Growth of B cell colonies independent of T cell contact.
One of the methods of obtaining B cell colonies involves physically admixing B cells in growth medium containing mitogens with accessory T cells treated with mitomycin C or low dose irradiation to prevent T cell colony formation. However, under these conditions T cells still have the capacity to form colonies in methylcellulose, which is most frequently used for colony assays, and B cells recovered from the colonies are contaminated by the large number of added T cells. Therefore, we have developed a method of growing B cell colonies by physically separating the enriched B cells from the T cells by using millicell-HA inserts (Millipore). The requirement for mitogens, culture conditions, development, and number of B cell colonies formed were similar to those observed with admixture of T and B cells. The advantage of this system is that it permits growth of B cell colonies without the confounding influence of T cell proliferation or treatment of T cells with mitomycin or irradiation, and relatively pure B cell colonies can be recovered for further characterization or functional studies.
Fernandez
L A
LA
Department of Medicine, Camp Hill Hospital, Halifax, Nova Scotia, Canada.
MacSween
J M
JM
Robson
D A
DA
D000944:Antigens, Differentiation, B-Lymphocyte / Q000032:analysis; D001402:B-Lymphocytes / Q000166:cytology* / Q000187:drug effects; D002469:Cell Separation; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay / Q000379:methods*; D046508:Culture Techniques / Q000379:methods; D006801:Humans; D008213:Lymphocyte Activation / Q000187:drug effects; D008222:Lymphokines / Q000378:metabolism / Q000494:pharmacology; D013205:Staphylococcal Protein A / Q000494:pharmacology; D013601:T-Lymphocytes / Q000378:metabolism
D000944:Antigens, Differentiation, B-Lymphocyte; D008222:Lymphokines; D013205:Staphylococcal Protein A