Middlebrook albumin dextrose catalase enrichment adc

M. bovis BCG Pasteur 1173P2,^{47 (link)} live M. tuberculosis attenuated vaccine MTBVAC,^{7 (link)}M. tuberculosis clinical isolate MT103, its phoP-mutant derivative SO2 (kanamycin resistant) and the phoP-complemented SO2 strain (SO2:phoP)^{20 (link)} were used in this study. MT103 strain was rendered fluorescent by the transfer of plasmid pMV361H gfp (green fluorescent protein).^{48 (link)} Mycobacteria were grown at 37 °C in Middlebrook 7H9 broth (BD Biosciences, Madrid, Spain) supplemented with 0.05% Tween 80 and 10% Middlebrook albumin dextrose catalase enrichment (ADC; BD Biosciences), and when required the medium was supplemented with 20 μg/ml of kanamycin or hygromycin. The virulence and quality of our stocks are always confirmed in vivo in mice. All virulent stocks used to perform these experiments showed high virulence in vivo.

Mycobacterial strains used in this study were grown at 37°C in Middlebrook 7H9 broth (BD Difco) supplemented with 0.05% Tween 80 (Sigma) and 10% Middlebrook albumin dextrose catalase enrichment (ADC, BD Biosciences) or on solid Middlebrook 7H10 agar (BD Difco) supplemented with 10% ADC (BD Biosciences).
BCG Tice was obtained and cultured from a commercial vial of OncoTICE. BCG Pasteur 1173P2 and BCG::RD1 were a kind gift from Roland Brosch (Institut Pasteur, France). GFP-expressing MTBVAC and BCG Tice strains were generated in the laboratory by transformation with the pJKD6 plasmid (a kind gift from Luciana Leite, Butantan Institute, Brazil). The MTBVAC strain lacking ESAT6 and CFP10 (MTBVAC ΔE6C10) was constructed and characterized in our laboratory.12 (link)