Counting efficacies of the CFU-enumerating method and microscopic counting method for mycobacteria located in cultured macrophages
Sato, K.
Kekkaku 67(10): 663-665
1992
ISSN/ISBN: 0022-9776 PMID: 1453568 Document Number: 391237
I compared counting efficacies of CFU-enumerating method for the number of mycobacteria (Mycobacterium intracellulare and M. fortuitum) locating in cultured macrophages with that of microscopic counting method. Zymosan A-induced macrophages from ddY mice were infected with either M. intracellulare or M. fortuitum by incubation in 10% FBS-RPMI 1640 medium containing the organisms for 1 hr, thereafter thoroughly washed to remove extracellular bacilli, and cultured for 3 to 5 days. At intervals, macrophages were thoroughly rinsed and subjected to either CFU-enumeration or microscopic counting, as follows. In the former method, macrophages were lysed with 0.2% Tween 80-distilled water by sonication using Handy Sonic and CFUs were counted on 7H11 agar plates. In the latter method, the number of acid-fast bacilli was counted by microscopy for macrophages after Ziehl-Toda's staining. The number of bacteria by the CFU-enumerating method was much greater than that by the microscopic counting method.