Use of anaerobic glove boxes for the cultivation of strictly anaerobic bacteria

Abstract

This paper discusses those features that are essential or at least highly desirable for the efficient and convenient operation of an anaerobic glove box. These include a flexible chamber with vacuum airlock, and a system for the continuous removal of oxygen from the chamber atmosphere, consisting of palladium catalyst in an atmosphere containing 10% H₂. Data reviewed and new data presented here indicate that agar plate media should contain palladium catalyst and cysteine, in order to permit the growth of strict anaerobes. A method is described that allows incorporation of palladium chloride catalyst into agar medium in a single step operation in the open laboratory. Data presented indicate that this medium was as efficient in the recovery of strictly anaerobic bacteria from the mouse cecum as a medium described earlier that contained palladium black and that required a more cumbersome method of preparation. The new type of medium may be stored for at least 1 month without anaerobic precautions prior to use in an anaerobic chamber, a feature that should permit inexpensive commercial production. It is concluded that the two principal features of the anaerobic glove box method are: 1) essentially standard bacteriologic techniques may be used to cultivate strict anaerobes, and 2) media may be prepared and stored in the conventional manner in the open laboratory. A glove box of proper dimensions may be designed to fit the special needs of any given laboratory. The entire process, from initial cultivation to final identification of strictly anaerobic isolates may be carried out in such a box using essentially the same procedures and requiring essentially the same degree of effort as one might expect to expend on the isolation of Salmonella from a stool specimen.