Immunoblot analysis of the humoral immune response to Leishmania donovani infantum polypeptides in human visceral leishmaniasis

Abstract

Using the immunoblot technique, we have compared the reactions of Leishmania donovani infantum polypeptides with the immunoglobulin G of human sera from patients with parasitologically proven L. d. infantum infection, with suspected visceral leishmaniasis, and with other leishmaniases, protozoiases, helminthiases, and fungal or bacterial diseases. A 94-kDa component reacted with all L. d. infantum-infected sera and with 75% of sera from patients with clinical and serological but no parasitological diagnoses. No reaction was observed with sera from patients in the other disease groups or with control sera. Studies of eight different isolates, subspecies, and species of the genus Leishmania demonstrated that the 94-kDa component was expressed in all strains examined.

Presence of antibodies in the aqueous humour and cerebrospinal fluid during Leishmania infections in dogs. Pathological features at the central nervous system

“…Although most of the bands were common to all sera we detected a certain variability in the antigen-binding pattern revealed by different samples, particularly in the 30-19 kD region. In agreement with previous reports for the humoral response to Leishmania donovani in human visceral leishmaniasis (Rolland-Burger et al 1991) we observed that the most intensively labelled bands corresponded to molecular masses of 94, 75, 70, 66-60, 40-30, 20-17. We also observed that the sera from these dogs strongly react with the Leishmania purified Hsp70 (70Kd), LiP0 (32kd), H2A (14kd), LiP2a and LiP2b (13kd) proteins (Laboratory data).…”

Identification of a 94‐kilodalton antigen on Leishmania promastigote forms and its specific recognition in human and canine visceral leishmaniasis

“…In a previous study (Rolland-Burger et al 1991), we reported that a 94-kDa component from L. infantum promastigotes was recognized in immunoblot by serum antibodies from VL-infected Greek patients, but not by sera from patients with other leishmaniases, protozoiases, helminthiases, and fungal or bacterial diseases. The present report confirms and extends these results.…”

“…Cross-reactions among various species of Leishmania are well known (McMahon-Pratt & David 1981, Lepay, Nogueira & Cohn 1983, Gardiner, Jaffe & Dwyer 1984, Anthony et al 1985, Colomer-Gould et al 1985, Edges et al 1985, Bouvier, Etges & Bordier 1987, allowing crossreactivities among VL and CL sera. In particular, we previously showed (Rolland-Burger et al 1991) that the 94 kDa antigen was detected by 1 : 200 diluted VL sera in L. infantum and L. donovani species, but also in those causing CL (L. tropica, L. major, L. guyanensis, L. mexicana). However, the 94 kDa antigen was not detected by 1 : 200 diluted CL sera.…”

“…The parts of membranes containing molecular mass standards were stained with India ink (Hancock & Tsang 1983). Immunoblot assays were performed as previously described (L. Rolland-Burger et al 1991). Briefly, strips were blocked with 5% skimmed milk in 100mM PBS, 0.3% Tween 20, washed in 100mM PBS, 0.1 % Tween 20 (washing buffer), then incubated for 2 h at 37°C (unless stated otherwise) with sera diluted 1 : 200 (unless stated otherwise) in washing buffer.…”

“…Previously we have shown by using the immunoblot technique, that a 94-kDa component, also present in eight Leishmania species/strains tested (promastigote forms), was recognized by sera from Greek patients with VL infection but not by sera from patients with other diseases, including cutaneous leishmaniasis (CL) and Chagas' disease (Rolland-Burger et al 1991, Rolland et al 1992). In the present study, we report that (i) circulating anti-94 kDa antibodies are present in the sera both from humans and dogs with VL from Old World (OW) as well as New World (NW), even at high dilutions, (ii) sera from patients with other infections than leishmaniases or without infection are negative, even at low dilution, and (iii) the immunostaining of the 94 kDa band by VL sera may be dependant of the stage (promastigote/amastigote) of the parasite.…”