An Apparatus for Grinding Bacteria at Low Temperatures

Abstract

The disrupting of bacteria by grinding in mills is a well established laboratory procedure, for which various forms of apparatus have been devised. The long grinding required to break most of the bacteria introduced into an ordinary mill changes the material so that some of the immunologically detectable components are no longer present, or at least not identifiable. In order to take advantage of the brittleness of cells at low temperature, Macfadyen and Rowland 1 used a grinding apparatus that was immersed in liquid air. Johlin and Avery 2 constructed a heavy steel cylinder in which a moist bacterial suspension was frozen with CO₂ ice; a tightly fitting plunger was introduced and struck 25 times with an 8 lb. hammer. These authors presented data indicating that this was a very efficient method of rupturing bacteria and obtaining the intracellular contents. For extracting tissues, a somewhat similar apparatus devised by Graeser, Ginsberg, and Friedemann 3 was used by Fox 4 to study the concentration of antibodies in tissues.

Recently Mudd, Shaw, Czarnetsky and Flosdorf 5 have devised an apparatus for disrupting bacteria at low temperatures after desiccation by the lyophil process. The grinding chamber consists of a hollow monel metal disc, milled out in such a manner that 2 large stainless steel balls 1½ inches in diameter fit accurately into the hollow. The bacteria are placed in this mill, frozen and dried by attaching it, with airtight connections, to a condenser which is immersed in a low-temperature freezing bath. Following the drying, the mill is rotated in a similar bath of CO₂ ice and methyl cellosolve. At least 80% of the bacteria are completely disintegrated within a short time.