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Disk Plate Assay for Diphtheria Antitoxin in Undiluted Horse and Human Serum and Infants' Whole Blood

From the Connaught Medical Research Laboratories and the School of Hygiene, University of Toronto, Toronto, Canada

Abstract

1. The disk plate assay has been adapted to the determination of diphtheria antitoxin in horse and human serum and infants' blood. Changes in the technical details previously described for polio-antibodies are given.

2. A linear relationship was found between zone diameter and the logarithm of antitoxin concentration from about 400 to 4 u/ml when indicating doses of toxin of 0.01 Lf or less were used. At concentrations below 4 u/ml marked curvature appeared. Over the whole range of dilutions, the relationship of the square of zone diameter and logarithm of antitoxin concentration showed no significant deviation from linearity, although minor systematic deviations occurred.

3. The estimated potencies on 85 samples of horse serum were in fair agreement with those obtained by flocculation assay. The standard deviation of an individual estimate was 12% of titer for 75 samples lying within the range of the reference serum used.

4. With 169 sera from children before and after a recall dose of DPT Polio Vaccine, the sensitivity of the disk plate assay was found to be similar to the rabbit skin "screening" test at 0.01 u/ml. Results by disk plate assay of 92 sera were compared with those by rabbit skin assay at the 0.001 u/ml level. The estimates by the rabbit skin assay were about one half of those found by disk plate assay for sera of low potency, the ratio tending to increase somewhat with the titer of antitoxin. The standard deviation of an individual estimate was about 10% of potency.

5. Microassay of infants' whole blood proved practical and gave more precise values than are readily obtained by rabbit skin assay. Within the limits of comparison, the values for whole blood by disk plate assay tended to be unexpectedly high.

6. The disk plate method of assay for diphtheria antitoxin proved to be simple, less laborious and more precise than conventional methods. It appeared to measure antitoxin per se, relatively uninfluenced by avidity.