A novel B cell stimulation/proliferation assay using simultaneous flow cytometric detection of cell surface markers and DNA content.
Assessment of the specific B cell proliferative response by thymidine incorporation following polyclonal stimulation (e.g. with pokeweed mitogen or B cell growth factor) is problematic whenever the proportion of B cells in a particular sample is low as in blood samples. Here we describe a simple assay consisting of identifying the cultured B cells with anti-CD19/CD20 fluorescein-conjugated antibody and simultaneously using propidium iodide DNA staining to determine the percent of S/G2/M phase cells among the gated B cells. This assay can be used to determine B cell responsiveness even in individuals with very low blood B cell counts (e.g., in recipients of bone marrow transplant), does not require laborious B cell purifications and does not involve radiation hazard.
Storek
J
J
Hart and Louise Lyon Laboratory, Department of Medicine, UCLA School of Medicine.
Schmid
I
I
Ferrara
S
S
Saxon
A
A
D000328:Adult; D000944:Antigens, Differentiation, B-Lymphocyte / Q000032:analysis; D001402:B-Lymphocytes / Q000166:cytology / Q000276:immunology*; D002453:Cell Cycle; D004247:DNA / Q000032:analysis; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D007075:Immunoglobulin M / Q000032:analysis; D008213:Lymphocyte Activation; D008875:Middle Aged; D011947:Receptors, Antigen, B-Cell / Q000032:analysis; D013936:Thymidine / Q000378:metabolism
D000944:Antigens, Differentiation, B-Lymphocyte; D007075:Immunoglobulin M; D011947:Receptors, Antigen, B-Cell; D004247:DNA; D013936:Thymidine