128 DIFCO MANUAL 



BACTO 



LOEFFLER BLOOD SERUM (B70) 



DEHYDRATED 



Beef Blood Serum 3 parts 



Dextrose Broth 1 part 



Bacto-Loeffler Blood Serum is employed in the cultural diagnosis of diphtheria. 

 The medium is of particular value in the determination of pigment production 

 by bacteria, and of the ability of anaerobes and other microorganisms to attack 

 protein, and for other special uses. Bacto-Loeffler Blood Serum has been prepared 

 in dehydrated form for the convenience of those laboratories where adequate 

 supplies of fresh blood serum are not available. 



Bacto-Loeffler Blood Serum is a modification of the horse serum, dextrose 

 broth medium described by LoefHer in 1887^ for the cultivation of Coryne- 

 bacterium diphtheriae. In our modern laboratories LoefHer Blood Serum still 

 retains its important position for the diagnosis of diphtheria. On Loeffler Blood 

 Serum C. diphtheriae grows luxuriantly and rapidly, developing morphologically 

 typical organisms, in 12-16 hours, aiding greatly in the diagnosis of diphtheria. 



Cleveland and Sanders,^ and Spector^ have used Bacto-Loeffler Blood Serum 

 in media for the cultivation of Endamoeha histolytica. Thompson* hydrolyzed 

 Bacto-Loeffler Blood Serum with sodium hydroxide and added this to a citrate 

 agar for the isolation of C. diphtheriae, on which diphtheria bacilli are stimu- 

 lated and other throat organisms are inhibited. 



To rehydrate the medium, dissolve 80 grams of Bacto-Loeffler Blood Serum in 

 1000 ml. of warm (42-45°C.) distilled water. 



During the dehydration process certain components of the medium become in- 

 soluble. The removal of this insoluble material by filtration on a Buchner funnel 

 with the use of filter aid or by centrifugation at high speed for 15-20 minutes 

 will result in better appearing final slants. The use of tubes that can be sealed 

 with a tight fitting closure such as a screw cap will produce slants free from 

 bubbles. It is recommended that the medium be distributed in tubes and coagu- 

 lated and sterilized in the autoclave as follows: 



1. Place not more than three rows of tubes in a slanting position, in wire 

 baskets, in the autoclave. 



2. Close all the ports and the door before turning on the steam to maintain 

 a mixture of air and steam for coagulation of the medium. If the exhaust of the 

 autoclave is equipped with an automatic thermo-element type trap, it is necessary 

 to have this line equipped with a valve which can be closed, thereby making it 

 possible to maintain a mixture of air and steam in the autoclave during coagula- 

 tion. 



3. Run the pressure as quickly as possible to 15 pounds and hold for 10 minutes 

 to coagulate the medium. 



4. At the end of coagulation, open the lowest port (or valve in thermo- 

 element type trap line if such is used) and replace the entrapped air with steam, 

 maintaining constant pressure while replacing the air-steam mixture with live 

 steam, as any appreciable change in pressure will cause the slants to be filled with 

 bubbles and render the medium useless. 



5. Maintain steam pressure for 15 minutes at 15 pounds (121°C.). 



6. After sterilization is completed, the source of steam is cut off. All ports and 

 the safety valve should be tightly closed as soon as the steam is shut off, allowing 

 the autoclave to cool slowly. Do not open any ports until the steam pressure 

 is nil. Final reaction of the medium will be pH 7.2. 



The autoclave sterilization methods of Hinkleman^ or Dupray® may be used 



