Abstract
The statistics of estimators used with the endpoint assay for virus titration were investigated. For a standard assay with 10 wells/dilution, the graphical estimator traditionally used was found to produce estimates with significant positive bias and a relatively low accuracy. Furthermore, the graphical estimator was found to be inconsistent. A superior estimator based on the maximum likelihood principle was developed. The results are discussed in relation to the choice between the endpoint titration assay and the plaque assay, and an alternative two-stage assay is presented.
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Abbreviations
- α:
-
concentration of infective virus particles (PFU/ml)
- v:
-
volume of sample in well (ml)
- λ=α.v:
-
virus particles in well (PFU)
- m:
-
no. of dilutions
- d≡(d1,d2,...,dm):
-
vector of dilutions
- n≡(n1,n2,...,nm):
-
vector of no. of wells scored for each dilution
- x≡(x1,x2,...,xm):
-
vector of observed no. of positive wells for dilutions
- X≡(X1,X2,...,Xm):
-
stochastical vector representing the observation vector
- E:
-
exponential series {...,10−2,10−1,100,101,102,...}
- λ=μɛ:
-
μ=mantissa of λɛ [1,10]; ɛ=exponent of lambda ɛ E
- δ=ɛd:
-
relative dilution vector
- t(X;d,n,λ):
-
estimator of λ
- λ=t(x;d,n,λ):
-
estimate of λ
- β:
-
relative bias
- γ:
-
variance-coefficient
- χ12(1−α):
-
(1-α)-percentile in χ2-distribution with 1 degree of freedom
References
Knudson DL and Tinsley TW (1974) Replication of a nuclear polyhedrosis virus in a continuous cell culture ofspodoptera frugiperda: purification, assay of infectivity, and growth characteristics of the virus. J. Virol. 14: 934–944.
Rao CR (1973) Linear statistical inference and its applications, 2nd Ed. John Wiley & Sons, New York.
Reed LJ and Muench H (1938) A simple method of estimating fifty percent endpoints. Am. J. Hygiene 27: 493–497.
Summers MD and Smith GE (1987) A Manual of Methods for Baculovirus Vectors and Insect Cell Culture Procedures. Texas Agricultural Experiment Station Bulletin No. 1555.
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Nielsen, L.K., Smyth, G.K. & Greenfield, P.F. Accuracy of the endpoint assay for virus titration. Cytotechnology 8, 231–236 (1992). https://doi.org/10.1007/BF02522040
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DOI: https://doi.org/10.1007/BF02522040