338 B. NAKIC, A. KASTELAN AND N. AVDALOVIC 



the "white graft" phenomenon in the secondary hosts. Less than 

 I X 10^ spleen or lymph node cells is sufficient to give a positive 

 reaction. 



All the tolerant animals tested by the above method were 

 found to be chimeras. An interesting fact emerged from these 

 studies. Though the parabionts were found to be chimeras as 

 long as they were tolerant, some of the animals that succeeded in 

 rejecting the cross-graft after a period of tolerance of at least two 

 weeks, were also found to be chimeras irrespective of whether the 

 test graft transplanted at the time when the test for chimerism was 

 performed, was rejected by the first- or the second-set reaction. 



Although the above test is an excellent one for the qualitative 

 demonstration of foreign cells, we found it necessary to devise a 

 test that could render possible both the qualitative and the 

 quantitative demonstration of donor cells in any rat strain. The 

 test that could best fulfil these qualifications was obviously a 

 cytological one and the possibility of using sex chromosomes as 

 "markers" was provided by genetic studies of S. Ohno and 

 co-workers (Ohno and Kinosita, 1955; Ohno, Kaplan and 

 Kinosita, 1958; Ohno, Kaplan and Kinosita, 1959). By joining in 

 parabiosis partners of opposite sex it was possible by this method 

 to identify male cells in a tolerant female parabiont and vice versa. 



The only way in which sex can be decided in mitotic meta- 

 phases of rat somatic cells (Ohno and Trujillo, personal communi- 

 cation) is by photographing the cell, then cutting out the chromo- 

 somes, pairing them and aligning them serially. In female somatic 

 cells there will be an additional pair of chromosomes similar to 

 the second, third and fourth largest pair of autosomes; this is the 

 X pair. In male somatic cells, after pairing has been completed 

 there will be one left-over chromosome similar to the second, 

 third and fourth largest autosomes, which is the X-chromosome, 

 and another left-over similar in size to the smallest acrocentric 

 pairs, which is the Y-chromosome. Fig. i shows a photomicro- 

 graph and an ideogram of a male cell recovered from the bone 



