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C L Jaffe

Publications and source records attributed to C L Jaffe.

At least 55 records · Page 3Linked to original sources
PubMed1989

Amastigote stage-specific monoclonal antibodies against Leishmania major.

Monoclonal antibodies were produced against gamma-irradiated amastigotes of Leishmania major. Five antibodies (T16 through T20) were selected which reacted in enzyme-linked immunoassays with the intracellular stage of the parasite. These antibodies did not react with promastigotes of L. major or Leishmania donovani. One of the monoclonal antibodies (T16) reacted with amastigotes of Leishmania mexicana amazonensis and L. donovani. Western blotting (immunoblotting) and immunoprecipitation of [35S]methionine-labeled amastigotes demonstrates that T16 reacted with multiple L. major amastigote components between 12 and 180 kilodaltons. Antibody T20 was shown to recognize a low-molecular-mass doublet (less than 26 kilodaltons) in both [14C]leucine- and [35S]methionine-labeled amastigotes. A protein of less than 180 kilodaltons was also weakly recognized by T17, T19, and T20 in metabolically labeled amastigotes. This protein reacted strongly with T16. The reactive antigens could be identified on the surface of amastigotes isolated from the lesions of infected mice and on newly transformed amastigotes within 24 h after the infection of mouse peritoneal macrophages by promastigotes. These monoclonal antibodies should prove useful for the diagnosis of L. major in human tissue biopsies.

Animals↗
PubMed1989

Autoantibody production by patients infected with Leishmania.

Sera from 29 patients with visceral leishmaniasis and 14 patients with cutaneous leishmaniasis were tested against a panel of nine nuclear antigens employing an enzyme-linked immunosorbent assay (ELISA). Anti- Sm, RNP, SS-A and SS-B antibodies were present in high titres in 83, 86, 36 and 73 per cent of the patients with visceral leishmaniasis and in 7, 14, 25 and 25 per cent of the patients with cutaneous leishmaniasis. One serum from a patient with visceral leishmaniasis which reacted strongly with Sm, RNP, SS-A and SS-B was examined by immunoblotting on extractable nuclear antigen from Hela cells. This serum binds to nine different antigenic bands (16, 23, 29, 30, 40, 50, 58, 100 and 115 kD). These same antigens were recognized by serum from a patient with systemic lupus erythematosus. The binding of visceral leishmaniasis serum antibodies to ribonucleoproteins was inhibited by prior incubation of serum with either leishmanial membrane antigens, from four different species of Leishmania, or intact cells of Leishmania donovani, implying molecular resemblance between common leishmanial antigens and ribonuclear antigens. It seems that appearance of autoantibodies to ribonucleoproteins in sera of patients infected with Leishmania is not only due to simply polyclonal activation of lymphocytes, but is also the result of a molecular mimicry between leishmanial antigens and ribonucleoproteins.

Antibodies, Antinuclear↗
PubMed1988

Purification of two Leishmania donovani membrane proteins recognized by sera from patients with visceral leishmaniasis.

Two Leishmania donovani membrane proteins recognized by sera from patients with visceral leishmaniasis were purified using species-specific monoclonal antibodies and characterized. The molecular weights of the proteins, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, were approximately 70,000 and approximately 72,000, respectively. The 70 kDa protein, which appears as a diffuse band on silver staining, was resolved into a doublet by Western blotting with monoclonal antibody. Though of similar molecular weight and amino acid composition, the two proteins were shown to be distinct by peptide mapping and Western blotting of the purified material. The two proteins are recognized specifically by human visceral leishmaniasis serum and not by serum from cutaneous leishmaniasis or Chagas' disease. These proteins will be useful in developing a direct serodiagnostic assay for visceral leishmaniasis.

Animals↗
PubMed1988

Temperature effects on molecular processes which lead to stage differentiation in Leishmania.

We have studied the effects of growth phase and temperature on the morphological and molecular processes that occur during stage differentiation of Leishmania. Parasites which differ in their ability to transform axenically were compared. A typical heat shock response is observed in strains that transform axenically. Heat shock proteins (HSPs) of 70 and 83 kd are transcribed and synthesized along with a decrease in cellular protein synthesis, including tubulin. Changes in the transcription of the tubulin gene are also noted. In strains which do not transform in culture HSPs are induced, but cellular protein synthesis is unaffected and no differences are observed in tubulin transcription and expression. HSPs 70 and 83 remain constitutively expressed in amastigotes. HSP 83 transcription increases along the promastigote growth curve. beta-Tubulin transcription is also affected by growth phase and temperature increase, though alpha-tubulin remains unaltered. An amastigote-like hybridization pattern is induced in heat-shocked promastigotes, in which a larger transcript for beta-tubulin (2.8 kb) becomes dominant and the promastigote transcript (2.4 kb) decreases. Tubulin expression is susceptible to temperature control in Leishmania mexicana amazonensis, though direct correlation to HSP expression was not demonstrated.

Animals↗
PubMed1988

Canine visceral leishmaniasis at Wadi Hamam, in Israel.

A preliminary survey of canine leishmaniasis was made in Israel by serodiagnosis, using a direct and competitive enzyme-linked immunosorbent assay. Three (4%) of the 75 dogs surveyed were seropositive. The infected dogs came from Wadi Hamam near Tiberias in northern Israel. Parasites isolated from one dog by needle aspiration were identified as Leishmania donovani sensu lato by their excreted factor.

Animals↗
PubMed1988

Use of purified parasite proteins from Leishmania donovani for the rapid serodiagnosis of visceral leishmaniasis.

Serodiagnosis of visceral leishmaniasis (VL) due to Leishmania donovani by using crude parasite antigen is complicated in many endemic areas because of cross-reactions with sera from humans with Chagas' disease. We used affinity-purified parasite proteins to develop a direct dot-blot assay for VL. Double-blind tests were carried out on sera from 40 patients with well-documented VL, from controls in endemic areas, and from patients with other diseases. Using gp70-2, 36 (90.0%) of 40 sera from patients with kala azar were correctly diagnosed; 1 (1.2%) of 86 sera from patients without kala azar was misdiagnosed. With dp72, 21 (100%) of 21 sera from patients with VL were correctly identified; 5 (7.0%) of 71 negative sera were misdiagnosed. None of the 18 sera from patients with Chagas' disease reacted positively against gp70-2. Our assay is rapid, simple, and specific and represents a new method for reliably diagnosing and monitoring VL.

Animals↗
PubMed1988

The identification of membrane glycoconjugates in Leishmania species.

The membrane glycoconjugates of 8 different species of Leishmania were compared by lectin blotting. Five different lectins with various sugar specificities were examined: concanavalin A, Lens culinaris, Ricinus communis, soybean agglutinin, and peanut agglutinin. Concanavalin A and Lens culinaris reacted with every Leishmania tested. The patterns observed for these 2 lectins, as well as the various species of parasites, were different. However, a common 41,000-52,000 and a 160,000-185,000 Mr component was present in almost all the parasite isolates examined. Ricinus communis only recognized a nondiscrete galactose-containing glycoconjugate similar to Leishmania-excreted factor. Soybean and peanut agglutinins reacted with a few low molecular weight parasite components. Soybean agglutinin reacted with all the Leishmania species tested, whereas peanut lectin only recognized 3 isolates. The latter lectin bound to discrete components migrating with the dye front and with Mr's of 35,000 and 52,000. Increased glycosylation was noted on avirulent L. major promastigotes and was associated with the appearance of several new peanut agglutinin-binding glycoproteins.

Agglutination↗
PubMed1987

Antigen-stimulated lymphokines from patients with cutaneous leishmaniasis induce monocyte killing of Leishmania major intracellular amastigotes.

The specific immune response of patients with cutaneous leishmaniasis including the ability of their lymphokines to enhance the monocytes' leishmaniacidal activity was studied. In 16 patients with cutaneous leishmaniasis, their concanavalin A-induced lymphocyte proliferative responses, interferon-gamma and interleukin 2 activities and the ability of their concanavalin A-induced lymphokines to kill monocyte intracellular amastigotes were not different from normal controls. Antigen-stimulated lymphocyte cultures showed that 13 of 13 patients had an increased lymphocyte proliferative response; 11 of 16 produced interleukin 2 and 12 of 13 produced interferon-gamma; in addition, 10 of 11 of these antigen-induced supernatants increased the monocytes' killing of Leishmania major amastigotes. Antibody levels to parasite membrane antigens determined by radioimmunoassay showed that 8 of 13 titers were greater than 10 and 4 of 13 titers were 2 to 10 times higher than control. Our findings demonstrate that patients with cutaneous leishmaniasis elicit a specific immune response to L. major antigens and part of this response is the production of lymphokine capable of promoting monocyte killing of intracellular amastigotes.

Animals↗
PubMed1987

Species-specific antibodies to Leishmania tropica (minor) recognize somatic antigens and exometabolites.

Monoclonal antibodies were raised against membranes of Leishmania tropica (formerly L. tropica minor). These antibodies do not react with membranes or homogenates prepared from any other species of Leishmania examined. The antigens identified by all four monoclonal antibodies reside on the external surface of the promastigote and are released into the culture medium. Immunoprecipitation of metabolically ([3H]glucose, -glucosamine, or [35S]methionine)-labeled, [125I]lactoperoxidase-labeled, or Western blot analysis of the parasites identified species-specific determinants residing on several different molecules with Mr between 12,000 and 140,000. All of the monoclonal antibodies appear to recognize determinants on a common [3H]sugar-labeled glycoconjugate, Mr = 32,000 to 44,000, but also react with additional components of 55,000, 80,000, 92,000, and 130,000 m.w. Antibody binding to the parasites is very sensitive to mild oxidation with periodate, suggesting that the species-specific determinants are carbohydrate in nature.

Animals↗
PubMed1987

Routine dot-blot assay of multiple serum samples using a simple apparatus.

This paper describes a simple method for carrying out dot-blot immunoassays simultaneously with a large number of different sera. No dot-blot manifold is necessary, only a modified 96-well flat-bottom microtiter plate. This apparatus is re-usable, inexpensive and especially suited for field testing of patient sera in serodiagnostic assays. One application of this technique, to the serodiagnosis of visceral leishmaniasis on a pure parasite protein, is demonstrated.

Antibodies↗
PubMed1987

Serodiagnostic assay for visceral leishmaniasis employing monoclonal antibodies.

A highly specific and sensitive competitive serodiagnostic assay for visceral leishmaniasis (VL) was developed using species specific Leishmania donovani monoclonal antibodies. This assay, either RIA or ELISA, is based on the specific inhibition of monoclonal antibody binding to a crude parasite homogenate by serum from patients with VL. 15 monoclonal antibodies were examined. The binding of 13 antibodies was significantly inhibited by VL serum and unaffected by normal serum. 3 species-specific monoclonal antibodies, D-2, D-13 and D-14, which recognize different parasite antigens, were chosen for use in the competitive serodiagnostic assay. In 90% of the positive cases, regardless of geographic origin, VL sera inhibited monoclonal antibody binding to the parasite antigen by more than 30%. No false positive was obtained with sera from Chagas disease, lepromatous leprosy, schistosomiasis, malaria, systemic lupus erythematosus, cutaneous or mucocutaneous leishmaniasis, even at serum dilutions (1:100) which cross-react strongly with Leishmania antigen in direct binding assays. Inhibition by negative control sera from areas endemic for VL and from non-endemic areas was negligible. The assay takes less than 24 h, requires minimum amounts of sera or antigen, and is easily standardized allowing interlaboratory comparison of test data. The competitive serodiagnostic assay will be especially useful in areas where Chagas disease is coendemic and the rapid diagnosis of VL by direct binding serodiagnostic assays presents a problem.

Antibodies, Monoclonal↗
PubMed1985

Brazilian Leishmania stocks phenotypically similar to Leishmania major.

Screening by enzyme electrophoresis of isolates of New World Leishmania from different geographic areas revealed a number of stocks with enzyme profiles different from those produced by reference strains of described subspecies of L. mexicana, L. braziliensis, and L. donovani. Analysis by six enzymes (aspartate aminotransferase; alanine aminotransferase; malate dehydrogenase; glucose-6-phosphate dehydrogenase; phosphoglucomutase; and glucose-phosphate isomerase) showed that these stocks have identical enzyme profiles and form a distinct zymodeme grouping. These observations were confirmed using the technique of schizodeme analysis and by comparing the k-DNA fingerprints produced by the restriction enzymes MspI, BspRI and AluI. The stocks were further analyzed by monoclonal antibodies and did not react with any of a large panel of L. mexicana, L. braziliensis, and L. donovani species- and/or subspecies-specific monoclonal antibodies using either an indirect radioimmune binding assay or immunofluorescence. These stocks did, however, react with a panel of monoclonal antibodies specific for L. major (formerly L. tropica major). Furthermore, the stocks could not be differentiated from L. major reference strains by enzyme electrophoresis nor could they be distinguished qualitatively from L. major based on their reactivity patterns using 10 Old World cutaneous species- and subspecies-specific monoclonal antibodies. Kinetoplast DNA restriction enzyme profiles, however, were different between these stocks and L. major reference strains. The implications of these results are discussed including the existence of other L. major-like stocks currently misidentified or uncharacterized.

Adult↗
PubMed1985

Subspecies- and species-specific antigens of Leishmania mexicana characterized by monoclonal antibodies.

Monoclonal antibodies have been produced that are specific for the reference stocks of Leishmania mexicana species and subspecies L. mexicana mexicana(L11, M379), L. mexicana amazonensis (WR303, H6, LV72), and L. mexicana pifanoi (L20). The specificities of these antibodies were confirmed by analyses employing an indirect radioimmune binding assay and 107 stocks of New World Leishmania. The molecules associated with these species- and subspecies-specific determinants have been characterized by Western blot analysis and consist of mainly low m.w. (11,000 to 50,000) membrane-associated components.

Animals↗
PubMed1984

Identification of cross-reactive promastigote cell surface antigens of some leishmanial stocks by 125I labeling and immunoprecipitation.

Externally oriented surface membrane constituents of promastigotes from several Leishmania species were radiolabeled with 125I. Autoradiographs of cell surface-labeled and sodium dodecyl sulfate-polyacrylamide gel electrophoresis-separated proteins of the stocks revealed distinctive patterns of bands in the molecular weight range of 6,000 to 240,000. Immunoprecipitation of detergent extracts of the labeled promastigote stocks with anti-Leishmania donovani membrane serum demonstrated that each of the stocks contained some antigenically cross-reactive determinants. The electrophoretic patterns of these determinants serve both to distinguish the parasite stocks (by unique, species-specific patterns) and to indicate antigenic similarities in stocks thought to be different by other biochemical criteria. At least 12 cross-reactive cell surface antigens in two New World leishmanias are recognized by polyvalent anti-L. donovani serum, suggesting that these common leishmanial antigens may account for the documented serological cross-reactivities among various Leishmania species. In all stocks tested, an iodinated protein was identified which had a relative molecular weight of 65,000 under reducing conditions but which demonstrated an increase in relative mobility in sodium dodecyl sulfate-polyacrylamide gels under nonreducing conditions. Distinctive patterns of the antigens common to the several stocks were also demonstrated with the use of monoclonal antibodies.

Animals↗
PubMed1984

Production and characterization of species-specific monoclonal antibodies against Leishmania donovani for immunodiagnosis.

Sixteen species-specific monoclonal antibodies were produced against membranes of Leishmania donovani. These antibodies only reacted with determinants present on L. donovani. No cross-reactions were found with any other species of Leishmania or with membranes of Trypanosoma cruzi. An extensive analysis of the binding specificities of selected antibodies was carried out by using whole promastigote homogenates as antigen. Monoclonal antibodies D-1, D-2, D-3, and D-4 correctly identified all 44 L. donovani stocks from a cross-panel of 84 New and Old World Leishmania stocks. Antibodies D-1 and D-2 were also useful for species classification by immunofluorescence. No cross-reactions were observed with any other Leishmania species examined. Based on either Western blot and/or radioimmunoprecipitation analyses, five distinct groups of molecules associated with L. donovani-specific antigenic determinants were identified. These molecules range in m.w. from 18 to 84 kilodaltons. The antigenic molecules recognized by antibodies D-2, D-10, and D-13 are also recognized by antibodies present in sera from patients with visceral leishmaniasis (kala-azar). Kala-azar sera obtained from cases in both the Old and New World specifically compete with these monoclonal antibodies for the appropriate antigenic determinants in Western blot analysis. These monoclonal antibodies and/or the purified protein antigens may be useful in the development of a serologic assay for the clinical diagnosis of visceral leishmaniasis caused by L. donovani and in epidemiologic studies of leishmaniasis.

Animals↗