Methods and Systems for Sample Analysis

Abstract

The present disclosure includes provides methods for analyzing biological samples to identify, classify, and/or quantify platelets in the sample. The present disclosure also provides systems and methods for analyzing a blood sample to determine presence of platelet clumps in the sample. Also provided are systems configured for performing the disclosed methods and computer readable medium storing instructions for performing steps of the disclosed methods.

1 - 14 . (canceled) 
     
     
         15 . A method of identifying a sample as comprising platelet clumps, the method comprising:
 incubating a sample with a reagent comprising a cell permeable fluorescent dye and a red blood cell lysis agent for a period of time sufficient to lyse red blood cells and to fluorescently stain nucleus-containing cells;   flowing the sample through a flow cell of the flow cytometer;   optically interrogating the sample flowing through the flow cell;   detecting optical signals from the interrogated sample;   identifying events associated with a fluorescent signal below a threshold, wherein identified events have a fluorescent signal below the fluorescent signal from white blood cells (WBCs); and   determining size of the identified events, wherein presence of events having a size larger than platelets indicates that the sample comprises platelet clumps.   
     
     
         16 . The method of  claim 15 , further comprising
 selecting the identified events that are of a larger size than that of platelets;   fitting a line to the selected events;   extending the line into signals associated with WBCs; and   reclassifying WBCs near the line as platelet clumps.   
     
     
         17 . The method of  claim 15 , wherein the signals comprise one or more of fluorescence (FL), intermediate angle scatter (IAS), polarized side scatter (PSS), or axial light loss (ALL). 
     
     
         18 . The method of  claim 15 , wherein the signals comprise fluorescence (FL) and polarized side scatter (PSS). 
     
     
         19 . The method of  claim 15 , wherein the sample is a whole blood sample. 
     
     
         20 . The method of  claim 15 , wherein optically interrogating the particles comprises exciting the cells using a laser. 
     
     
         21 . A non-transitory computer readable medium storing instructions that, when executed by a computing device, cause the computing device to:
 detect optical signals from an interrogated sample, wherein the optical signals are generated from flowing the sample through a flow cell of a flow cytometer and optically interrogating the flowing sample, wherein the sample has been incubated with a reagent, comprising a cell permeable fluorescent dye and a red blood cell lysis agent, for a period of time sufficient to lyse red blood cells and to fluorescently stain nucleus-containing cells;   identify events associated with a fluorescent signal below a threshold, wherein identified events have a fluorescent signal below the fluorescent signal from white blood cells (WBCs); and   determine size of the identified events, wherein presence of events having a size larger than platelets indicates that the sample comprises platelet clumps.   
     
     
         22 . The non-transitory computer readable medium of  claim 21 , wherein the instructions comprise instructions that cause the computing device to:
 select the identified events that are of a larger size than that of platelets;   fit a line to the selected events;   extend the line into signals associated with WBCs; and   reclassify WBCs near the line as platelet clumps.   
     
     
         23 . The non-transitory computer readable medium of  claim 21 , wherein the signals comprise one or more of fluorescence (FL), intermediate angle scatter (IAS), polarized side scatter (PSS), or axial light loss (ALL). 
     
     
         24 . The non-transitory computer readable medium of  claim 21 , wherein the signals comprise intermediate angle scatter (IAS) and polarized side scatter (PSS). 
     
     
         25 . A flow cytometer, comprising:
 a flow cell;   an optical particle interrogation system optically coupled to the flow cell; and   the non-transitory computer readable medium of  claim 21 .