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The Agar Diffusion Precipitin Technique: A Comparison of the Simple and Double Diffusion Methods¹

From the Departments of Pathology, Veterans Administration Hospital, Dearborn, Michigan, and Wayne State University, College of Medicine, Detroit, Michigan

Abstract

The simple and double diffusion methods were studied experimentally by means of specially designed plates. The 2 methods were compared with respect to their sensitivity, reaction range, reaction time, resolving power and susceptibility to concentration and temperature artifacts. The following observations were made:

1. The potential sensitivity of the double diffusion method is somewhat greater than that of the simple diffusion method.

2. The reaction range of the simple diffusion method is unlimited in the antigen excess zone but is restricted to only a narrow segment of the antibody excess zone. The reaction range of the double diffusion method is unlimited in both directions.

3. The effective reaction time, i.e., the time interval during which a precipitate band persists in the column, is the same for both methods at intermediate or high concentrations of antigen and antibody, but at relatively low concentrations of the reactants the effective reaction time of the simple diffusion method is shorter than that of the double diffusion method.

4. The resolving power of the double diffusion method for mixed precipitin systems is considerably better than that of the simple diffusion method.

5. The sensitivity of the simple diffusion method to concentration and temperature changes is much greater than the sensitivity of the double diffusion method. Under average laboratory conditions temperature artifacts in double diffusion columns or plates have never been observed, but special precautions must be taken to prevent such artifacts in simple diffusion columns.

Footnotes

This investigation was supported by a research grant RG-4892 from the National Institutes of Health, Public Health Service.