Cellular Analysis of Body Fluids

Abstract

Herein is provided a simple, reliable and accurate method for cellular analysis on hematology analyzers. In various aspects, the methods provide separation and/or differentiation between red blood cells (RBCs) and white blood cells (WBCs) by utilizing a fluorescent dye to selectively stain WBCs such that they emit stronger fluorescence signals. The method provides optimal detection limits on WBCs and RBCs, thereby allowing analysis of samples with sparse cellular concentrations. As few as one reagent may be used to prepare a single dilution for body fluid analysis, in order to simplify the body fluid analysis. Minimal damage to WBCs is attained using the lysis-free approach described in aspects of the disclosure.

1 .- 21 . (canceled) 
     
     
         22 . A hematology system comprising:
 a sample holder for holding a sample;   a mixing receptacle for mixing a staining composition with at least a portion of the sample;   a storage receptacle for storing the staining composition, wherein the staining composition comprises a dye that permeates a cell membrane and binds to a nucleic acid, and wherein the dye is fluorescent when bound to a nucleic acid;   a flow cell;   at least one energy source for applying electromagnetic energy to the flow cell;   a detector configured to detect fluorescence originating from within the flow cell to differentiate cells with nuclei from cells without nuclei based only on the presence or absence of the fluorescent dye; and   one or more visible light detectors for detecting scattered visible light.   
     
     
         23 . The hematology system of  claim 22 , further comprising an aspirating mechanism for aspirating at least a portion of the sample from the sample holder and moving it to the mixing receptacle. 
     
     
         24 . The hematology system of  claim 22 , further comprising an aspirating mechanism for aspirating the staining composition and moving it from the storage receptacle to the mixing receptacle. 
     
     
         25 . The hematology system of  claim 22 , wherein the at least one energy source produces monochromatic light at a wavelength λ1. 
     
     
         26 . The hematology system of  claim 25 , wherein the dye absorbs light at wavelength λ1 and emits light at wavelength λ2 when the dye is bound to a nucleic acid. 
     
     
         27 . The hematology system of  claim 26 , wherein the detector detects light at wavelength λ2. 
     
     
         28 . The hematology system of  claim 22 , further comprising at least one additional energy source for providing non-monochromatic visible light. 
     
     
         29 . The hematology system of  claim 22 , wherein the sample is a body fluid. 
     
     
         30 . A method for preparing a hematology system, the method comprising:
 providing a mixing receptacle for mixing at least a portion of a sample to be analyzed with a staining composition;   providing a storage receptacle for storing the staining composition, wherein the staining composition comprises a dye that permeates a cell membrane and binds to a nucleic acid, and wherein the dye is fluorescent when bound to a nucleic acid;   providing a flow cell;   establishing a stable core stream to allow the sample to pass through the flow cell;   positioning an energy source for providing electromagnetic energy to the flow cell; and   positioning a detector for detecting fluorescence emitted from within the flow cell.   
     
     
         31 . A method for preparing a receptacle for use in analyzing a body fluid, the method comprising adding to the receptacle a composition comprising a dye that permeates a cell membrane and binds to a nucleic acid, wherein the dye is fluorescent when bound to a nucleic acid, and wherein the receptacle is an integrated or modular part of a hematology system.