424 BACTERIOLOGICAL CHEMISTRY 



conditions, such as heating at 50° to 60° C, or solution in 

 urea or alkali, which cause the protein chains to unfold, 

 and the conditions then restored to normal in the presence 

 of an antigen, the polypeptide chains would refold in a 

 manner complementary to the antigen ; in other words 

 antibodies would be formed. He claims, on reasonable 

 but not rigid evidence, to have produced in vitro in this 

 way antibodies to 1 : 3-dihydroxybenzene-2 : 4 : 6-tri-jt)- 

 azophenyl arsonic acid, methyl blue and the specific 

 polysaccharide of Type III pneumococcus. 



The Separation of Antibodies. — Many attempts have 

 been made to separate antibodies from the inactive con- 

 stituents of the antiserum. These depend on (1) non- 

 specific methods and (2) specific methods : — 



1 (a) AUemtio7i of the Salt Content. — If serum is half- 

 saturated with ammonium sulphate the globulin is 

 precipitated, leaving albumin in solution ; the albumin 

 is precipitated on complete saturation of the serum with 

 ammonium sulphate. Globulin can be further split into 

 euglobulin which is insoluble in distilled water, and into 

 pseudoglobulin which is soluble in distilled water. Dialysis 

 of serum to remove all the salts, therefore causes the, 

 precipitation of euglobulin. Dilution of serum with 9 or 

 10 volumes of distilled water and slight acidification, for 

 example by passing carbon dioxide into the solution, 

 precipitates most of the euglobulin fraction and a small 

 proportion of the pseudoglobulin. Euglobulin, which 

 can also be precipitated by one-third saturation with 

 ammonium sulphate, comprises chiefly ^- and y-globulins 

 Avhich are the slower moving components in an electric 

 field (see p. 425). The pseudo -globulin is composed 

 mainly of the fast moving « -globulin fraction. It has been 

 found that the antibodies are almost entirely associated 

 with the glol3ulin fraction, but there is little agreement as 

 to the distribution between euglobidin and pseudo- 

 globulin, even when dealing with the same antibody. 



