Abstract
This paper contains a report of a rapid and simple method for removing bacteria from broth cultures. Large volumes can be handled conveniently. Preliminary tests indicate that the antigenic properties of the bacteria remain unaffected.
The method consists in the formation of a precipitate of calcium phosphate in the broth. This adsorbs the bacteria, settles quickly, and can be removed by siphoning away the supernatant liquid and lightly centrifuging the remaining sediment. The calcium phosphate may then be redissolved by suspending the precipitate in water and acidifying to about pH 5.0. The bacteria can be recovered by centrifugation and then washed.
The amounts of bacterial culture employed are limited only by the available apparatus. A preliminary test is made of the amount of calcium phosphate necessary to effect practically complete removal of the microörganisms. This may be done by preparing a series of tubes containing 10 cc. of broth to which is added 0.1-1.0 cc. of the calcium reagent, followed by N NaOH, to pH 7.0. After light centrifugation, the supernatant liquid will be crystal-clear in the tubes containing enough calcium. In the routine treatment of any one type of culture, a single test should suffice.
We conduct the precipitation of 5-liter quantities of culture by adding half the required amount of 0.5 M acid calcium phosphate,∗ and half the amount of N NaOH that will be required to bring the pH to 6.5-7.0 (Nitrazine paper). The rest of the calcium phosphate and NaOH are then added. This is probably more efficient than adding (the whole amount at one time, and helps to prevent large clumps of precipitate from forming. For the same reason, the mixture should be agitated vigorously during the precipitation.
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