PubMed Health⌕ Search

Biomedical subjects

F M Ubeira

Publications and source records attributed to F M Ubeira.

At least 55 records · Page 3Linked to original sources
PubMed1995

A sandwich immunoassay to quantify low levels of turbot (Scophthalmus maximus) immunoglobulins.

We have developed an enzyme-linked immunosorbent assay for the quantification of turbot (Scophthalmus maximus) immunoglobulin (Ig). The capture antibody is a rabbit polyclonal antiserum to turbot Ig, and the detector antibody a monoclonal antibody (UR3) to the turbot Ig heavy chain. Both antibodies bind nearly 100% of turbot Ig. The assay allows detection of turbot Ig in serum at concentrations as low as 0.16 micrograms ml-1 and takes less than 4 h. Precision is satisfactory, with intra-assay coefficients of variation (CVs) ranging from 2.1 to 16.6%, and inter-assay CVs ranging from 5.8 to 24.6%. We used the assay to determine Ig concentrations in the sera of healthy turbot of different weights. Mean serum Ig concentration was 3.35 +/- 0.74 mg ml-1 for fish weighing 15-25 g and 11.14 +/- 1.87 mg ml-1 for fish weighing 1000-2000 g.

Animals↗
PubMed1995

Anisakis simplex: stage-specific antigens recognized by mice.

To investigate antigenic differences between the developmental stages of Anisakis simplex, somatic, excretion-secretion and detergent-solubilized surface antigen preparations of third- and fourth-stage A. simplex larvae, and a somatic antigen preparation of adults, were characterized by SDS-PAGE and immunoblotting using mouse immune serum containing antibodies against the two larval stages. Excretion-secretion and surface antigen preparations from third-stage larvae behaved very differently from those of fourth-stage larvae, in both SDS-PAGE and immunoblotting; this suggests that certain excretion-secretion and surface antigens are highly stage-specific. By contrast, somatic components of third- and fourth-stage larvae were found to have very similar banding patterns, suggesting conservation of these components during the development of the parasite in mice. The SDS-PAGE and immunoblotting results for adult somatic components seem to support this hypothesis.

Animals↗
PubMed1994

Monoclonal antibodies to turbot (Scophthalmus maximus) immunoglobulins: characterization and applicability in immunoassays.

Five monoclonal antibodies (mAbs) to immunoglobulins (Igs) of the turbot Scophthalmus maximus were produced and characterized. All the mAbs (denominated UR1, UR3, UR4, UR6 and UR7) are of isotype IgG1/kappa and show good anti-turbot Ig reactivity in enzyme-linked immunosorbent assay (ELISA) and immunoblotting. Results of competitive ELISA and immunoblotting analysis indicate that these five mAbs react with at least three different epitopes on the turbot Ig H chain. Except in the case of UR1, reactivity with periodate-treated purified turbot Ig was much lower than with the untreated Ig, suggesting that carbohydrate residues are involved in epitope recognition. All the mAbs showed reactivity with sera from the closely related species Scophthalmus rhombus but not with sera from species of other flatfish genera. One of these mAbs (UR3) has been successfully applied for the detection of antibodies against Vibrio anguillarum in ELISA.

Animals↗
PubMed1994

Molecular analysis of antigens targeted by protective antibodies in rapid expulsion of Trichinella spiralis.

Rapid expulsion is a protective immune mechanism which eliminates as much as 99% of a challenge infection of Trichinella spiralis muscle larvae from the gastrointestinal tract of suckling rats. Protective monoclonal antibodies (mAbs) generated against larval excretory-secretory antigens (ESA) specifically recognize a 45-kDa glycoprotein, gp45, in addition to a distinct profile of other cross-reactive antigens that are also recognized by non-protective mAbs. Recent data indicate that protective mAbs recognize carbohydrate epitopes. To complement biochemical studies on the target(s) of rapid expulsion, we describe here the cloning and characterization of the cDNA, TspE1, which belongs to a multigene family and encodes several larval proteins in the 40-50-kDa range. A second cDNA, TspM6 encodes a 45-kDa antigen and is homologous to the published sequence of gp45. Anti-TspE1 antibodies detected antigens within beta- and gamma-stichocytes while anti-TspM6 antibodies detected antigens within alpha-stichocytes of the secretory organs of muscle larvae. Sequence analysis has provided no functional information on the encoded gene products. Neither recombinant antigen is recognized by the mAbs but native parasite molecules with peptide homology to both the TspE1 and TspM6 recombinant antigens bear the glycan recognized by the protective mAbs. These molecules are candidate targets in rapid expulsion.

Amino Acid Sequence↗
PubMed1994

Glycans as targets for monoclonal antibodies that protect rats against Trichinella spiralis.

We have investigated the role of glycans on Trichinella spiralis antigens in recognition by rat monoclonal antibodies (mAbs) which protect rat pups against challenge with the parasite. In pups born to infected dams or pups passively immunized with mAbs, antibodies eliminate a challenge dose from the intestine within hours ('rapid expulsion'). Because such dramatic protection can be afforded by mAbs, we have sought to characterize the parasite antigens they target. In this report we show that protective antibodies were unable to bind excretory/secretory (ES) antigens deglycosylated with trifluoromethanesulphonic acid (TFMS). In addition, oligosaccharides isolated from glycoproteins by alkaline hydrolysis or peptide: N glycosidase F (PNGase F) digestion were bound by protective, but not non-protective, mAbs. Glycans affinity purified with protective mAb 9D bound to all but one protective mAb. These antibodies have been shown previously to bind to the surfaces of intact larvae, indicating that the glycan is exposed on the parasite surface. Candidate glycans that may be involved in binding protective mAbs have unusual tri- and tetra-antennary structures with terminal tyvelose moieties (Reason et al., Glycobiology, 4, 000-000, 1994). Coating of the larval surface with such glycans may serve to protect the parasite and its secreted products from enzymatic attack as the parasite travels to and resides in its epithelial niche.

Animals↗
PubMed1993

A new cell culture method (the lateral diffusion system) suitable for the induction of antibody-forming cells in vitro.

The lateral diffusion system (LDS) is a new cell culture method suitable for the in vitro induction of antibody-forming cells, based upon the diffusion of nutrients and other molecules between two contiguous compartments separated by a vertical microporous membrane. We compared an LDS cell culture plate with two commercially available diffusion culture units, the Marbrook chamber (C.A. Hendley Ltd.) and the Millicell-HA cell culture unit (Millipore Corporation), in terms of ability to support spleen cells responding to in vitro exposure to a soluble antigen (phosphorylcholine (PC) coupled to keyhole limpet haemocyanin) and two particulate antigens (a PC-rich microsomal fraction obtained from the parasite Trichinella spiralis, and sheep red blood cells). We found that, at high cell culture densities, similar numbers of plaque-forming cells were induced using the three units, but that the LDS is superior in versatility and ease of use to any of the devices currently used for PFC induction in vitro.

Animals↗
PubMed1993

Requirements for the induction of cross-reactive anti-Trichinella IgE antibodies in mice.

Mice primed with Trichinella spiralis or T. pseudospiralis and reinfected with either the homologous or the heterologous species produced high levels of IgE antibodies that cross-reacted with the non-inducing strain in passive cutaneous anaphylaxis assays. Cross-reactive antibodies were not induced by primary infection. Cross-reactivity persisted for more than 6 months following secondary infection or destruction of encysted larvae with mebendazole. Both the prevention of larvi-position by thiabendazole and the interruption of infection using naphthalophos indicated that the presence of the pre-adult stage alone provided sufficient priming for the induction of detectable levels of cross-reactive IgE by subsequent reinfection. These results suggest the existence of two sets of Trichinella allergens, one comprising species-specific major allergens (MAs) and the other comprising minor allergens (mAs) evoking a cross-reactive IgE response that occurs to a detectable extent only when the response to MAs has reached its ceiling. These findings are relevant to the design of experiments investigating the role played by IgE antibodies in protection against reinfection in rodents.

Animals↗
PubMed1993

Quantification of low levels of rainbow trout immunoglobulin by enzyme immunoassay using two monoclonal antibodies.

An enzyme immunoassay has been developed for quantitation of low levels of trout immunoglobulin (Ig). This assay uses two monoclonal antibodies, one as capture antibody and the other as detector, directed against two non-overlapping epitopes on the heavy chains of trout Ig. The assay shows high reproducibility and can detect 0.12 micrograms trout Ig ml-1. Coefficients of intra- and interassay variation ranged from 3.8 to 7.1% and from 7.9 to 17.4%, respectively. Analysis of 37 healthy trout showed increasing serum Ig concentration with size. The mean Ig concentration was 0.67 mg ml-1 for trout of about 20 g and 9.1 mg ml-1 for trout weighing more than 125 g.

Animals↗
PubMed1993

Anisakis simplex: antigen recognition and antibody production in experimentally infected mice.

The kinetics of antibody response to intraperitoneal infection of mice with third stage larvae of Anisakis simplex was investigated by ELISA. Maximum antibody response to excretion-secretion (ES) antigens was reached before maximum response to somatic (SA) antigens. Total immunoglobulin (Ig) production (consisting mainly of IgM and IgG1 isotypes) was very similar in both cases. Immunoblotting was used to characterize the antigens recognized by the host in the presence or absence of the metabolic products released by the parasite in vivo. Sera from mice infected with live larvae (anti-L3 L serum) and immunized with dead larvae (anti-L3 D serum) recognized a similar pattern of bands in immunoblots of ES and SA antigen preparations. In the latter, however, three bands at 14, 17 and 18 kD were only recognized by the anti-L3 L serum. A possible explanation is that these low molecular weight antigens are ES products released only in vivo. Finally, the immune response in mouse was compared using ELISA and immunoblotting with the response of a human anisakiasis reference serum, and was found to display considerable similarities. This suggests that the mouse may be a useful model for studying the immunobiology of A. simplex in man.

Animals↗
PubMed1991

Anatomical location of phosphorylcholine and other antigens on encysted Trichinella using immunohistochemistry followed by Wheatley's trichrome stain.

This work investigated the location on the parasite of Trichinella antigens recognized by the mouse immune system and the question as to which of them bear the epitope phosphorylcholine (PC). Wheatley's trichrome stain (initially developed for faecal smears) proved to be excellent for visualization of Trichinella structures, enabling four types of stichocyte to be distinguished. By applying this stain on infected muscle sections after immunocytochemistry using (a) anti-PC BH8 monoclonal antibodies, (b) serum from mice that had been infected twice in the presence of 0.05% thiabendazole (to prevent reproduction by adult females) and then bled on day 7 post-reinfection, (c) serum from infected mice that were bled on day 14 postinfection, or (d) serum from infected mice that were bled on day 42 postinfection, we found (1) that PC is an abundant structural epitope on the hypodermis/muscle, genital primordium and intestinal tract but is absent from the cuticle and stichosome; (2) that the principle secretory cells of adult worms are delta- and beta-stichocytes, whereas those of migrating and encysted L1 larvae are alpha-stichocytes; and (3) that Trichinella antigens recognized in the encysted phase of the parasite's life cycle are present in parasitized myofibres in the sarcoplasmic matrix and in the nucleoplasm of hypertrophic nuclei. The significance of these findings is discussed.

Animals↗
PubMed1991

Specific immunosuppression by Trichinella: fine specificity and effect on lymphocyte function in vivo.

Muscle-phase Trichinella larvae depress the immune response of mice to the phosphorylcholine (PC)-bearing Trichinella antigen FCp without affecting responses to other PC-bearing or non-PC antigens. The depressive activity is independent of antigen dose and Trichinella species and, in adoptive cell transfer experiments with lethally irradiated recipient mice, depended on the state of the recipient (infected recipients had a depressed response even a month after their encysted larvae had been killed and regardless of whether the donor had been exposed to FCp) but not on the state of the transferred cells. We conclude that lymphocytes are not permanently altered by the depressive action, that the agent responsible persists in the host at least a month after the death of the encysted Trichinella larvae, and that the alteration does not eliminate lymphocyte immunological memory.

Animals↗
PubMed1988

Immunomodulation by Trichinella spiralis: primary versus secondary response to phosphorylcholine-containing antigens.

The present work was designed to determine in detail the capacity of the nematode Trichinella to modulate the plaque-forming cell (PFC) response of BCF1 mice to the parasite's own antigens. To this end, we studied the PFC responses shown by infected and non-infected BCF1 mice using as the target antigen phosphorylcholine, an epitope which is found in the parasite. From the results presented here, the following conclusions can be drawn: i) Trichinella spiralis is capable of modulating the immunoresponse to thymus-dependent (TD), but not to thymus-independent (TI), parasite antigens; ii) Trichinella spiralis suppresses the PFC response to the parasite-derived TD antigen FCp1 (a particulate antigen containing PC) during the muscle stage of its life cycle, but does not affect the responses to other parasite-derived PC-bearing antigens; this seems to indicate that the suppressive activity exerted by Trichinella is highly specific; iii) anti-PC PFC production in the secondary response was also suppressed by the parasite. Finally, the inability of the FCp1 antigen to induce detectable anti-PC PFC, other than IgM, is discussed.

Animals↗
PubMed1988

The effect of the intestinal worms and migrating L1 larvae of Trichinella spiralis on the production of antiparasitic IgE antibodies.

The effect of the adult worms and migrating L1 larvae of Trichinella spiralis on the production of specific IgE antibodies was determined in BCF1 mice. To achieve this, we combined the effect of two anthelminthics: thiabendazole, to produce chemosterilization of adult females, and napthalophos, to expel adult worms from the intestine of infected mice on the desired day. Our results demonstrate that when the natural route of infection is used the production of IgE antibodies is not dependent on the infection dose or the number of migrating L1 larvae, and that both intestinal worms and migrating L1 larvae contribute to the production of reaginic antibodies. In addition to this, an extended period of antigenic stimulation (10-12 days) is required for the induction of a detectable, specific IgE response by adult worms. Finally, our results seem to indicate that although the effects of adult worms and migratory L1 larvae on the IgE production are not additive, the presence of adult worms in the intestine of mice may stimulate a secondary exposure to common antigens released by the migrating L1 larvae.

Animals↗
PubMed1988

Allergenic cross-reactivity of several strains of Trichinella in mice and rats by the passive cutaneous anaphylaxis technique.

The cross-reactivity of antiparasite IgE antibody responses induced by allergenic antigens obtained from Trichinella spiralis and Trichinella pseudospiralis was determined by the passive cutaneous anaphylaxis (PCA) technique in BCF1 mice infected with T. spiralis, Trichinella nelsoni, Trichinella nativa, and T. pseudospiralis and in rats infected with T. spiralis and T. pseudospiralis. Our results demonstrate that when the antigen used for the PCA challenge is derived from muscular L1 larvae of T. spiralis, high IgE antibody titres can be detected from sera of animals infected with T. spiralis, T. nelsoni, or T. nativa, but not with T. pseudospiralis, during the entire life cycle of the parasite. However, when homologous antigens are used in the PCA test, we obtained comparable values of IgE titres in rats and mice infected with both T. spiralis and T. pseudospiralis strains. These results suggest the existence of a high degree of immunologic identity between the allergenic antigens of T. spiralis, T. nelsoni, and T. nativa strains, but not T. pseudospiralis.

Allergens↗
PubMed1987

The antiphosphorylcholine plaque-forming cell responses induced by the nematode Trichinella in BWF1 mice.

We have tested the ability of the nematode Trichinella to infect young and old (NZB x NZW) F1 (BWF1) mice. We report the capacity of these mice to respond to the parasite antigens containing the epitope phosphorylcholine. The values obtained in adult worm intestinal retention rates and in the number of encysted larvae on the 35th day postinfection showed that the old BWF1 mice are more susceptible than young BWF1 and control (BALB/c x CBA/j)F1 (BCF1) mice to Trichinella infection. However, unlike the BCF1 mice, young BWF1 mice were unable to produce a good anti-phosphorylcholine plaque-forming cell response after the killing of Trichinella larvae by the anthelminthic mebendazole. Old BWF1 mice presented a discrete response which is discussed. Finally, our results seem to indicate that the lack of anti-phosphorylcholine response in young BWF1 mice after mebendazole treatment may be related to the high susceptibility of these mice to the suppressive properties of encysted Trichinella larvae against their own antigens.

Age Factors↗
PubMed1987

Immune response to Trichinella epitopes: the antiphosphorylcholine plaque-forming cell response during the biological cycle.

Phosphorylcholine (PC), an immunodominant component of the cell wall of certain bacteria, fungi and nematodes, is known to induce low anti-PC antibody levels during natural infection by Trichinella spiralis. This article reports a study in which spleen cells from BCF1 mice infected with Trichinella sp. larvae were found to produce large numbers of direct haemolytic plaques in response to PC conjugated to sheep red blood cells (SRBC) after muscle-encysted larvae had been killed by treatment with mebendazole. Inhibition of the response by PC-chloride, immunodiffusion and immunoelectrophoretic studies with the anti-PC IgA (TEPC-15) and anti-idiotype T15 serum assays showed the plaque-forming cell (PFC) response to be specific for PC. The absence of haemolytic plaques when unconjugated SRBC or TNP-SRBC were used as indicator cells ruled out involvement of a polyclonal response. Greatest anti-PC PFC response was found to be associated with a microsomal fraction designated FCp1, a particulate fraction behaving as a thymus-dependent antigen. The FCp1 fractions from all four strains of Trichinella employed induced anti-PC PFC responses when injected into mice. These results suggest that FCp1 is a suitable antigen for use in detailed studies of immune responses to Trichinella and related parasites.

Animals↗