The Isolation of Microfilariae from Blood for Use as Antigen

Abstract
This report describes the isolation of microfilariae from the blood of dogs infected with Dirofilaria immitis and their use as test antigen in sero-logical tests. Citrated blood is hemolyzed with saponin and the microfilariae are washed, shaken and centrifuged free of the circulating blood elements and from the soluble blood substances. The microfilariae then isolated are in a conc. of 200 or more times that of the original blood, and have remained viable for 2 weeks or longer in a menstruum of balanced salt soln. or plasma. The antigen used is a saline extract of the microfilariae which has been prepd. by freeze drying. It was calculated that there were approx. 500 million microfilariae per gm. dry wt. Precipitin tests were made on the sera of 20 dogs with varying degrees of microfilaremia and on the sera of 8 men known to have filariasis. The tests were positive at dry wt. dilutions of antigen, ranging from 1/1600 to 1/25,000. Parallel tests set up with extracts made from dried D. immitis gave results similar to those with the microfilarial antigen, but at a lower titre. What appeared to be an agglutination phenomenon was noted when a suspension of living microfilariae was added to infected serum. This phenomenon was checked by observing suspensions of living microfilariae in infected serum, normal serum, Tyrode and saline soln. in hanging drop prepns. and in thin film prepns., and it was concluded that the "lining up" phenomenon was due to surface tension in a thin film rather than an agglutination reaction.