Search PubMed⌕ Search

Biomedical subjects

F Celada

Publications and source records attributed to F Celada.

At least 55 records · Page 3Linked to original sources
PubMed1984

Follow-up of circulating immune complexes in the course of acute viral hepatitis, and correlation with serologically relevant parameters.

Type A, type B and type non-A, non-B hepatitis patients were followed up. Several parameters were checked at ten day intervals. Circulating immune complexes (CIC) were detected in a large percentage of patients by using the PEG test and an assay that makes use of bovine conglutinin (K) as recognition unit, and an enzymatically labelled immune complex as the probe. The decrease in the mean level of CIC in the patients correlated with the decrease in serum transaminases and bilirubinaemia in type A and type B hepatitis. Although the pattern of the mean values of the two assays was similar for type A and type B hepatitis, when the two CIC assays were compared for each patient, no significant correlation was found. In light of these and previous results, the necessity for performing CIC monitoring with more than one assay is also discussed.

Acute Disease↗
PubMed1983

An enzymatically active antigen-antibody probe to measure circulating immune complexes. II. E. coli beta-galactosidase in the probe and C1q as the recognition unit.

An enzymatically active probe (beta-galactosidase-anti-beta-galactosidase complex) is used to measure circulating immune complexes (CIC), in a competition assay where probe and CIC are confronted with a 'recognition unit'. The latter is bovine conglutinin in the original description of this method. Here we describe a version utilizing human or bovine C1q. The two techniques are compared for their sensitivity and specificity, on both in vitro formed tetanus toxoid-anti-toxoid complexes and on sera from patients with selected diseases. The results confirm that the two recognition units are sensitive to families of CIC that only partially overlap. The parallel use of conglutinin and C1q yields both quantitative and qualitative information on the nature of CIC in individual sera.

Animals↗
PubMed1983

Activation of human B lymphocytes: frequency of antigen-specific B cells triggered by alloreactive or by antigen-specific T cell clones.

Human T cell clones specific for tetanus toxoid (TT) or for alloantigens were isolated and expanded in culture using antigen stimulation and soluble growth factors. When stimulated by the specific antigen, the clones were able to proliferate in the absence of exogenous growth factors and to provide help to B cells. The alloreactive and TT-specific clones were compared for their capacity to help an anti-TT as well as a polyclonal antibody response. The frequency of the B cells activated to the production of specific antibody was determined by limiting dilution analysis in cultures containing limiting numbers of responding B cells and optimal numbers of cloned T helper cells. In these conditions a single activated b cell was able to produce about 10 ng of antibody. TT-specific clones, in the presence of low TT concentrations, selectively induced a number of TT-specific B cells to produce IgG antibody in the absence of a detectable polyclonal B cell activation. On the other hand the alloreactive clones activated higher numbers of TT-specific B cells as a part of a strong polyspecific B cell activation. In this case the antibody production did not require the presence of TT in culture; furthermore cells producing antibodies of unrelated specificities were also activated in allostimulated cultures. IgG anti-TT were produced only by immune donors, while IgM anti-TT were produced by immune and nonimmune donors, providing the B cells were activated by alloreactive clones. These data demonstrate that human memory B cells can be triggered by both antigen-specific and alloreactive T cells to antibody production. The alloreactive clones can therefore be used to analyze at the clonal level the repertoire of a pool of B cells that contain most, if not all, the memory antibody specificities.

Antibody Formation↗
PubMed1983

Cold-precipitable immune complexes in collagen diseases: evidence for the coexistence of multiple types of circulating complexes in the same serum.

In patients with systemic lupus erythematosus, mixed cryoglobulinemia, and rheumatoid arthritis, the presence of cold-precipitable immune complexes (IC) was investigated by means of two different methods, i.e., the Clq-binding activity (ClqBA) and a competitive enzyme immunoassay, based on solid-phase bovine conglutinin (K). Cold precipitability of IC ranged between 0 and 100% with K and between 0 and 71% with ClqBA. No correlation existed either between the levels or the cold precipitability of the IC measured by the two systems in the same sera. On the whole, cold-precipitable IC were better determined by the K method than by ClqBA and in mixed cryoglobulinemia cryocrit levels correlated with IC levels determined with K, but not ClqBA. These data provide direct evidence of the coexistence of several types of circulating IC in the same serum and that the two methods recognize, at least in part, different IC in the same specimen. It might be hypothesized that different IC present in a serum may have a distinct biological significance.

Antigen-Antibody Complex↗
PubMed1983

Recognition of donor fibroblast antigens by lymphocytes homing in the human grafted kidney.

A human transplanted kidney, surgically removed because of untreatable chronic rejection, was used as the source of lymphocytes (K-L) of recipient origin that were expanded with interleukin-2 (IL-2), and of kidney fibroblasts (K-F) of donor origin that were maintained as an established line. Cytotoxicity assays were performed using K-L and peripheral blood lymphocytes (PBL) as effectors, and K-F and donor PBL as targets. From the results the following conclusions can be drawn: (1) cytotoxic lymphocytes, presumably involved in the process of chronic graft rejection, home in the kidney (from which they can be recovered) but are not detected in the circulation; (2) cytotoxic lymphocytes can be generated from peripheral lymphocytes by mixed lymphocyte culture (MLC) and further expansion in vitro with IL-2 (MLC-L); and (3) although both K-L and MLC-L are cytotoxic toward K-F, the former are not cytotoxic toward donor PBL. This suggests that although MLC-L recognize antigens shared by K-F and PBL, K-L recognize antigens specific for K-F only. These results, if confirmed, indicate that antigens not present on PBL, and possibly tissue-restricted are important in graft rejection. Thus, while monitoring transplanted patients, a lack of cytotoxicity in the recipient PBL may be misleading because the relevant cytotoxic effector cells may have disappeared from the periphery and the appropriate antigenic target may be absent on donor PBL.

Adolescent↗
PubMed1982

Human T cell lines with antigen specificity and helper activity.

Human T blasts, obtained by stimulation of peripheral blood mononuclear cells (PBM) with tetanus toxoid, diphtheria toxoid or Candida albicans, were expanded in long-term culture using alternate periods of antigen restimulation and growth in media containing interleukin 2. The cells gave a proliferative response only to the antigen originally used for stimulation. Such as response was strictly dependent upon the presence of autologous but not of allogeneic mitomycin C-treated mononuclear cells. When added to autologous PBM depleted of E-rosetting cells together with the specific antigen, the T blasts induced a polyclonal proliferation and differentiation of B cells. Allogeneic B cells were activated by antigen-stimulated T blasts only in the presence of irradiated mononuclear cells autologous to the responding T blasts. The above responses seemed not to be regulated solely by the release of soluble factors; apparently cell to cell interactions had to take place to obtain an efficient B cell activation.

Antibody-Producing Cells↗
PubMed1981

Requirement of the immunogen (E. coli beta-galactosidase) for the response towards a determinant responsible for antibody-mediated enzyme activation, while antibodies binding some other site can be elicited by mitogen alone.

Virgin mouse spleen cells cultured in vitro without antigen or mitogen produced a measurable amount of IgM binding E. coli beta-galactosidase (beta-gal). Lipopolysaccharide (LPS) and LPS plus antigen enhance this response, which cannot be considered truly polyclonal since it does not include the production of antibodies directed towards a conformation-dependent determinant, responsible for the activation of a mutant beta-gal, and known otherwise to be highly immunogenic. By priming in vivo with alum-treated beta-gal (unable to elicit activating antibodies), an activating response is obtained in vitro by LPS plus antigen, but not by LPS alone. These results are compatible with a two-signal requirement for the activation of B cells and may be explained as follows: (a) the mitogen, in absence of immunogen, stimulates those clones which have received a specific signal from cross-reacting structures in the environment: (b) instead, no cross-reaction are available for the conformation-controlled structure of the "activating" determinant; thus, intact immunogen is required as well as mitogen. Because of this "uniqueness", the molecule of beta-gal offers a highly specific tool to probe carrier-hapten relationships in native proteins.

Animals↗
PubMed1981

Antibody-mediated activation of genetically defective Escherichia coli beta-galactosidases by monoclonal antibodies produced by somatic cell hybrids.

Six hybridomas producing monoclonal antibodies against Escherichia coli beta-galactosidase (beta-D-galactoside galoctohydrolase, EC 3.2.1.23) have been derived from two separate somatic cell fusions. Three of these antibodies can activate defective enzymes produced by strains of E. coli carrying Z-gene point mutations. In antigen excess, one monoclonal antibody shows similar enzyme binding and mutant-activating capacity. Characteristically, the former reaction has a 200-fold higher equilibrium constant. These data provide direct evidence that the enzyme-activation reaction is a single-hit event in which one antibody site favors the correct conformation of one active center of the enzyme. Because each "activating" hybridoma is able to activate several but not all point mutant enzymes tested, it appears that the correction of the genetic defect is produced by binding key sites of the protein three-dimensional structure rather than the sites affected by the mutation.

Animals↗
PubMed1981

Patients with high titers of circulating immune complexes are most likely to benefit from plasmapheresis treatment.

Intermittent flow centrifugation system was utilized in the management of 17 patients with IC diseases. 14 out of 14 patients that had high levels of ICs prior to plasmapheresis showed both clinical and immunochemical evidence of improvement with plasmapheresis. 3 patients had no evidence of circulating ICs prior to plasmapheresis: in these cases no apparent result was achieved after the apheretic therapy. Although the striking correlation between IC removal and clinical result is not a formal proof of responsibility of CIC in the pathogenesis of the disease, it is suggested that quantitative determination of CICs (utilizing several different recognition units) may be a useful parameter in the decision to apply plasmapheresis in the therapy of selected autoimmune diseases.

Adolescent↗
PubMed1979

A dimer--dimer binding region in beta-galactosidase.

alpha Complementation in beta-galactosidase is the restoration of enzyme activity by addition of the alpha donor CNBr2, from amino acid residues 3--92 of the polypeptide, to inactive M15 protein from the lacZ deletion mutant strain M15. M15 protein lacks residues 11--41 and is a dimer; the active complex, like native beta-galactosidase, is tetrameric [Langley, K. E., & Zabin, I. (1976) Biochemistry 15, 4866--4875]. A dimer--dimer binding region in beta-galactosidase has been identified by proteolytic and immunologic studies of alpha-complementation. Proteolytic experiments were carried out with trypsin. Treatment of native beta-galactosidase with trypsin, followed by reaction of the mixture with cyanogen bromide, yields intact CNBr2 as measured by its ability to complement M15 protein. Active CNBr2 is not obtained when urea-denatured beta-galactosidase is treated in the same way. Therefore the segment corresponding to CNBr2 is apparently buried within the folded protein. Immunologic experiments were carried out with antibodies against CNBr2, tryptic peptide T8 (residues 60--140), and CNBr3 (residues 93--187). Anti-CNBr2 and anti-T8 bind to M15 protein but not to beta-galactosidase, indicating that this area is exposed in the dimer. Anti CNBr2, but not anti-T8 or anti-CNBr3, inhibits the formation of alpha-complemented enzyme. These results indicate that an early part of the sequence, within the segment corresponding to CNBr2, is involved in dimer--dimer interaction.

Antibodies↗
PubMed1978

Probes of beta-galactosidase structure with antibodies. Reaction of anti-peptide antibodies against native enzyme.

Antibodies were prepared against 18 tryptic and cyanogen bromide peptides from beta-galactosidase ranging in size from 15 to 96 amino acid residues representing more than 80% of the polypeptide chain. They were tested for binding capacity and affinity toward their homologous antigens and toward the whole native protein. Nine antisera bound to beta-galactosidase; these had been raised against certain peptides from the central and carboxyl-terminal regions of the poly-peptide chain. Based on these results a preliminary model of the three-dimensional structure of the folded protein is suggested.

Antibodies↗
PubMed1978

Immune response against the beta-galactosidase enzyme of E. coli at precursor cell level. I. Analysis of the secondary repertoire in BALB/c mice.

The BALB/c secondary response against the beta-galactosidase (beta-gal) enzyme of E. Coli was analyzed at the precursor cell level by using the splenic focus technique. Our results indicate that in immunized mice, one out of 18 000 B cells is able to recognize beta-gal. Among the families of anti-beta-gal monoclonal antibodies, a subset of specific antibodies was detected which is capable of protecting the enzyme from heat denaturation. The frequency of clones making protecting antibodies is 1 out of 90 000 and appears to be fairly constant among different individual mice. Further, the degree of heterogeneity of protecting antibodies analyzed in one individual is very high (250-fold difference in affinity) but comparable to other secondary repertoires. Specific frequencies are compared with previous findings relative to secondary responses against artificial haptens. It is suggested that a different type of recognition exists between protein determinants and artificial haptens. In addition, the relatively high proportion of clones making antibodies of the protecting type suggests that only a small proportion of antigenic sites on the beta-gal is actually able to stimulate an immune response.

Animals↗
PubMed1978

Long-term antibody synthesis in vitro. VI. Anti-allotype sera as probes of clonal products in affinity maturation.

A new experimental system is described for measuring the allotypic product of rabbit B cells during long-lasting in vitro antibody responses. The immunoenzymatic assays described allow determination of several parameters mapping in different regions of the same molecule, which can be measured and combined to yield a multidimensional picture of the time-course dynamics of antibody synthesis. The rabbit immune system responding to Escherichia coli beta-D-galactosidase was sample and disassembled by (a) culturing lymph node microfragments and (b) sorting out from among all anti-enzyme antibodies only those activating a mutant enzyme, AMEF, which bore the b4 or b9 allotype. A considerable simplification of the response was achieved in the microcultures as documented by cultures of heterozygous cells which produced only one allotype and by the fact that each culture showed a distinctive pattern when antibody titre, association constant, heterogeneity index, L-chain type, and k-chain allotype were considered together. This array of patterns was not an artifact but the result of disassembling a representative sample of the rabbit immune system into small components, since the b4/b9 ratio obtained by averaging the results of all cultures from a heterozygous rabbit lymph node was the same as the serum ratio. Despite the Poisson distribution of the responder microcultures, none of them was monoclonal; i.e. no antibodies homogeneous by all parameters tested were observed, This finidng supports the notion that in normal lymphoid tissue in its native tridimensional arrangement, one T cell can trigger several B cells clustered in one antibody-forming unit. This natural arrangement would ensure the monospecificity of the cluster (dictated by the T cell) while allowing for variation in affinity (depending upon the array of B cells in the unit). Accordingly our findings would results from the fact that as the size of the microfragments was reduced, the cells diluted out first were T cells, but as long as one of them was present, several B-cell clones were triggered. The b4/b9 pattern of any given culture remained constant over several months, but the ratio kappa/lambda underwent changes. An increase in molecules with non kappa-chains (which could not be reacted with anti-kappa-chain allotype antisera) was usually associated with a parallel decrease in antibody affinity. This occurred by the end of the antibody cycle and might be related to the regulation of antibody synthesis by T-cell suppressor factors.

Animals↗
PubMed1976

Affinity and specificity of penicillin-antibody interaction determined by an enzyme (E. coli beta-galactosidase) immunoassay.

An enzyme immunoassay (EIA) for a penicillin derivative is described with a sensitivity at least at the nanogram level. The label, E. coli beta-galactosidase is a macromolecule of 540 000 daltons: the size of the enzyme and the ease of linking penicilloyl residues to it make it an interesting model to study the effect of the degree of haptenic substitution (DS) in the tracer on the parameters of EIA. Our results show that the affinity of the binding reaction between antibody and tracer is proportional to the DS but the sensitivity of inhibition is not affected, at least not between 1 and 10 penicilloyl residues per GZ molecule. The theoretical consequences and practical applications of multivalent tracers in EIA are discussed.

Antigen-Antibody Reactions↗