Separation of tuberculin-active proteins from the infected mouse lung tissue.

Mice were infected intravenously with 0.5 mg of a virulent strain of M, bovis. The lungs were separated from the moribund mice 3 to 4 weeks after infection. The lungs from 640 mice were homogenized and the bacilli removed therefrom by sequential centrifugation. The supernatant was heated at 120 C for 20 min and subjected to filtration and centrifugation to remove the denatured and aggregated materials. The supernatant was then fractionated with ammonium sulfate at 50 %-saturation, by gel-filtration on a Sephadex G-100 column and by chromatography on a DEAE-cellulose column, The protein fraction finally obtained had a high tuberculin activity and behaved like PPD in disc electrophoresis in polyacrylamide gel.