Transfection and gene expression in normal and malignant primary B lymphocytes.
Although many different protocols for transfection of lymphoid cell lines exist, successful DNA transfer into primary B lymphocytes has, to date, not been demonstrated. We now describe a simple method for gene transfer into highly purified normal and malignant B lymphocytes by electroporation. Using a powerful expression vector containing two copies of the cytomegalovirus (CMV) immediate early enhancer linked to the human T cell lymphotropic virus I (HTLV I) promoter, we could demonstrate transfected gene expression in both high density small 'resting' B cells and in low density 'activated' B cells. Successful transfection was detected by expression of chloramphenicol acetyl transferase and by immunofluorescence. The neoplastic cells of B cell chronic lymphocytic leukemia could also be transfected with an efficiency of 5-10%, but only after preactivation. This method of transfection will permit analysis of the contribution of individual genes and their products to normal and malignant B cell growth and differentiation.
Ludwig Institute for Cancer Research, St. Mary's Branch, London, U.K.
M
M
Buschle
M K
MK
Brenner
I S
IS
Chen
H G
HG
Drexler
S M
SM
Gignac
C M
CM
Rooney
D000943:Antigens, Differentiation; D001402:B-Lymphocytes / Q000201:enzymology / Q000276:immunology; D018956:CD5 Antigens; D002469:Cell Separation; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D003587:Cytomegalovirus / Q000235:genetics; D004558:Electric Stimulation; D004742:Enhancer Elements, Genetic / Q000235:genetics; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D004854:Herpesvirus 4, Human / Q000235:genetics; D015368:Human T-lymphotropic virus 1 / Q000235:genetics; D006801:Humans; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000235:genetics / Q000276:immunology*; D008213:Lymphocyte Activation / Q000187:drug effects; D010957:Plasmids; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection; D014407:Tumor Cells, Cultured
D000943:Antigens, Differentiation; D018956:CD5 Antigens; D015500:Chloramphenicol O-Acetyltransferase; D013755:Tetradecanoylphorbol Acetate