IgG subclasses in parasitic infestations.
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Humans↗
Biomedical subjects
D Catty
Publications and source records attributed to D Catty.
Presence of kappa 2 light chain in normal rabbits and as induced auto anti-allotype antibody in kappa 1 light chain suppressed subjects.
Using an antiserum raised in b(bas)/b(bas)-suppressed rabbits to kappa 2 light chain we have shown that the kappa 2 light chain is an isotype of kappa 1, present in the majority of, if not all, rabbits. It probably exists in at least 2 allelic forms. It is capable of producing a functional antibody molecule (auto anti-b6) and compensates for the absence of kappa 1 light chain in homozygous b6-suppressed rabbits.
Animals↗
Improved methodology for the production of monoclonal antibodies against parasites.
A modification of the standard fusion methodology is described which results in greatly increased yields of monoclonal antibodies against certain organ-specific parasites. Several fusions were carried out using mice infected with Schistosoma mansoni or Nematospiroides dubius, using B lymphocytes harvested from either the spleen or the mesenteric lymph nodes. Results indicated a greatly improved yield of positive clones using the lymph nodes as a source of B cells for fusion. A 7-fold increase in the number of positive clones was seen with N. dubius injections, while S. mansoni fusions showed a 2-fold increase.
Animals↗
Demonstration of the clonality and specificity of an auto-antibody response to a suppressed immunoglobulin allotype of the rabbit kappa chain b locus.
A regulatory idiotypic network is proposed to control allotype expression in normal rabbits. We have used suppression of the kappa chain b6 allotype in an attempt to restrict the number of regulatory idiotypes involved in an induced auto anti-allotype response. These auto anti-b6 antibodies were examined by an agarose imprint immuno-fixation IEF technique using iodinated allotype-bearing IgG. All totally b6-suppressed rabbits produced a clonally complex response which was generally spectrotypically unique - thus contradicting previous claims of a dominant common idiotypic pattern. The compensating light chains thus have available as many regulatory V genes as does the kappa 1 light chain. However, the b6/b6 homozygotes breaking b6 suppression produce an auto anti-b6 antibody which does not interact with their "escaping" molecules and which is clonally restricted. We propose a regulatory mechanism limiting the V genes utilised to produce the autoantibody, the latter then allowing only molecules bearing non-interactive allotopes to "escape" suppression.
Animals↗
A secondary immune deficiency in the Fatty/Orl-op rat.
The response of T and B lymphocytes to the mitogens PHA and LPS was studied in the Fatty/Orl-op rat. Whereas T and B cells from op/op spleen showed no lack of responsiveness compared with that of normal (op/+) littermate rats, the response of thymic cells to the T cell mitogen was lower in op/op rats over 12 days of age compared with normal rats of the same age. Osteopetrotic rats of 10 days and younger did not show this T cell deficiency. The responsiveness of T cells from op/op lymph nodes to PHA was less than that of normal rats at 12 days and older. Thus, though splenic T and B cell populations were well maintained in the adult op/op rat despite the severe depletion of marrow cells in this mutant, the thymus and lymph node T cell populations were qualitatively or quantitatively deficient. Since osteopetrotic obliteration of the marrow cavities precedes the appearance of the immune deficiency we suggest that the immune deficiency may be caused by a failure of T cell supply to the thymus and lymph nodes.
Animals↗
Quantitative and spectrotypic analysis of paternal IgG2a expression in normal and allotype-suppressed mice.
The synthesis and clonal diversity of IgG2a molecules bearing the paternally inherited immunoglobulin allotype have been examined in the offspring of matings between BALB/c mothers (Igh-1a) and SJL or C57BL/10 males (both Igh-1b) using a sensitive quantitative single radial immunodiffusion in gel assay and isoelectric focusing with autoradiography. In normal litters, the first detectable paternally-marked IgG2a is extensively polyclonal in both F1 crosses (i.e. diversity precedes expression); however, there is a delay of 2-3 weeks in the first appearance of the clonally diverse set of molecules when these are coded by the SJL genome, compared with the C57BL/10. Delayed maturation of allelically-excluded Igh-1b-expressing B cells in the (BALB/c X SJL)F1 may explain the unique susceptibility of these offspring to chronic allotype suppression when exposed to maternal anti-Igh-1b antibodies in early life. We find that, although such suppressed mice may begin life with a (delayed) synthesis of polyclonal IgG2a of paternal allele (Igh-1b), the condition of chronic suppression later imposed in the majority of mice is associated with spectrotype (clonal) simplicity.
Animals↗
Clonal diversity and homology of latent and nominal group a immunoglobulin allotypes in the rabbit.
Latent VH immunoglobulin allotypes are expressed unexpectedly and transiently at low concn in the serum of rabbits. Latent group a1 molecules in sera from rabbit colonies in Philadelphia (U.S.A.) and Birmingham (U.K.) were examined for a1 specificity and clonal diversity using reference nominal allotypic reagents and isoelectric focusing (IEF) autoradiography. Latent a1 molecules from rabbits of both colonies had diverse spectrotypic patterns in the pI range 5.5-8.3, as identified with 125I-labelled affinity-purified specificity-tested, anti-nominal a1 antibody. Comparisons of spectrotypes between nominal a1 antigen and latent a1 focused molecules revealed a marked correspondence in banding over the pI range. Reference anti-nominal a1 antibodies could be absorbed out substantially by the IgG fraction of serum from two rabbits containing latent a1 molecules; in a reciprocal fashion absorption with nominal a1 molecules reduced the binding of focused latent a1. The latent a1 molecules from both U.S.- and U.K.-bred rabbits displayed strikingly similar IEF spectra and their antigenic similarities were confirmed by similar absorption capacities of the reference anti-a1 serum. When sequential serum samples from one (U.S.) latent a1 rabbit were compared by IEF, some bands, e.g. those between pH 7.75 and 8.3, appeared to fluctuate in their presence, whereas others, e.g. between pH 5.3 and 7.4, were expressed continuously. We can conclude that latent a1 molecules are clonally complex and some are consistently produced in small amounts. As they also show antigenic similarity, if not identity with nominal a1, we believe that they are probably the product of the same gene (or genes) with an equivalent capacity to be associated with specificity-determining genes even though the level of synthetic activity is lower and possibly governed differently. Anti-a1 antibody was raised in a rabbit in which latent a1 allotype had been previously detected. This antibody was of low avidity and, while inhibitable on RIA by nominal a1 it was not inhibitable by the donor's latent a1 or by a second latent a1 of the same (partially inbred) U.K. colony, but was inhibitable by a latent a1 serum from the Philadelphia, U.S.A. colony. This result suggests that a1 molecules are the products of more than one gene.
Animals↗
Spectrotypic analysis of passively acquired and newly synthesised IgG antibodies in the neonatal and young mouse.
Isoelectric focussing with autoradiography has been used to analyse the selective nature of passive transmission of specific anti-Ig allotypic antibodies from the mother to the young mouse, and to study the generation of spectrotypic (clonal) diversity of autologous IgG2a (carrying the paternal inherited Igh-1b allotype) in BALB/c X SJL/J F1 (Igh- 1ab heterozygote) mice. Transmission of anti-allotypic antibody to neonatal mice was found to be pI restricted, with selection favouring electrophoretically fast IgG. Comparison of the antibody spectrotype in maternal serum, milk and neonatal serum revealed that the pI restriction in transmission operates at the level of the neonatal gut. Analysis of the paternally inherited Igh-1b IgG2a molecules as they are first synthesised and secreted into the neonatal serum revealed an extensive polyclonality on first detection by the very sensitive focussing assay. It can be deduced that IgV region diversity is generated by IgG2a-synthesising cells prior to, or at the time of, the first secretion of this class of antibody into the serum.
Aging↗
Transmission of immunoglobulin to foetal and neonatal mice.
Transmission of immunoglobulin (Ig) classes and subclasses from mother to foetus and to neonate, and the survival of maternal Ig in the circulation of the young mouse up to 40 days after birth, has been quantitated in Balb/c homozygous and (Balb/c X SJL/J)F1 matings using isotype-specific heteroantisera in radial immunodiffusion in gel assays. The transfer of anti-allotype (anti-Ig-1b (gamma 2a] antibodies from immunised Balb/c mothers (Ig-1a) to F1 heterozygote (Ig-1ab) offspring was measured by passive haemagglutination of Ig-1b target allotype-coated sheep red blood cells. A small but significant level of transmission of Ig to the foetus occurs by the 15th day of gestation (5 days before birth) but the bulk of passively acquired Ig is derived from the milk after birth. All Ig acquired in utero and later across the intestinal barrier is exclusively of IgG isotypes (gamma 1, gamma 2a, gamma 2b) even though the milk has a large predominance of IgA. An appreciable level of maternally derived antibody is maintained in the circulation of the young mouse 24 days or more after gut 'closure' on the 16th day post-partum.
Amniotic Fluid↗
The measurement of antigens released by radiation-attenuated Trichinella spiralis larvae.
A radioimmunoassay has been developed that uses antisera raised to different excretory-secretory antigens of infective larvae of Trichinella spiralis (LESA) to measure accurately the output of these antigens following gamma irradiation at doses from 10 to 120 Krads. In the lower range (up to 20 Krads) irradiation results in the increased export of antigens to the culture supernatant in a subsequent 3 h period, without obvious or gross damage to the worms. Higher doses (greater than 40 Krads) suppress antigen release over the same period compared with the activity of untreated (control) cultures. This work makes two contributions. It describes a sensitive assay system which detects and measures parasite antigens that may be important both in protection and in serodiagnosis, and it offers for the first time an explanation for the special properties of the lower dose range larval irradiation-attenuated vaccine in inducing a high degree of reinfection resistance, as reported in older literature and recently confirmed by us.
Animals↗
The use of cellulose carbonate-based immunoadsorbents in the isolation of minor allotypic components of rabbit immunoglobulin populations.
Cellulose trans-2,3-carbonate has been used as a new insoluble matrix for the simple coupling of a1- and b4-positive rabbit immunoglobulin to make immunoadsorbents capable of purifying from serum, with great efficiency, alloantibodies to these allotypic determinants. The antibodies have themselves been conjugated to prepare specific antibody immunoadsorbents of high binding activity for their allotypic target molecules. With these anti-allotypic solid-phase reagents it has been possible to affinity purify a1- and b4-positive immunoglobulin molecules and to deplete serum immunoglobulin of these molecules to leave in the eluates only the allotypically uncontaminated minor immunoglobulin components which are a-negative or b-negative (lambda chain-bearing) molecules. lambda chain molecules were also purified in very small quantities by affinity chromatography on a sheep anti-rabbit lambda chain column. This method of purifying minor populations of rabbit immunoglobulin from normal serum by special immunoadsorbent applications offers new opportunities to study the products of rarely expressed immunoglobulin genes in normal rabbits.
Animals↗
Early precursors of B lymphocytes. I. Rabbit/mouse species differences in the physical properties and surface phenotype of pre-B cells, and in the maturation sequence of early B cells.
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Animals↗
Early precursors of B lymphocytes. II. Exploitation of the singular properties of rabbit pre-B cells in their purification.
Rabbit pre-B cells are almost uniformly large and low in density and bear a modest number of Fc receptors (McElroy, P.J., Willcox, N. and Catty, D., Eur. J. Immunol. 1981. 11:76). A simple density gradient centrifugation procedure consistently and rapidly enriches them to 20-35% frequencies. At the same time, it depletes them of most of the B cells, phagocytic and erythroid cells that are also common in bone marrow, and are of progressively higher densities. The low-density fractions include 45-55% of large blast-like cells and have 35-45% of cells with Fc receptors. By isolating the large Fc receptor-bearing cells from low-density fractions of baby rabbit bone marrow on a fluorescence-activated cell sorter, pre-B cells were obtained at approximately 60% frequency and virtually uncontaminated by B cells (or plasma cells). This level of purity is perhaps adequate for the critical experiments necessary to establish the precursor status of pre-B cells definitively, and it might also prove valuable for studying the mechanisms of the genetic rearrangements that occur in early B cell development.
Animals↗
Mice treated with strontium 90: an animal model deficient in NK cells.
Treatment of BALB/c mice with radioactive isotopes of the bone-seeking element strontium reduces the percentage of specific NK-cell cytotoxicity to only 2.6%, compared with 13.6% for normal BALB/c and 36.3% for athymic (nude) BALB/c. The syngeneic plasmacytoma NS-1 was used as target in a 4th in vitro NK-cell microassay. Marrow cellularity in treated mice is reduced to 12.5% of controls, but haemopoietic and stem-cell functions are taken over by the spleen and the peripheral blood picture remains relatively normal. Allogenic (H-2k) tumour transplants are rejected normally with good anti-H-2k alloantibody response. Haemopoietic and T- and B-cell functions are therefore substantially intact, and the defect seems confined to NK cells. In vivo, after s.c. inoculation of 10(6) NS-1 cells, 8/12 controls grew a solid tumour after a mean delay of 30.5 +/- 1.25 (s.e.) days, whereas 5/6 90Sr-treated mice grew the tumours after a delay of only 10.5 +/- 1.8 days. This markedly reduced delay in the 90Sr-treated mice lends support to suggestions that NK cells play an important role in resisting the establishment of tumour foci (i.e. in antitumour surveillance). Mice treated with 90Sr could be useful in evaluating the in vitro role of NK cells.
Animals↗
Properties of monoclonal antibodies to human immunoglobulin kappa and lambda chains.
Hybridomas have been produced from mice immunized with human IgG. Culture supernates were assayed for the presence of antibody-producing cells by passive haemagglutination. Hybridomas producing antibodies to human kappa (kappa) and lambda (lambda) light chains have been cloned and grown as ascitic tumours in BALB/c mice. The antigen-binding characteristics of the monoclonal antibodies, contained in the ascitic fluid, were assessed by haemagglutination inhibition, ELISA and radioimmunoassay systems and by the binding of radiolabelled antigen in analytical flat-bed iso-electric focussing gels. One monoclonal anti-kappa reacted better with free than with combined kappa chains; for another the reverse was true. Antibody fractions separated by DEAE chromatography of ascitic fluids were coupled to ox red cells with chromic chloride and compared with polyclonal antibodies for the detection of cell-surface immunoglobulins.
Animals↗
Surface Ig on rabbit lymphocytes. Rabbit B and T cells are distinct populations.
Rabbit peripheral blood lymphocytes (PBL) were analyzed by immunofluorescence using anti-T cell conjugates and anti-Fab, anti-a1 allotype, anti-IgM and anti-IgA conjugates. In addition, T cells were demonstrated by rosetting with papain-treated homologous erythrocytes. Control experiments, using acid treatment and incubation at 37 degrees C for 18 h after or without pronase treatment, revealed the endogenous origin of all surface determinants tested. A good correlation was found between results obtained with the two anti-T cell conjugates used and the T rosette test on PBL and on lymphoid cells isolated from various organs. In lymphocytes isolated from peripheral blood and from various lymphoid organs, the percentages of T and B cells were respectively 45 and 38 for PBL, 10 and 46 for bone marrow, 27 and 31 for appendix, 40 and 45 for spleen, 42 and 46 for Peyer's patches, 96 and 0.3 for thymus and 70 and 16 for peripheral lymph nodes. The percentage of "null" cells in lymphocyte populations derived from bone marrow and appendix is rather high. The final percentages of T and B cells in rabbit PBL depend to a significant extent on the method of isolation, especially isolation by Ficoll-Hypaque centrifugation results in a depletion of T cells. Moreover, a rather impure lymphoid cell suspension is obtained. In double incubation experiments, T cells (as defined by T cell antigen(s) or rosette formation) and B cells (Fab-bearing cells) were entirely different subpopulations. Allotypes of the a locus could not be detected on the surface of T cells. The results are discussed with respect to genetic coding of antigen receptors on B and T cells.
Animals↗
Multiple Ig classes on rabbit B lymphocytes.
Rabbit PBL were studied regarding the presence of different classes of s-Ig, under experimental conditions, ensuring the endogeneous origin of these proteins. About 40% of the lymphocytes are B cells (Fab positive and a1 positive in a1 homozygous rabbits). IgG positive lymphocytes could be found, but only using and anti dll conjugate (allotype located on the Fd gamma region of IgG). Anti Fc gamma conjugates were negative. Most of the B cells are IgM positive, most of these IgM cells however were also positive for either IgG or IgA. Np lymphocytes were found bearing both IgG and IgA. Careful analysis of the percentages of various isotypes found on B cells suggests that some "IgD" positive lymphocytes could be present. Results are discussed in relation to B cell differentiation.
Animals↗
Clonal heterogeneity and other properties of auto anti-allotype antibodies produced in allotype-suppressed rabbits.
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Animals↗