Improved protocol for colorimetric detection of complement-mediated cytotoxicity based on the measurement of cytoplasmic lactate dehydrogenase activity.
The conventional cytotoxicity detection protocols based on the lactate dehydrogenase (LDH) activity assays rely upon the quantitation of the enzyme activity released into the assay medium upon cell lysis. In the case of complement-mediated cytotoxicity this results in the need to take into account the high and variable background LDH activity from the serum added to the cells. Using primate-derived COS-7 and pig kidney PK15 cell lines we show that, in the case of adherent cells, it is possible to overcome this drawback and to measure cell death caused by complement attack, by quantifying the amount of LDH activity retained by the undamaged cells. The modified assay is therefore quicker to carry out than the conventional procedure and cheaper because fewer control readings must be taken.
Sepp
A
A
Department of Immunology, Royal Postgraduate Medical School, Hammersmith Hospital, London, UK.
Binns
R M
RM
Lechler
R I
RI
D000818:Animals; D002021:Buffers; D019556:COS Cells; D002452:Cell Count; D003124:Colorimetry; D003165:Complement System Proteins / Q000502:physiology*; D003593:Cytoplasm / Q000201:enzymology* / Q000276:immunology; D003601:Cytotoxicity Tests, Immunologic / Q000379:methods*; D004789:Enzyme Activation / Q000276:immunology; D000882:Haplorhini; D007668:Kidney / Q000166:cytology; D007770:L-Lactate Dehydrogenase / Q000276:immunology* / Q000378:metabolism; D013552:Swine
D002021:Buffers; D003165:Complement System Proteins; D007770:L-Lactate Dehydrogenase