A New Method for the Isolation of M. tuberculosis

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Abstract
This method is superior to the 2 most commonly employed to date, primarily by elimination of contaminants. For outpatients, 3-5 ml of 1:1000 Zephiran (Winthrop-Stearns, Inc., a mixture of alkyldimethyl-benzyl-ammonium chlorides), prepared with distilled water, should be added to 50-ml screw-capped bottles and sent to outpatients for collection of sputum specimens. For hospitalized patients'' sputums, gastrics, urine or feces and the returned outpatient sputum specimens, a digestant consisting of 10% trisodium phosphate (23% Na3PO4.12H2D) in a 1:1000 solution of Zephiran (prepared with distilled water) may be used. Five to ten ml of specimen are placed in a 50-ml glass-stoppered centrifuge tube with 10 to 15 ml of digestant, and allowed to digest 15 minutes at room temperature. Sterile water is added and the tubes centrifuged at 3000 rpm for 30 minutes. The supernatant is poured off, the sediment resuspended in sterile distilled water, and recentrifuged. The sediment obtained may now be inoculated on Petragnani''s medium (Frobisher''s modification) and slides made to be stained by the Ziehl-Neilsen''s technique. Media other than Petragnani''s or Bacto-Peizer TB medium should be used with caution, and NaOH should never be used with Zephiran.