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Anaplasma marginale-infected erythrocytes were examined for the presence of maturation, isolate-restricted, and isolate-common antigens by indirect immunofluorescence with monoclonal antibodies. A panel of 18 monoclonal antibodies was used; none of the antibodies reacted with Anaplasma ovis, Babesia bigemina, babesia bovis, Trypanosoma brucei, Trypanosoma congolense, or uninfected bovine erythrocytes. Antigens common to all six A. marginale isolates were detected by nine antibodies. Single isolates from Florida, Southern Idaho, Northern Texas, and Virginia and two isolates from Washington state had four patterns of reactivity with a second panel of nine antibodies. Antigenically distinct stages were not detected, as sequential smears taken daily during acute infection had the same pattern of reactivity. The results demonstrate antigenic heterogeneity among isolates of A. marginale and the presence of common antigens. This information allows grouping of different isolates and, more importantly, provides a method for the identification and isolation of common antigens for diagnostic tests.

Bovine erythrocytes infected with Anaplasma marginale were cultured for 1 to 5 days in a CO2 incubation chamber, pulse-labeled with [3H]thymidine and [14C]methionine, lysed, and fractionated by differential centrifugation and continuous density gradient centrifugation in Renografin. Anaplasma and associated fragments of stroma formed two distinct bands in the dense region of the gradient. Electron microscopic examination of pelleted material from the bands from cells cultured for 1 day revealed the presence of organisms that were morphologically intact or in various states of degeneration. Examination of fractions from the gradient for incorporation of label revealed that analplasma present in erythrocytes can incorporate both [3H]thymidine and [14C]methionine. Subsequent experiments demonstrated that organisms cultured for 3 and 5 days incorporated the radiolabeled compounds also, but to a lesser extent. The experiments demonstrate that it is possible to culture analplasma in vitro for short periods of time and monitor their growth characteristics.