108 DIFCO MANUAL 



Bacto-Dubos Oleic Agar Base is recommended for the preparation of a solid 

 medium for the isolation of Mycobacterium tuberculosis. It is prepared accord- 

 ing to the formula described by Dubos and Middlebrook.^ Bacto-Dubos Oleic 

 Agar Base enriched with Bacto-Dubos Oleic Albumin Complex may also be 

 used as a basal medium on which to test the sensitivity of M. tuberculosis to 

 chemotherapeutic agents. 



Dubos and Middlebrook in a discussion of media for the cultivation of tubercle 

 bacilli described an agar medium suitable for diagnostic primary isolations and 

 also for studies of colony morphology. This medium is prepared without glycerol 

 or dextrose in order to discourage the growth of commensal organisms that 

 might not have been killed during the concentration process. Asparagine like- 

 wise is added in minimal quantities to provide maximum selectivity. The basal 

 agar is enriched with Bacto-Dubos Oleic Albumin Complex, (page 275). The 

 sterile base is cooled to 50-55 °C., and oleic acid albumin complex and penicillin 

 as selective agent are added. The medium is mixed and distributed in sterile 

 tubes or plates as desired. About 15 ml. of medium is used per 95 mm. dish 

 and following solidification two drops of an adequately diluted suspension of the 

 tubercle bacilli or of pathological material are used as an inoculum. Roberts, 

 Wallace and Erlich^ in a study of methods of isolation of the tubercle bacilli re- 

 ported that Dubos Oleic Albumin Agar Medium gave better results than other 

 media, even superior to guinea pig inoculations. Byham,^ comparing different 

 media for primary isolation work, reported best results with Middlebrook's 

 medium.* Commensal organism contamination was reduced by the penicillin and 

 early growth of the tubercle bacilli was obtained. Foley,^ in a limited survey, 

 too small to permit analysis reported that four strains grew on the solid Dubos 

 medium, but failed to develop in the liquid Dubos medium. He pointed out that 

 colonial morphology is strikingly different on the Dubos Agar than on coagu- 

 lated egg media. 



To rehydrate the medium suspend 4.2 grams Bacto-Dubos Oleic Agar Base 

 in 180 ml. of cold distilled water. Heat to boiling to dissolve the medium com- 

 pletely. Sterilize in the autoclave for 15 minutes at 15 pounds pressure (121°C.). 

 Cool to 50-55 °C. and add the contents of one tube of Bacto-Dubos Oleic 

 Albumin Complex (20 ml.) and 5,000 to 10,000 units of penicillin (25 to 50 units 

 per ml.), under aseptic conditions. Mix thoroughly and distribute in sterile tubes 

 or plates as desired. Keep uninoculated medium in the refrigerator. 



One pound of Bacto-Dubos Oleic Agar Base will make 21.6 liters of medium. 



2 Am. Rev. Tuberculosis, 61:563:1950. 27:28:1947. 



3 Am. J. Clin. Path., 20:678:1950. ^ J. Lab. Clin. Med., 32:842:1947. 



^ Am. Rev. Tuberculosis, 56:334:1942. * Proc. N.Y, State Assoc, of Pub. Health Lab., 



61:563:1950. 

 1950. 



BACTO 



TB BROTH BASE (B291) 



DEHYDRATED 



Bacto-Yeast Extract 2 g. 



Proteose Peptone No. 3, Difco 2 g. 



Bacto-Casitone 2 g. 



Disodium Phosphate 2.5 g. 



Monopotassium Phosphate 1 g. 



Sodium Citrate 1.5 g. 



Magnesium Sulfate 0.6 g. 



Tween 80 0.5 g. 



Bacto-TB Broth Base is a modification of the medium described by Dubos 

 and Davis^ and is recommended for the preparation of a liquid medium for 



