352 C. E. SCHWERDT 



Friedewald and Pickels (1944) estimated the mimber of influenza virus 

 particles required to infect mice and eggs as well as agglutinate chick red 

 cells. Virus particle comit was determined indirectly by dividing the dry 

 weight mass of an aliquot of purified virus by the dry mass of a single particle 

 estimated from physicochemical data. Donald and Isaacs (1954a) and Isaacs 

 and Donald (1955), in more extensive studies, counted particles of representa- 

 tive strains of aU four viruses of the myxovirus group by electron microscopy, 

 using both the spray droplet and red cell adsorption techniques. The results 

 of both groups, in summary, indicated that at the 50 % agglutination or 

 partial agglutination end points the number of influenza A virus particles 

 equaled the number of red ceUs present. They also agreed on an estimate of 

 approximately 10 particles corresponding to one ID50 from titrations in 

 embryonated eggs conducted under optimal conditions. These results 

 compare favorably with those of Werner and Schlesinger (1954), who found 

 an average ratio of 1 .2 virus particles per cell at the partial agglutination end 

 point, and of Graham and McClelland ( 1 950) and Miller and Schlesinger ( 1 955), 

 who estimated 16 and 10 particles, respectively, per ID50 in chick embryos. 



Such findings receive additional support from the many observations, 

 reviewed by Isaacs (1957), that the ratio of the number of infective doses per 

 hemagglutinating dose (I/HA) averages 10^'^ for virus preparations of high 

 infectivity (i.e., virus harvested from eggs infected at low multiplicities to 

 avoid the production of noninfective or "incomplete" virus (von Magnus, 

 1946)). Since, as inferred from above, 10'-^^ physical particles constitute one 

 hemagglutinating dose, an I /HA ratio of 10^-^ indicates that approximately 7 

 particles correspond to one ID50. 



"V^Hien influenza virus particle comits are made by the so-caUed "absolute 

 assay" hemagglutination method (Section III, B, 2) devised independently by 

 Levine et al. (1953) and Horsfall (1954), lower ratios of particles per ID50 and 

 per hemagglutinating dose by pattern test were found, m contrast to the 

 results cited above. These workers estimated that 1 or 2 virus particles 

 represent 1 egg-infective unit and that only 1 virus particle per 20 red cells is 

 present at the partial agglutination end point by pattern test. Tyrrell and 

 Valentine (1957) have made a comparison of the indirect "absolute assay" 

 method of coimting particles with direct electron microscope comits and 

 found that the indirect method yielded, on the average 10-fold lower 

 estimates of particle numbers. It would appear that the hemagglutuiation 

 method of enumerating virus particles has approximately a 10 % rather 

 than 100 % efficiency in forming red cell dimers. 



4. Other Viruses 



With the new, direct techniques for enumerating virus particles by electron 

 microscopy now available, studies on the quantitative relationship between 



