THE PHYSICAL PROPERTIES OF INFECTIVE PARTICLES 293 



for electron microscopy by Williams is simply an application of freeze- 

 drying in which particularly rapid freezing rates and low sublimation tem- 

 peratures are employed. It is gratifying to note that these two methods, so 

 widely different in principle, have been found to yield essentially identical 

 results when applied to the same kinds of biological objects. The application 

 of these methods has revised our notions of the morphology of viruses by 

 revealing, primarily, that several of the "spherical" ones are actually poly- 

 hedral. 



ii. Virus Suspensions, Quantitative. It is frequently useful to observe 

 samples of suspensions of virus particles in a quantitative manner, where 

 the observation of the details of their structures is not important. This type 

 of investigation arises when one is interested in morphological identifications 

 of viruses, where quantitative assay of specific viral infectivity is sought, 

 and where particle weights of viruses are being determined. The criteria to 

 be met in preparations made for this kind of work are: (1) that the specimen 

 field is representative of the entire suspension, and (2) that the volume of 

 suspension giving rise to a particular specimen field is known. 



Three methods of satisfying the above criteria are now in common use. 

 In order to have a representative specimen field it is necessary either to have 

 a large field, any portion of which is statistically representative of the sus- 

 pension, or a very small field that is representative and that can be micro- 

 graphed in its entirety. It turns out that in the method of agar-inhibition 

 the virus particles remaining on the collodion film are spread out at random 

 with no sign of clumping (Kellenberger and Kellenberger, 1955). The volume 

 of suspension corresponding to a given field of virus particles can be calcu- 

 lated by mixing with the suspension some indicator particles, such as poly- 

 styrene latex in known number concentrations. A second method (Sharp 

 et al., 1952) is to deposit upon some sort of substrate surface, by high-speed 

 centrifugation, all the virus particles in a known volume of suspension. 

 The particles apparently stick where they impinge upon the surface and a 

 random array is secured. The method in principle is an absolute one, 

 requiring no indicator particles. The third method is the one now most 

 commonly employed and involves the deposition of suspended material, 

 mixed with indicator particles, in the form of mmute droplets (Backus and 

 Williams, 1950). The droplets are representative of the suspension and each 

 dried pattern is small enough to allow it to be recorded on one micrograph 

 (Fig. 8). It is evident that by any of the methods one may count the numbers 

 of particles of any distinct morphological class. If previously identified, virus 

 particles may be counted, and if an aliquot concentration is biologically 

 assayed, a measurement of specific viral infectivity can be secured. Also, if 

 the suspension is reasonably monodisperse, and if contaminant material is 

 known to be absent, the particle weight of a virus may be obtained by 



