Improved detection of terminal transferase (TdT): the use of detergents on glutaraldehyde-fixed non-dehydrated cells prevents denaturation and diffusion artifacts.
TdT as an intranuclear enzyme mainly of immature lymphoid cells is commonly determined immunologically using air-dried cell smears fixed with methanol. Both cell dehydration and alcohol fixation were found here to denature TdT and surface antigens. This could be prevented by using non-dehydrated cells bound electrostatically to poly-L-lysine-coated slides, fixed minimally with glutaraldehyde and rendered permeable to antibodies by the non-ionic detergent Brij 56. Crosslinking glutaraldehyde in addition prevented diffusion of TdT to extranuclear sites. By avoiding artifacts of denaturation and diffusion, a higher sensitivity in the detection of TdT was achieved despite considerably lower quantities of antibody.
D002463:Cell Membrane Permeability; D004253:DNA Nucleotidylexotransferase / Q000032:analysis*; D004254:DNA Nucleotidyltransferases / Q000032:analysis*; D003902:Detergents / Q000494:pharmacology*; D004058:Diffusion; D005976:Glutaral; D006651:Histocytochemistry; D006801:Humans; D007938:Leukemia / Q000201:enzymology; D008722:Methods; D011489:Protein Denaturation; D013501:Surface-Active Agents / Q000494:pharmacology*; D013601:T-Lymphocytes / Q000201:enzymology
D003902:Detergents; D013501:Surface-Active Agents; D004254:DNA Nucleotidyltransferases; D004253:DNA Nucleotidylexotransferase; D005976:Glutaral