VIROLOGICAL METHODS 273 



cumstances a fourfold or more rise in antibody titer in the convalescent 

 serum is considered significant. This conclusion assumes that the test 

 was conducted with precision and with adequate controls. 



The antihemagglutination test is also used in the antigenic analysis 

 of antigenically related viruses. The principle is not too different from 

 cross agglutination among the bacteria. There is an important differ- 

 ence, however. In viral hemagglutination, using the proper menstruum, 

 the antigen-antibody combination interferes with a result that would 

 otherwise occur. On the surface this may seem inconsequential, but it 

 could obviate the very quantitative end hoped for. If antigenically 

 related viral particles A and B possess hemagglutinin activity X and 

 2X, respectively, it may take twice as many viral A particles to equal 

 a hemagglutinin dose as those of virus B. If the antibody-binding 

 ability of hemagglutinin groups are disproportionate in the opposite 

 direction or not directly proportionate in the same direction, direct 

 comparison of the two viruses from an antigenic standpoint as elicited 

 by the antihemagglutination test would be in error. 



If the antibody functioned indirectly, i.e., not acting against the 

 hemagglutinin but against other portions of the virus particle and thus 

 preventing hemagglutination by not allowing the hemagglutinin to seat 

 properly with the surface of the red cell, antigenic comparisons by this 

 test could be in even greater error. Since some virus particles are 

 known to fix firmly to red cells in the presence of considerably greater 

 quantity of antibody than is necessary to prevent hemagglutination, the 

 above hypothesis may not be so unlikely as might be presumed (Hul tin 

 and McKee, 1951). The fact remains that with three major reacting 

 entities, virus, red cell, and antibody, the direct conclusions drawn from 

 a two-way system, antigen-antibody reaction, should be viewed with 

 caution. 



To discover qualitative antigenic differences among related viruses 

 antibody adsorption much as is employed in bacteriology may be used. 

 Advantage being taken of the fact that adequate centrifugal force will 

 sediment the majority of viral particles from a given viral preparation 

 and that a number of different viruses apparently form a firm union 

 with their specific antibody, the antibody-adsorption technic can be 

 carried out. Both the adsorption and centrifugation should be carried 

 out in the cold (4°C) unless the virus and antibody are known to be 

 very stable. 



Dilution of the antibody may be essentially avoided if the viral particles 

 are sedimented first and then resuspended in the antibody. The time 

 to be allowed for adsorption will vary, as will the gravitational force 

 necessary to sediment the viruses with the virus and antibody employed. 

 Experimentation and consultation with previous works will have to be 



