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Appl Environ Microbiol. 1964 January; 12(1): 7-9
Copyright © 1964 by the American Society for Microbiology. All Rights Reserved.

Single-Dose Assay Technique for Variola Virus^1,2

U.S. Army Biological Laboratories, Fort Detrick, Frederick, Maryland

ABSTRACT

A biological assay for variola virus was needed that would (i) require a minimum of time and (ii) have acceptable precision. Preliminary titrations made in 11-day-old embryonated eggs and in suckling mice (6 to 24 hr of age) demonstrated a linear relationship between the concentration of variola virus injected and the mean reciprocal time to death (MTD) of both hosts. This linear response indicated that the injection of a single dose of virus suspension and the measurement of the MTD should result in an acceptable assay method. Seven replicate samples of liquid preparations (20% chorioallantoic membrane in Heart Infusion Broth) and the freeze-dried material obtained with these suspensions were assayed in triplicate for pock infectious units and for MTD. The variance of the pock counts was far greater than was expected from a Poisson distribution, and coefficients of variation ranged from 25 to 61%. Variances obtained with the single-dilution assay were all far below that expected from a Poisson distribution, and coefficients of variations ranged from 5.2 to 13.6%. The use of the MTD assay resulted in a saving of time, a saving in the number of hosts necessary per assay, and increased precision.

¹ Presented in part before the American Society of Microbiology, 63rd Annual Meeting, Cleveland, Ohio, 5-9 May, 1963.

² In conducting the research reported herein, the investigators adhered to "Principles of Laboratory Animal Care" as established by the National Society for Medical Research.