INFLUENCE OF SAMPLE PREPARATION ON CELLULAR GLUTATHIONE RECOVERY FROMADHERENT CELLS IN CULTURE

During the last decade, the unbound glutathione content of cultured ad herent cells has become a very important biological marker for many ph armacological and toxicological in vitro studies with regard to the pr otective role of the tripeptide in its reduced form (GSH). However, th e literature does not provide extensive information on the influence o f sample preparation on cellular GSH and thiol analyses. Using the fib roblast-like V79 cell line as model, we undertook a comparative study of the efficiency of different procedures reported in the literature w ith respect to GSH recovery. Depending on the preanalytical step, up t o 10-fold discrepancies could be observed in the recovery of intracell ular GSH. Different parameters that must be controlled in order to max imize GSH recovery are discussed The optimal strategy consisted in rap id perchloric acid deproteinization performed directly in the dish, wh ich was extremely valuable for preparing GSH samples from adherent cel ls, and especially from cells expressing elevated gamma-glutamyl trans ferase activity.