Ronald L. Anthony

On-site diagnosis of Plasmodium falciparum, P. vivax, and P. malariae by using the Quantitative Buffy Coat system.

The Quantitative Buffy Coat (QBC) system was used for the detection and identification of malaria parasites in blood specimens from 570 residents of Oksibil, an isolated highland valley in the eastern Jayawijaya Mountains of Irian Jaya (Indonesian New Guinea). The availability of a battery-powered centrifuge and a fiberoptic Paralens enabled us to complete and interpret the assay in this remote environment. Of 322 QBC tubes examined for 2-4 min each, results of 295 (92%) concurred with findings on the matched Giemsa-stained thick smear (GTS). The 27 discrepant results included 13 QBC+/GTS- that, upon reexamination, were found to be GTS+. When using the corrected GTS results as the standard, the sensitivity and specificity of the QBC were 94% and 96%, respectively. Because electricity was available only 3 hr per day, it was decided to decrease the examination for an additional 248 QBC to a maximum of 90 sec per tube. This shortened inspection time resulted in a reduction of sensitivity to 53% but specificity was preserved at 89%. Forty-two of 45 conflicting results, QBC-/GTS+ from cases of light Plasmodium falciparum infections with < 1 trophozoite or gametocyte per field, were resolved by reexamination of the QBC in the laboratory. Tubes held at 4 C could be reexamined, without noticeable loss of fluorescence, for at least 6 wk after collection. Despite some difficulty in the identification of Plasmodium species, it was concluded that the QBC is an easy, sensitive method for the rapid diagnosis of malaria in the field and that it provides the inexperienced microscopist with an additional means for on-site identification of individuals needing treatment.

Production of a Monoclonal Antibody against the Sea Nettle Venom Mouse Lethal Factor

Abstract A monoclonal antibody to the sea nettle, Chrysaora quinquecirrha, venom mouse lethal factor was prepared using the crude venom as the antigen. Cloned hybridomas were selected by an enzyme-linked immunosorbent assay (ELISA) using partially purified lethal factor as the antigen. Ascites fluid from the cloned hybridoma-bearing mouse had an ELISA titer of 12.800 and the capability to neutralize the lethal activity (2 LD50) of the intravenous injection of crude venom into mice after overnight incubation.

Long-Term Culture of Hamster Duodenal Explants and Cells

Explants from adult Syrian hamster duodenum have been maintained in organ culture on gelatin sponges using CMRL 1066 serum supplemented media for 30 days. There was necrosis of the tall villi architecture during the 1st week in culture while columnar and mucous cells survived, migrated, and replicated along portions of the explant basement membrane and in the gelatin sponge matrix. Cells in the sponge multiplied and formed epithelial sheets which showed villus projections and cyst configurations. The cells in these epithelial structures were attached to one another by junctional complexes. Epithelial cells were isolated from the sponge matrix by collagenase digestion and were successfully grown in culture. These duodenal explants and cells show potential for use as in vitro models for experimental studies, involving acute and chronic response of cells to injury or carcinogen-induced transformation.

Measurement of carcinoembryonic antigen in serum of patients by using a technique of passive hemagglutination inhibition

Abstract A technique of passive hemagglutination inhibition (PHI) which can detect nanogram levels of carcinoembryonic antigen (CEA) in serum of patients has been developed. Human O-negative erythrocytes were sensitized with CEA recovered from primary adenocarcinomas of the colon and rectum. Sera from patients were then examined for their capacity to inhibit the agglutination of the sensitized erythrocytes in the presence of a predetermined amount of goat anti-CEA serum. Positive sera were defined as those which produced inhibition of agglutination at a dilution of 1:8 or greater. The percentage of positive sera was 82% for primary adenocarcinomas, 55% for all other cancers, 62% for benign diseases of the gastrointestinal tract, 60% for alcoholic cirrhosis, and 17% for normal healthy controls. Results are presented for 240 patients and attempts have been made to correlate PHI titers with CEA levels determined by radioimmunoassay.

Flow Cytometric Analysis of Leishmania Surface Membrane Antigen Expression

The advent of cell fusion technology for the construction of hybridomas synthesizing monoclonal antibodies (Kohler and Milstein, 1975) has provided the immunoparasitologist with the means to detect subtle antigenic differences on the surface membrane of the leishmania parasites which cause human disease. Use of these antibodies in radioimmune binding assays (McMahon-Pratt and David, 1981; Jaffe and McMahon-Pratt 1983; Jaffe et al., 1984; Pan et al., 1984; McMahon-Pratt et al., 1985; Grimaldi et al., 1987), immunofluorescent antibody assays (Handman and Hocking, 1982; de Ibarra et al., 1982; Lemesre et al., 1985) and enzyme linked immunosorbant assays (Fong and Chang, 1982; Anthony et al., 1985) has now culminated in the identification of a sizeable composite of species-specific, subspecies-specific and stage-specific epitopes. Most importantly, the ability to demonstrate such epitopes on parasites taken directly from the patient1s lesion (Lynch et al., 1986; Anthony et al., 1987) permits a prompt diagnosis of the infection. Such diagnoses are critical when attempting to decide regimens of treatment, management of the patient and prognosis.