106 



Journal of Applied Microscopy. 



Agar- Agar. The Preservation of 

 Culture Media. 



. In spite of the various methods pub- 

 lished from time to time for malcing 

 agar, it cannot be denied that it is still 

 a process dreaded by the average bac- 

 teriologist, and with good reason. In 

 almost all laboratories the tedious 

 methods taught in Europe are still 

 adhered to, the result being an infinite 

 amount of labor wasted, and often a 

 very poor quality of agar produced. 

 Three years ago, after several experi- 

 ments in an attempt to lessen the labor 

 of the process, I hit on the following 

 plan, which I have used and taught ever 

 since with the best results. The only 

 drawback to the plan is that it requires 

 an autoclave, but as this piece of appa- 

 ratus is now found in all well appointed 

 laboratories, it can hardly be considered 

 a great disadvantage. To make one liter 

 of agar-agar take 



A. Dried peptone (1%), 10 grams. 

 Common salt (.5%), 5 grams. 

 Liebig ext. (.5%), 5 grams. 

 Water, 500 cc. 



Boil for three minutes and neutral- 

 ize. 



B. Agar-agar (1.2%), 12 grams. 

 Water, 500 cc. 



Chop the agar and put into autoclave. 

 Run autoclave up to two atmospheres of 

 pressure, giving 121.4° C. of heat. As 

 soon as this pressure is reached, turn 

 out the flame, and allow the autoclave 

 to cool until below 100° C. before opening. 

 The two solutions A and B are then 

 mixed, cooled to 60° C, the whites of two 

 eggs beaten in 50 cc. of water added, 

 well stirred in, and the whole then boiled 

 and filtered through paper. 



The whole process requires only an 

 hour and a quarter to an hour and a 

 half, and the result is a most excellent 

 jelly. Instead of the white of egg, blood 

 serum may be used, which seems to add 

 also to the nutritive value of the medium. 

 Agar made with meat extract will often 

 form a precipitate during the steriliza- 

 tion, which is objectionable if one wishes 

 to use it in the pouring of petri dishes, 

 or the making of Esmarch's roll-tubes. 



To make an absolutely and perma- 

 nently clear agar, fresh meat should be 

 used as follows: 



To make one liter, take 



A. Chopped meat, 500 grams. 

 Water, 500 cc. 



Mix and place in cool place over 

 night, then strain through towel. 



B. Agar-agar (1.2%), 12 grams. 

 Water, 500cc. 



Put in autoclave, run up to two atmos- 

 pheres of pressure, put out flame, and 



allow to cool until below 100° C. before 

 opening. Let the solution of agar cool 

 still further to about 75° C, and then mix 

 A and B, add (1%) 10 grams dried peptone 

 and (.57c) 5 grams common salt, bring to 

 a boil for about three minutes, neutralize 

 and filter. The product is an absolutely 

 clear jelly, which never forms any pre- 

 cipitate. The whole process, with the 

 exception of the time the meat is 

 steeping, requires only about one hour 

 and a half. In both the above methods 

 of making agar, the filtration is very 

 quick — from ten to twelve minutes for 

 the liter. I never use a hot-water 

 funnel, but wet the filter paper with 

 boiling water immediately before pour- 

 ing in the agar. In the process with 

 fresh meat the clarification is effected 

 by the coagulation of the albumen in the 

 meat water, hence solution B must not 

 be added to A until cool enough to avoid 

 coagulation. In general the fresh meat 

 is to be recommended, and the process 

 is easier than with the meat extract, 

 though the latter has the advantage of 

 cheapness, and convenience, since the 

 meat extract can always be kept on 

 hand, and the time lost in soaking the 

 fresh meat is saved. 



The two processes above given have 

 been thoroughly tried by myself and 

 others, and can be confidently recom- 

 mended, as being the quickest and 

 simplest methods yet devised for the 

 making of this most necessary culture 

 medium, while the result is all that could 

 be desired. 



Before closing, let me advise a simple, 

 cheap, and satisfactory inethod of pre- 

 serving culture media, for which I am 

 indebted to Messrs. Kanthack and Drys- 

 dale. It is the use of tin-foil over the 

 mouth of the tube. It is applied as soon 

 as the tubes are filled, the cotton plug 

 being pushed in, and any excess of 

 cotton cut off. The tubes can then be 

 sterilized in the usual way, the tin-foil 

 undergoing sterilization at the same 

 time. The covering is not, of course, 

 absolutely air-tight, but very little evap- 

 oration can take place, and I have kept 

 media for seven months in perfectly 

 good condition by this method. If 

 desired the tubes can be put into a large 

 museum jar with perfect safety after 

 being capped in the above manner, and 

 preserved thus indefinitely. 



Mazyck p. Ravekel, M. D. 



Bacteriologist to the State Live Stock 

 ■ Sanitary Board of Pennsylvania; In- 

 structor in Bacteriology, Veterinary 

 Department, University of Pennsylvania. 



Please do not overlook the blank on 

 page vii. Secretaries of societies kindly 

 send full lists. 



