PRODUCTION OF ANTIPNEUMOCOCCIC SERUM 553 



that concentration of antipneumococcic serum by water precipita- 

 tion might be developed into a practical method. Water subjects 

 the antibody to as little physical injury, perhaps, as any of the 

 processes employed. If modifications were made so that a high 

 yield of antibody free from reactive substances could be obtained, 

 the method would also possess the added advantage of being highly 

 economical. 



Carbon Dioxide : Avery reported that after diluting antipneumo- 

 coccic serum with one-half its volume of water saturation with car- 

 bon dioxide resulted in an incomplete precipitation of the anti- 

 bodies. Felton 396 repeated the experiment and as Avery had found, 

 noted that the yield in protective substance was never more, and 

 usually less than one-half the amount contained in the original se- 

 rum. The method of concentration with carbon dioxide has little 

 value from the practical standpoint. 



Sodium Chloride: Avery found that saturation of serum with 

 sodium chloride, as with carbon dioxide, resulted in a low yield of 

 antibody, since the immune substances were apparently associated 

 with both the euglobulin and pseudoglobulin separated by this 

 treatment. Banzhaf (1925) 70 corroborated the observation and 

 determined that the euglobulin so obtained contained only 8 to 10 

 per cent of the immune bodies. 



Alcohol Precipitation : From the sodium sulfate method, Felton 

 (1931) 408 turned to the use of ethyl alcohol as a precipitant in 

 concentrating antipneumococcic serum. Briefly, the technique rec- 

 ommended is as follows: Ethyl alcohol (95 per cent) and the 

 serum are cooled to 0°, and the alcohol is added to the serum in the 

 proportion of 20 cubic centimeters of alcohol to 100 cubic centi- 

 meters of serum. The mixture is allowed to stand for eighteen 

 hours in the cold, and the precipitate which forms is then recov- 

 ered either by centrifugation or filtration. The precipitate is 

 washed with one and one-half volumes of cold distilled water to re- 

 move the water-soluble protein. The insoluble fraction is dissolved 

 to the volume desired in isotonic salt solution. The so-called acid 



