Experimental Studies of the Agar-Plate Precipitin Test of Ouchterlony

Abstract

The agar-plate precipitin test of Ouchterlony was studied experimentally in order to evaluate the effects of the following factors: Initial concentration of agar, size of Petri dish, initial NaCl concentration in agar, pH of agar, temperature of incubation, arrangement of basins, absolute and relative concentrations of antigen and antibody, discontinuous diffusion.It was found that precipitin reactions in liquid media did not provide reliable information as to the optimum concentrations of antigen and antibody in the agar-plate test.No spurious precipitate lines were observed as a result of repeated simultaneous refilling of all reservoirs with fractional amounts of antigen and antibody in optimum proportions.Temperature changes did not significantly alter the appearance of the precipitate lines. The effect of temperature changes under conditions differing from those of our experiments remains to be studied.The precipitate lines developed better in agar containing no sodium chloride than in agar containing 0.9% or higher concentrations of sodium chloride. However, omission of the sodium chloride can not be recommended for routine use until its effect on the precipitation pattern has been more fully explored.Separation of the specific precipitate into multiple zones was noted with high relative concentrations of antigen or antibody, and non-specific precipitation of highly concentrated γ-globulin was observed in salt-free agar. The significance of these findings was briefly discussed.