INGREDIENTS OF CULTURE MEDIA 281 



Coombs test. The prozones commonly experienced in titrations using untreated 

 cells in albumin solutions, were completely eliminated when trypsin-modified 

 cells were employed. Wheeler^o found Trypsin, Dif co, 1 : 250 and Bacto-Trypsin 

 1% to be eminently satisfactory for use in treating erythrocytes employed in Rh 

 determinations. The latter preparation was preferable because of its greater 

 purity, ready solubility and ease of solution. Wiener and Katz^^ in their 

 studies on the use of enzyme-treated red cells in tests for Rh sensitization found 

 that the use of trypsinated red cells was a sensitive and specific method for testing 

 Rh-Hr antibodies and antibodies of other specificities. Enzyme-treated cells gave 

 more sensitive results in tests for Rh sensitization than other standard tests 

 hitherto introduced and equalled them in specificity. The titers obtained using 

 trypsinated erythrocytes were four or five times as high as by the albumin plasma 

 conglutination technique. In no case of Rh sensitization did the test with enzyme- 

 treated cells fail to demonstrate the presence of Rh antibodies although in several 

 of these cases the other tests were negative. Furthermore, the trypsinated cell 

 technique was observed to be a reliable method for demonstrating autosensitiza- 

 tion even in the absence of clinical manifestations. These authors used Trypsin, 

 Dif CO, 1 : 250 and noted no damage to any of the hemagglutinogen loci by this 

 enzyme. 



Unger, Tortora and Mappi^^ jn their isoimmunization studies in rabbits used 

 Trypsin, Difco, 1:250 for conditioning the erythrocytes and observed that the 

 trypsinated cell method was more sensitive than either the saline agglutination 

 or albumin plasma methods for the detection of isoantibodies. Rosenfield and 

 Vogel^^ studied the effect of trypsinated erythrocytes on the identification of 

 hemagglutinins. They confirmed Wheeler's observations that the prozone phenom- 

 enon obtained with certain high titer sera in albumin is not observed with 

 optimally trypsin-treated erythrocytes. They found trypsinated cells to be sensitive 

 to agglutination by extremely small amounts of Rh antibody. Like Wiener and 

 Katz^^ these authors did not encounter an instance of sensitization to Rh-Hr that 

 was not detectable by this technique. This fact makes the enzyme technique just 

 as valuable in routine tests for Rh sensitization as the indirect antiglobulin test. 

 Rosenfield and Vogel also observed that trypsin treatment of the cells influenced 

 the A and B factors so that agglutination was of better avidity and the titration 

 values were higher than in saline. The O factor was not changed. The M and N 

 factors were largely destroyed by enzyme activity. The S factor was enhanced by 

 trypsin treatment. The Fy^ (Duffy) factor was destroyed by enzyme treatment. 

 Trypsin improved the Le^ and Le^ agglutination. Trypsin was also found to 

 accentuate hemolysin activity upon the red cells treated with this enzyme. 

 Trypsin enhanced agglutination with Anti-P sera. Agglutinogen for cold auto- 

 agglutinins was altered by enzyme treatment resulting in higher titration values 

 for cold agglutinins in general and a wider thermal amplitude. They recom- 

 mended careful standardization of the technique of treating cells with trypsin so 

 as to avoid false positive reactions. Rosenthal, Damelshak and Brukhardt^* used 

 Trypsin, Difco 1 : 250 in treating red blood cells in their study of cells in acquired 

 hemolytic anemia. Morris^^ used Trypsin, Difco 1 : 250 in his study of hemag- 

 glutination by murine encephalomyelitis virus. He reported that virus which 

 failed to agglutinate human group O erythrocytes at 20° C. did so in the presence 

 of trypsin. Bacto-Trypsin 1% is ideally suited for these determinations. 



Bacto-Trypsin 1% for Hemagglutination is reconstituted by adding 10 ml. 

 Bacto-Hemagglutination Buffer to each 10 ml. vial. The resultant solution is 

 sparklingly clear and equivalent in tryptic activity to a 1 per cent solution of 

 Trypsin, Difco, 1 :250. The reaction of the final solution will be pH 7.2. The re- 

 constituted solution should be kept refrigerated at 2-6° G. or colder until used. 

 Bacto-Trypsin 1% for Hemagglutination when reconstituted is used according to 



