Serologic Analyses of Cell-Surface Antigens of Acanthamoeba spp. with Plasma Membrane Antisera*†

SYNOPSIS. Antisera were raised against plasma membrane-enriched fractions of the species Acanthamoeba castellanii and Acanthamoeba culbertsoni to determine whether cell-surface antigens would facilitate species identification of Acanthamoeba isolated from the environment or in human infections. Acanthamoeba castellanii and A. culbertsoni plasma membranes were purified, after homogenization, by differential and isopycnic centrifugation. Electron microscopic examination of purified membrane samples showed an enrichment of membranes with a typical trilaminar structure. Occasionally, mitochondria were recognized in the electron microscope preparations. 5′-Nucleotidase, Mg²⁺-ATPase, and alkaline phosphatase were enriched 11-fold, 2-fold, and 7-fold, respectively, in the A. castellanii membranes, as determined from analyses of the enzyme activities in whole cell homogenates and membrane preparations. 5′-Nucleotidase was not detected in A. culbertsoni, but the activities of Mg²⁺-ATPase and alkaline phosphatase were increased 2- to 3-fold. Both membrane preparations showed no glucose-6-phosphatase activity and less than 5% contamination with succinic dehydrogenase. From assays of acid phosphatase activity, the most apparent contamination of the plasma membrane preparations was with membranes of phagocytic vacuoles. Acanthamoeba castellanii membrane antisera produced significant agglutination and fluorescence of homologous cells to titers of 1:8192 and 1:1024, respectively. Acanthamoeba polyphaga and Acanthamoeba rhysodes gave the most cross-reactions in heterologous tests. They were agglutinated to a titer of 1:128 and positively fluoresced to titers of 1:32 and 1:64, respectively. Antisera of A. culbertsoni membrane agglutinated homologous cells at a dilution up to 1:4096 and produced homologous fluorescent titers up to 1:512. Other than agglutination of A. polyphaga to a titer of 1:128, these antisera did not cross-react significantly with any remaining heterologous species. Three new isolates were identified with these plasma membrane antisera: 2 of them, contaminants from tumor tissue cultures, were identified as A. culbertsoni. Preliminary information is also given on the use of the membrane antisera for species identification of Acanthamoeba in several new cases of amebic encephalitis.