A rapid sample preparation technique for flow cytometric analysis of immunofluorescence allowing absolute enumeration of cell subpopulations.

Journal of Immunological Methods
L W TerstappenM R Loken

Abstract

A simple and rapid method was developed for immunofluorescence measurements of cells by flow cytometry which does not require washing procedures, permitting absolute enumeration of cell subpopulations. Peripheral blood cells were labeled with fluorescein and phycoerythrin conjugated monoclonal antibodies and the nucleic acid stain LDS-751. Distilled water was added following incubation to induce erythrocyte lysis by hypotonic shock. After lysis for 30 s the tonicity of the sample was increased followed by measurement on the flow cytometer. The leukocyte populations were clearly resolved in the correlation of forward and orthogonal light scattering. The immunofluorescence resolution of the labeled leukocytes was equivalent to NH4Cl and a commercial lysing preparation. Absolute number of leukocytes and percentage of leukocyte subpopulations determined with this procedure correlated well with the results obtained with a clinical hematology analyzer. Cell recovery and preservation of cellular characteristics of three different procedures for lysing the human erythrocytes were compared. The LDS-751 permitted the discrimination of intact cells from residual erythrocyte ghosts, platelets and damaged nucleated cells. A considerable los...Continue Reading

References

Mar 1, 1981·The Journal of Experimental Medicine·T TimonenR B Herberman
Aug 1, 1980·Proceedings of the National Academy of Sciences of the United States of America·R A HoffmanG Goldstein

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Citations

Nov 25, 1992·Journal of Immunological Methods·S Stemme, B Kallberg
Nov 1, 2001·Journal of Immunological Methods·D S Snyder, P L Small
Sep 7, 2002·Journal of Immunological Methods·Yan-Yi Wang, Xiao-Xiang Zheng
Mar 20, 1993·Annals of the New York Academy of Sciences·G T StelzerM R Loken
Feb 18, 2011·Applied Physiology, Nutrition, and Metabolism = Physiologie Appliquée, Nutrition Et Métabolisme·Michael De LisioGianni Parise
Jun 6, 2007·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Iva SlaninováEva Táborská
Dec 7, 2007·Cytometry. Part B, Clinical Cytometry·Sven BjörnssonPer Simonsson
Oct 9, 2007·Developmental Biology·Irina KirillovaZipora Yablonka-Reuveni
Nov 23, 2016·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Curt I CivinJames C Sturm
Oct 1, 2005·Arteriosclerosis, Thrombosis, and Vascular Biology·Elvire HenrichotChristoph A Meier
Jul 11, 2008·Medicine and Science in Sports and Exercise·Yuko TanimuraRyuichi Ajisaka
May 17, 2005·European Biophysics Journal : EBJ·Stefan Trapp, Richard W Horobin
Mar 25, 2005·Biological Research for Nursing·Leslie S RitterPaul F McDonagh
Sep 5, 2008·Current Protocols in Cytometry·Paul J SmithRachel J Errington
Feb 9, 2019·Scientific Reports·Vaskar GnyawaliMichael C Kolios
Oct 22, 2019·European Journal of Immunology·Andrea CossarizzaArturo Zychlinsky
Nov 5, 1998·Clinical and Diagnostic Laboratory Immunology·P SchlenkeG Bein

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