Download Red Blood Cell Lysis

Flow cytometry whole blood samples: Red
blood cell Lysis
Flow cytometry is often used for detection of various protein/antigens in blood samples, particularly for immunophenotyping.
Analysis of the results is usually much simpler and more accurate when the red blood cells are not included in the sample.
They need to be lysed.
Procedure
Red cells in whole blood samples for flow cytometry can be lysed using 0.1% Triton X-100 in PBS for 30 minutes (after brief
fixation of whole blood with 2% or 4% formaldehyde).
There are also several commercially available red blood cell lysing kits available (the manufacturer’s kit protocol should be
followed).
Research
Several detergents have been reported to affect detection of surface and internal antigens of blood cells. In the following
reference, the impact of different detergents and detergent concentrations on RBC lysis and on WBC resolution using light
scatter were investigated. Only three detergents, TX-100, NP-40, and Brij-58, showed significant RBC lysis after 2%
formaldehyde fixation for 10 min at room temperature. These detergents had a deleterious effect on WBC light scatter at
detergent concentrations above 0.2%, whereas concentrations below 0.1% resulted in incomplete RBC lysis. Thirty minutes
of incubation at room temperature with 0.1% detergent was optimal.
2005 International Society for Analytical Cytology Cytometry Part A 67!:4-17 2005
Whole Blood Fixation and Permeabilization Protocol with Red Blood Cell Lysis for Flow
Cytometry of Intracellular Phosphorylated Epitopes in Leukocyte Subpopulations.
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