Elimination of mycoplasmas from cultured human cell lines utilizing a combination of two antibiotics, quinolone and a pleuromutilin derivative

Forty three cultured human cell lines were treated with a combination of 2 antibiotics to eliminate contaminant mycoplasmas. One course of treatment was composed of consecutive 3 or 4 cycles. Each cycle grew cells in BM-1 (pleuromutilin derivative; Boehringer Mannheim) containing medium (10 micrograms BM-1/ml culture) for 3 days, alternating with MC-210 (quinolone; Dainihon Pharmaceutical) containing medium (0.625 micrograms MC-210/ml culture) for 4 days. No treatment failure was encountered with this procedure. Before treatments, 18 (90%) of 20 cell line samples were contaminated with mycoplasma, as tested by DNA hybridization method (MYCOPLASMA T.C. RAPID DETECTION SYSTEM; Gen-Probe Inc.). Out of 43 cell lines treated, 7 were reduced in growth and dropped out. Among the other 36 cell lines, 27 became negative, 5 borderline and 4 slightly positive to the mycoplasma detection. All of the latter 9 cell lines, treated with one more similar course, found to be free from mycoplasma. Six of the dropout lines were cured of mycoplasma by a second treatment, under modified culture conditions. The last cell line (NATO) was successfully treated with another lot of FCS. Thus, the procedure proved successful even in treating promiscuously infected cell lines.