252 HILDEMANN, LINSCOTT AND MORLINO 



distress or disease following the injection. These animals were 

 also weighed regularly for a month after injection and all showed 

 normal weight gains. 



A more direct test for chimerism in A/Jax survivors of runt 

 disease was made by the cytotoxic isoantibody method devised 

 by Gorer and Boyse (1959). Briefly, potent A anti-Csy and C57 

 anti-A isoimmune sera were obtained following four intraperi- 

 toneal injections totalling about 300 milhon leucocytes (lymph 

 node, spleen, and bone marrow) per recipient. Each test tube 

 contained o-i ml. of antiserum dilution (1:4-1:64), o-i ml. of 

 I per cent lymph node cell suspension, and 0-05 ml. of guinea pig 

 complement previously absorbed with mouse cells at o°c to 

 remove naturally-occurring antibodies. Appropriate serum and 

 complement control tubes were included. After 60-90 minutes' 

 incubation at 30°, 0-05 ml. of i per cent Eosin-Y was added to 

 each tube; 200 or more cells from each test mixture were then 

 scored and the proportion of stained cells determined. AU dilu- 

 tions were made with Medium 199 (Hyland Laboratories), since 

 preliminary tests indicated that cell viability persisted for many 

 hours in this medium. A total of 14 A/Jax survivors of the runting 

 syndrome was tested by the above method. In all instances, a high 

 proportion of the lymph node cells was stained by Eosin-Y after 

 reaction with C57 anti-A serum, although there was no evidence 

 of cytotoxic reactions after exposure to A anti-Csy serum. 

 Typically, 60-80 per cent of the test cells were stained in the 

 presence of anti- A/Jax serum, and 10-15 ^^^ cent were stained 

 after incubation with anti-C57 serum. This low percentage of 

 stained cells in the latter tests was also observed in the serum and 

 complement control tubes. Some cell death is unavoidable as a 

 result of teasing the cells from the nodes. The potency of the A 

 anti-C57 serum was verified by its high cytotoxicity for C57BL/6 

 lymph node cells under identical conditions. These experiments 

 then clearly support the assumption that no substantial number of 

 C57BL/6 lymphoid cells persisted in A/Jax mice that recovered 



