A simple and rapid immunological technique for visualising chromosome-mediated gene transfer (CMGT).
A method is described for visualising chromosome-mediated gene transfer (CMGT) by detecting chromosomes labelled with bromodeoxyuridine (BrdU) using a monoclonal antibody to BrdU. In this experiment, the CCRF-CEM T cell line was grown in the presence of BrdU and the labelled chromosomes were isolated and transfected into human embryonic fibroblasts. Uptake and retention of chromosomes were compared for transfection with either PEG or DMSO treatments. Following transfection the labelled chromosomes could be visualised in recipient cells using a monoclonal antibody to BrdU, followed by immunoperoxidase staining. Chromosome uptake into cells was similar for both DMSO and PEG treatments and was a relatively frequent event; about 1 in 5 recipient cells had labelled material present. This technique can be used to assess the technical aspects of the earliest stages of chromosome-mediated gene transfer.
Pittman
S
S
Kanematsu Laboratories, Royal Prince Alfred Hospital, Camperdown, NSW, Australia.
Warneford
S
S
Repka
E
E
Callaghan
T
T
Zbroja
R A
RA
Vincent
P C
PC
Young
G A
GA
D000911:Antibodies, Monoclonal / Q000276:immunology; D001973:Bromodeoxyuridine / Q000032:analysis* / Q000276:immunology; D002478:Cells, Cultured; D002875:Chromosomes / Q000032:analysis* / Q000276:immunology; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D005347:Fibroblasts / Q000648:ultrastructure; D006801:Humans; D007124:Immunoenzyme Techniques; D011092:Polyethylene Glycols / Q000494:pharmacology; D014170:Transformation, Genetic / Q000187:drug effects
D000911:Antibodies, Monoclonal; D011092:Polyethylene Glycols; D001973:Bromodeoxyuridine; D004121:Dimethyl Sulfoxide