technique to correct these difficulties and secondly to add more data
bearing on the specificity of the test. Soon after their publication Funk
became interested and coming to the same conclusions as to specificity
devised a centrifugating method for measuring the yeast growth. Williams
also improved his original method and devised a gravimetric method for the
same purpose. From the viewpoint of methodology we now have methods which
are suitable as quantitive procedures for determining the effect of
extracts of unknown substances on yeast growth and hence if the
stimulatory substance is vitamine "B," a means of determining within a
space of twenty-four hours the approximate content of stimulatory material
in a given source. Since the Funk method is the simplest of these and
illustrates the principles involved it will suffice to describe that.

_Funk method of yeast test with Eddy and Stevenson modification_

1. To a basal diet of 9 cc. of sterile culture medium such as a von Nageli
solution [Footnote: von Nageli's solution consists of the following
ingredients NH_4NO_3, 1 gram; Ca_3(PO_4)_2, 0.005 gram; MgSO_4, 0.25 gram
dextrose 10.0 grams made up to 100 cc. with distilled water. Other culture
media may be used and such combinations will be found in any text on
yeasts. They all permit a certain amount of growth but all are apparently
stimulated by the addition of vitamine extracts.] in a sterile test tube
is added 1 cc. of the sterile, neutral, watery extract of the source of
the vitamine. A pure culture of Fleischman's yeast (Funk prefers brewer's
yeast) is maintained on an agar slant and twenty-four hours before the
test is to be made, a transplant is made to a fresh agar slant. One
standardized platinum loopful of the twenty-four hour yeast growth is then
used to inoculate the contents of the tube, the tube stoppered with cotton
and incubated for from twenty-four to seventy-two hours at a temperature
of 31 C. The seventy-two hour incubation period yields nearly optimum