838 IMMUNOGENETICS 



macroscopic reading is performed bv shaking the pellet of sedimented cells gently back 

 into suspension, meanwhile checking for agglutination by examining the suspension for 

 clumps against a light background, for these and similar tests, a special small centri- 

 fuge called the Sero-fuge (Clay-Adams, Inc., N.Y.) provides a significant saving in 

 time. 



Variants of this test system are numerous. For example, for particular reagents 

 the cellular suspensions may need to be heavier than 2 per cent; the temperature may 

 be optimally either higher or lower than room temperature; a slanting capillary tube 

 technique, with examination for the sedimentation pattern on the side of the capillary 

 tube, is a sensitive technique for some systems and conserves valuable test sera. For 

 some tests, such as Rh tests with particular reagents, slide agglutination is used. Direc- 

 tions for special techniques are provided with commercial reagents if these are to be 

 purchased. 



2. Other systems for agglutination. Antisera that fail to agglutinate red cells in 

 saline, or that do so only weakly or unreliably, may produce strong and reliable 

 agglutination if a medium other than saline is employed, or if the cells are treated in 

 particular ways. One of the most common modifications is to use 20 per cent bovine 

 albumin as a diluting fluid for the test serum and as a medium for the suspension for 

 test red cells. Frequently, a somewhat heavier red-cell suspension is used in such 

 systems — often of the order of 4 per cent. These tests are often incubated for an 

 interval of two hours or so, at 37° C, before being read. Normal serum is sometime 

 used as a diluting fluid ; if the normal serum contains antibody, this can be absorbed 

 out with washed red cells before it is used. Dextran, gelatin, and polyvinylpyrrolidone 

 are also sometimes used. 380, 665, 829 



Murine antibodies generally produce only irregular and uncertain agglutination 

 of murine red blood cells in saline media. A major methodologic contribution, which 

 made it possible to bring the mouse into productive use for investigations of red-cell 

 immunogenetics, was the development by Gorer and Mikulska 459 of the Dextran 

 method lor hemagglutination. In this method, the test red cells are suspended in a 

 1/2 dilution with saline of normal human serum, which has been absorbed with washed, 

 pooled, murine red blood cells in order to remove any normal, human-anti-mouse red- 

 cell antibody. We ordinarily make up the test cell suspension in saline to proper 

 concentration and just in the amount needed, then centrifuge down the cells, aspirate 

 off the supernatant saline, and replace it with the absorbed and diluted human serum. 

 The test reagents, mouse-anti-mouse antisera, are diluted with a saline dilution of 

 Dextran, the latter at a concentration of 2 percent. Unfortunately, different Dextran 

 preparations vary in their desirability for this purpose; some of them agglutinate cells 

 themselves without the addition of antibody in nonspecific fashion, while others fail to 

 promote the agglutination of antibody-sensitized cells. The Dextran of choice is 

 Tntradex, produced by Glaxo Laboratories, Ltd. (Greenford, Middlesex, England) but 

 this is difficult to obtain. 



After adding a drop oi' the normal, serum-suspended, test cells to a drop of the 



