[Reprinted from The Medical News, December 10, 1892.] 
A NEW METHOD FOR THE CULTURE OF DIPH- 
THERIA-BACILLI IN HARD-BOILED EGGS.1 
By WYATT JOHNSTON, M.D., 
OF MONTREAL?" 
All who have had experience in the diagnosis of 
diphtheria by culture-methods agree in praising 
their accuracy and promptitude. Unfortunately, 
the general practitioner, who must feel most of all 
the need of some accurate method for the prompt 
diagnosis of doubtful cases, does not seem dis- 
posed to avail himself of the new process, and the 
prophecy of Roux and Yersin, that the method 
would come into general use, appears still to be far 
from fulfilment. 
Thinking that the chief obstacle lay in the dif- 
ficulty of obtaining serum for the culture-medium, 
M. Sakharof2 recently suggested a simple plan by 
which slices of hard-boiled eggs, cut with a steril- 
ized knife and placed in sterilized tubes, could be 
made to replace the serum. 
Of this method I have no personal experience, 
but should imagine that the main objection would 
still exist, as the physician might not have test-tubes 
about him at the time when they were most needed. 
1 From the Pathological Laboratory of McGill College, 
2 Annales Inst. Pasteur, June, 1892. 2 
I have, during the past two months, made use of 
a method which may be regarded as a modification 
of Sakharof's, and which does away with the 
necessity both of test-tubes and the preparation of 
media before they are actually needed for use. 
I employ hard-boiled eggs, from which a part of 
the shell is removed with ordinary forceps, after 
being tapped so as to break it. In this way shell 
and shell-membrane can readily be peeled off from 
one extremity (by selecting the narrow extremity 
the air-chamber is avoided), leaving a smooth, 
glistening, moist surface, which offers a most tempt- 
ing spot for making cultures. These are made, as 
in the case of serum, by touching the diphtheritic 
exudation with a sterilized needle and drawing the 
latter lightly from three to six times across the ex- 
posed white of the egg. Instead of the regulation 
platinum needle mounted in a glass rod, I employ 
either an ordinary needle or a bit of silver suture- 
wire held in an artery forceps. To guard the cul- 
ture against contamination the egg has only to be 
placed upside down in a common egg-cup; it can 
afterward be wrapped in paper and transported, if 
necessary. The interior of the cup can be sterilized, 
if desired, by allowing a flame to enter it for a 
second or two, though I have not found this neces- 
sary, as the nutrient surface does not come in contact 
with the inside of the cup. The egg and shell are, 
of course, both sterilized by the act of boiling. 
Five minutes' boiling suffices, and if the opera- 
tion has to be done "while you wait," the egg can 
be cooled in a still shorter time by placing it in 
cold water. Strict attention to aseptic details is 3 
unnecessary, as the diphtheria-bacillus outstrips in 
its growth the contaminating organisms likely to 
lead to confusion. The appearance of the diph- 
theria-colonies at the expiration of twenty-four 
hours is the same as when they are grown in serum, 
but I have found the growth even more rapid, so 
that a colony is already visible in twelve hours. 
Confusion with micrococci is, of course, to be 
guarded against. The reliability of this method 
seems to be the same as that of the methods of 
Haffter and E. Roux. I have found one bacillus 
which attains visible dimensions within the same 
period, but as this also grew on blood-serum in the 
manner characteristic of the diphtheria-bacillus, 
the great value of the method here described is not 
invalidated by that fact. 
Although this minor modification of a now well- 
tried procedure might enable it to be employed by 
those destitute of laboratory outfits, I do not think 
it likely that this means of diagnosis will be utilized 
by physicians not habituated to laboratory methods. 
It may be of interest to state here that the con- 
stant temperature of about 350 C., needful to insure 
the rapid and characteristic growth of the diphtheria- 
bacillus, can readily be obtained by placing in a 
cupboard or box with the culture, a large jar or 
pail of warm water, which is renewed from time to 
time, thus making an impromptu thermostat.