IMMUNOLOGICAL STUDIES OF MICROSOMAL STRUCTURE AND FUNCTION 21 7 



with anti-microsomal serum. In the cell sap, the appearance of labelled 

 serum proteins is again slower. Nevertheless, after a long period of 

 incubation, the cell sap also contained a complete spectrum of strongly 

 labelled serum proteins. It should be emphasized that the concentration 

 of the serum proteins was practically the same in the cell saps of all four 

 experiments. Moreover, Fig. 3(</) shows, that even additional antigens in 

 the cell sap finally become considerably labelled, provided the period of 

 incubation with the isotope is sufficiently long. Thus, experiments of this 

 type suggest that the rate of isotope incorporation in the serum proteins 

 is faster than that of most other liver antigens reacting in our systems. 



When the pattern of incorporation of ^"^C amino acids into liver antigens 

 is studied after incubation of mitochondria-free homogenates, or of isolated 

 microsomal particles [24], a number of interesting differences are observed. 

 Even in these systems, a reasonable labelling is obtained, both of serum 

 protein-like antigens, and of other antigens extracted from the microsomal 

 fraction. However, only trace amounts of radioactive serum proteins can 

 be detected in the cell sap. Instead, this fraction, and, to a still higher 

 extent, the ribonucleoprotein particles, contain radioactive material which 

 precipitates in the agar around the basins, but without reacting with anti- 

 bodies. The nature of this material is unknown. The significance of these 

 findings will be discussed elsewhere [9]. 



Fig. 3. Autoradiographs of precipitin reactions of lubrol extracts (Lu) and 

 EDTA-extracts (EDTA) of microsomes, and of the cell sap (CS), of rat liver, with 

 various antisera (a-m : antiserum against liver microsomes ; a-cs : antiserum against 

 liver cell sap; a-pH 5: antiserum against cell sap fraction precipitated at pH 5; 

 a-rs: antiserum against rat serum). The antigen solutions were obtained from 

 four separately incubated (P*C]-L-leucine, L-isoleucine, and L-valine) and sub- 

 sequently fractionated aliquots of a preparation of rat liver slices. 



{a) 15 min. incubation with isotope. Time of exposure of autoradiograph : 

 3 months. The radioactivity of the TCA-precipitated protein of an aliquot of the 

 total homogenate was 135 c.p.m./mg. protein and that of an aliquot of the isolated 

 cell sap 68 c.p.m./mg. 



{b) 30 min. incubation with isotope. Time of exposure of autoradiograph: 6 

 weeks. Radioactivity: total homogenate 402 c.p.m./mg. protein; cell sap: 280 

 c.p.m./mg. protein. 



(f) 60 min. incubation with isotope. Time of exposure of autoradiograph: 6 

 weeks. Radioactivity: total homogenate 478 c.p.m./mg. protein; cell sap: 380 

 c.p.m./mg. protein. 



{d) 120 min. incubation with isotope. Time of exposure of autoradiograph: 

 3 weeks. Radioactivity: total homogenate 2190 c.p.m./mg. protein, cell sap 

 15 10 c.p.m./mg. protein. 



(Autoradiographs obtained after 6 months exposure of the plates corresponding 

 to the experiments of Figs. 2{a)-{c) did not show a greater number of labelled 

 precipitates.) The reactions obtained with the EDTA extracts and with a-pH 5 

 as well as other details are discussed elsewhere. From Morgan et al. [10]. 



