channels;

    (h) characterizing the target particles in the sample by analyzing the pixels independently identified in steps (f) and (g); and

    (i) calculating at least one of;

    (I) a volume of the sample scanned; and

    (II) an absolute particle count;

    wherein particles are initially identified and analyzed in channels with above-threshold pixels located in patterns diagnostic of said particles, and said particles are then independently analyzed in all remaining channels by locating pixels in the same positions as the above-threshold pixels initially identified.

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