Heterologous Antisera Research Articles

Increased expression of monocyte chemoattractant protein-1 in anti-thymocyte antibody-induced glomerulonephritis

The infiltration of monocytes-macrophages in the glomerulus is one of the hallmarks of glomerulonephritis and may play an important pathogenetic role. Monocyte chemoattractant protein-1 (MCP-1) and colony stimulating factor-1 (CSF-1) are monocyte-specific cytokines with chemoattractant and activating activities for monocytes. MCP-1 and CSF-1 can be generated by several cell types, including glomerular mesangial cells, and can be stimulated by cytokines and immune complexes. To study the expression of CSF-1 and MCP-1 in a model of proliferative glomerulonephritis we used Northern blot analysis and immuno-histochemistry. The glomerular lesion was induced in rats by the i.v. injection of a heterologous anti-thymocyte antiserum (ATS), directed against an antigen which is localized on glomerular mesangial cells. Northern blot analysis revealed comparable amounts of CSF-1 in glomeruli isolated from control untreated rats, and from rats after 30 minutes to three weeks of injection of ATS antibody. In control glomeruli no mRNA levels for MCP-1 were detectable, but increased markedly 30 minutes after the induction of the nephritis, were then reduced at 24 hours and increased again at 5 and 21 days after induction of the disease. The increase in mRNA levels for MCP-1 30 minutes or 24 hours after ATS injection was markedly attenuated if rats were complement depleted by cobra venom injection. These time points following antibody injection were associated with mesangial immune complex formation (30 min), mesangiolysis (24 hr) and proliferative glomerulonephritis (5 and 21 days). By immunohistology the presence of MCP-1 was demonstrated in glomeruli with a predominant mesangial distribution. The mesangial immunofluorescence for MCP-1 followed a pattern similar to that of the mRNA for MCP-1 after induction of the disease process, that is, it increased after 30 minutes, decreased after 24 hours and was increased again at three weeks. Within 30 minutes of the antibody injection an increased infiltration of monocytes-macrophages was observed in the glomeruli, which was maintained up to three weeks of induction of the glomerulonephritis. When the rats were decomplemented with cobra venom factor prior to the i.v. injection of ATS, the expression of MCP-1 in glomeruli remained low and the influx of monocytes/macrophages did not appear. We conclude that MCP-1 is increased early on in glomeruli of rats with immune-mediated mesangial proliferative glomerulonephritis. This increase is mediated by complement activation secondary to the in situ immune complex formation at the glomerular mesangium.(ABSTRACT TRUNCATED AT 400 WORDS)

Biogenesis of petunia and carnation corolla chloroplasts: changes in the abundance of nuclear and plastid‐encoded photosynthesis‐specific gene products during flower development

To study the biogenesis of the photosynthetic apparatus in corollas, as well as compositional differences between corolla and leaf chloroplasts, the levels of 12 nuclear‐ and plastid‐encoded thylakoid and stromal components were analyzed by western blotting using heterologous antisera. Relative levels of the thylakoid polypeptides analyzed in petunia (Petunia hybrida cv. Hit Parade Rosa) and carnation (Dianthus caryophyllus cv. White Sim) corollas increased, per unit chlorophyll, in parallel to corolla development, peaking at the mature stage in both flower types, with the exception of subunit V of the photosystem I (PSI) core complex, which continued to accumulate even after anthesis. The photosystem II (PSII) major light‐harvesting chlorophyll a/b protein accumulated in corollas to a level, per unit chlorophyll, similar to that in leaves in both petunia and carnation plants. Components of the cytochrome b6/f complex were found to accumulate to higher levels in corolla chloroplasts than in leaves. The opposite trend was found for components of the PSI core complex, as well as for the stromally located small and large subunits of ribulose‐1,5‐bisphosphate carboxylase, which accumulated in corollas to levels ca 2. 5 times lower than their respective levels in leaves. The latter subunits accumulated coordinately in corollas of both plant types during flower development. Data obtained from immunological studies were correlated with those at the mRNA level. Nothern blotting revealed that, in petunia corollas, the steady‐state transcript level of genes coding for the small subunit of ribulose‐l,5‐bisphosphate carboxylase was identical to that of genes coding for the major light‐harvesting chlorophyll a/b protein. Levels of both transcripts, as well as those of plastid‐encoded genes for the PSII reaction center's Dl and D2, were ca two‐thirds of their respective levels in leaves. In contrast, the level of transcript for the large subunit of ribulose‐1,5‐bisphosphate carboxylase was reduced 5‐fold in petunia corollas, as compared to leaves.

Immunocytochemical localization of photosystem I and the fucoxanthin-chlorophylla/c light-harvesting complex in the diatomPhaeodactylum tricornutum

iserum against two polypeptides of the major fucoxanthin-chlorophylla/c light-harvesting complex of the diatomPhaeodactylum tricornutum and heterologous antiserum against purified photosystem I particles of maize were used to localize these two complexes on the thylakoid membranes ofP. tricornutum. As in many chromophyte algae, the thylakoids are loosely appressed and organized into extended bands of three, giving a ratio of 2∶1 for appressed versus non-appressed membranes. Immunoelectron microscopy demonstrated that the fucoxanthin-chlorophylla/c light-harvesting complex, which is believed to be associated with photosystem II, was equally distributed on the appressed and non-appressed thylakoid membranes. Photosystem I was also found on both types of membranes, but was slightly more concentrated on the two outer non-appressed membranes of each band. Similarly, photosystem I activity, as measured by the photooxidation of 3,3′-diaminobenzidine, was higher in the outer thylakoids than in the central thylakoid of each band. We conclude that the thylakoids of diatoms differ from those of green algae and higher plants in their macromolecular organization as well as in their morphological arrangement.

Immune-activation in recurrent oral ulcers (ROU).

Tissue specimens from eight patients with recurrent oral ulcers (ROU) were analyzed for possible signs of active lymphocyte involvement. A total of 13 specimens were analyzed, eight (8) taken at the ulcer site and five (5) from clinically unaffected mucosa at a site opposite the ulcer. Monoclonal antibodies or heterologous antisera were applied using the avidin-biotin-peroxidase complex (ABC) or peroxidase-antiperoxidase complex (PAP) methods to visualize cell-activation-associated marker proteins. In specimens from the ulcer area, approximately 43 +/- 18% of all inflammatory cells were positive for the MHC locus II coded Ia antigen. Furthermore, markers for cycling cells, interleukin-2 (CD25, 13 +/- 6%) and transferrin (CD71, 23 +/- 14%) receptors, were frequent in the specimens studied. Staining for CD1 (5 +/- 2%) disclosed dendritic intraepithelial cells in diseased and in clinically unaffected mucosa. Mobilization of such cells is suggested by their presence in submucosa in ROU specimens, but not in clinically unaffected mucosa. The presence of CD1 cells, presumably denoting their identity as potent antigen-presenting Lagerhans' cells, and the rich presence of Ia suggest that local conditions are favorable for induction of T-cell-mediated responses. The simultaneous presence in such infiltrates of activation marker positive T-cells suggests activation de facto. This together with the rarity of activated B-cells, i.e. plasmablasts/cells containing cytoplasmic immunoglobulin, suggests active involvement of the local cells of the T-lymphocyte lineage in the pathogenesis of ROU.

Specificity of polyclonal antibodies to different antigenic preparations of Pseudomonas syringae pv. pisi strain UQM551 and Pseudomonas syringae pv. syringae strain L

Polyclonal antibodies were produced against sonicated and heat‐killed cells of Pseudomonas syringae pv. pisi strain UQM551 and Pseudomonas syringae pv. syringae strain L, and their specificities were compared. Evidence is presented that the serological specificity between these two pathovars lies in surface antigens. Of the surface antigens purified and tested, only flagella and lipopolysaccharide from the cell wall showed no cross‐reactivity with heterologous antisera. Antisera to glutaraldehyde‐fixed flagella of the two strains showed a high level of specificity. At a species or genus level, antisera prepared from heat‐killed cells ofP. syringae distinguished this species from all other bacterial species and genera tested, including strains of Pseudomonas fluorescens, Escherichia coli, Agrobacterium and Rhizobium.

Comparison of in vitro translation products of Sarcocystis gigantea and Sarcocystis tenella

RNA was purified from cystozoites of Sarcocystis gigantea and Sarcocystis tenella and used to in vitro translate polypeptides in a wheat germ and a rabbit reticulocyte translation system. The in vitro translated polypeptides were compared by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The S. tenella mRNA translated at least two polypeptides (mol. wt about 80,000 and 21,500) in both translation systems that were not translated by the S. gigantea mRNA. To study co-translational and initial post-translational processing in Sarcocystis, the poly(A) + RNA preparations were in vitro translated in the rabbit reticulocyte translation system in the presence or absence of canine microsomal membranes. Based on electrophoresis, there appeared to be modification of at least some Sarcocystis polypeptides in the mol. wt range 17,000–30,000. In addition, the translation products were immunoprecipitated with a homologous and a heterologous antiserum. The immunoprecipitated polypeptides were compared by electrophoresis and the S. tenella translation products contained at least one unique antigenic polypeptide with a mol. wt of about 34,700 that was not processed by the microsomal membranes. These results suggest that there is at least one polypeptide that is a candidate for use as an antigen for the differentiation of S. gigantea and S. tenella infections in sheep.

Purification of the DNA-dependent RNA polymerase from the myxobacterium Stigmatella aurantiaca.

The DNA-dependent RNA polymerase (EC 2.7.7.6) of the myxobacterium Stigmatella aurantiaca has been purified. It shows three main polypeptide bands with apparent molecular weights of 146,000, 105,000, and 40,000 in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. beta and beta' subunits of the S. aurantiaca polymerase were shown to migrate in the 146,000-molecular-weight polypeptide band and the main sigma factor was shown to migrate in the 105,000-molecular-weight band by using heterologous antisera.

Characterization of the structural proteins of hemorrhagic enteritis virus.

The structural proteins of hemorrhagic enteritis (HEV), a turkey adenovirus, were analyzed by polyacrylamide gel electrophoresis (PAGE) and Western blotting using polyspecific, monospecific and monoclonal antibodies for detection. In purified HEV preparations, eleven polypeptides with apparent molecular weights ranging from 96,000 to 9,500 (96k to 9.5k), were specifically recognized by convalescent turkey serum. Six of these polypeptides were further characterized by PAGE, Western blotting, ELISA, sucrose gradient centrifugation and electron microscopy. The 96k polypeptide was identified as the hexon polypeptide which is a monomer of the major outer capsid or hexon protein. The 51/52k and 29k polypeptides, identified as the penton base and fiber polypeptides respectively, were the components of the vertex or penton protein. The 57k polypeptide was identified as a homologue of the human adenovirus type 2 (Ad 2) IIIa protein with which it shares a common epitope. Two core proteins with molecular weights of 12.5 and 9.5k were present in purified HEV nucleoprotein cores. The proteins of two HEV isolates, one apathogenic (HEV-A) and one virulent (HEV-V), resembled each other in most respects. However, differences between HEV-A and HEV-V were found in electrophoretic migration of the penton base protein both under native and denatured conditions, and in the electrophoretic migration of the 43/44k polypeptide. Moreover, homologous antiserum against the fiber protein reacted stronger than heterologous antiserum in an ELISA. Single fibers were detected by electron microscopy attached to the penton base proteins of HEV virions and in isolated pentons. The feature of having single fibers is shared with the mammalian adenoviruses and the avian egg drop syndrome 1976 virus (EDS 76 V), but not with the fowl adenoviruses which have double fibers attached to their penton base proteins.

Inhibitory effect of NZ-107 on anaphylactic bronchoconstriction in guinea pigs and rats.

We studied the effect of NZ-107 in a number of animal models of anaphylactic bronchoconstriction. In conscious guinea pigs, pretreated with indomethacin, pyrilamine and propranolol, passively sensitized with heterologous anti serum, NZ-107 in doses of 10-30 mg/kg per os inhibited the aerosolized antigen-induced cough and collapse. NZ-107 in a high dose of 100 mg/kg per os significantly prevented aerosolized antigen-induced anaphylactic collapse, but not cough in actively or passively sensitized conscious guinea pigs and also significantly protected aerosolized histamine-induced collapse, but not cough in conscious guinea pigs. This compound had little inhibitory effect on aerosolized acetylcholine-induced cough and collapse. In anesthetized animals, the effect of NZ-107 on bronchoconstriction induced by intravenous administration of antigen and various agonists was examined by the method of Konzett and Rössler. In doses of 10-50 mg/kg per os, NZ-107 inhibited antigen-induced bronchoconstriction in anesthetized guinea pigs. NZ-107 when intravenously administered to the anesthetized guinea pigs inhibited not only leukotriene D4-induced bronchoconstriction, but also thromboxane A2 mimetic U-46619-, platelet-activating factor- and histamine-induced bronchoconstriction. In anesthetized rats, NZ-107 in a dose of 300 mg/kg per os tended to inhibit the antigen-induced bronchoconstriction, but this effect was not significant. These results indicate that NZ-107 acts as a spasmolytic agent which inhibits bronchial responses to antigens or various other bronchoconstrictors in animal models, suggesting that NZ-107 may be potentially beneficial in the treatment of bronchial asthma.

Intracerebroventricular immunization with TRH-antiserum lowers blood pressure in spontaneously hypertensive rats.

Centrally administered thyrotropin-releasing hormone exerts a well documented hypertensive effect. In this study, the possible physiological role of thyrotropin-releasing hormone in the central cardiovascular regulation was evaluated in spontaneously hypertensive rats receiving long-term (8-14 days) intracerebroventricular infusion of a heterologous antiserum to thyrotropin-releasing hormone. The effect of this passive immunization on the blood pressure was monitored from conscious animals by the tail-cuff method. Thyrotropin-releasing hormone antiserum significantly decreased the systolic arterial pressure in adult rats with established hypertension. No alterations in serum thyroid hormone status were observed indicating that the antihypertensive effect of immunological blockade of thyrotropin-releasing hormone was not due to changes in the serum thyroid hormone levels. These results provide evidence for a role of endogenous brain thyrotropin-releasing hormone in the maintenance of hypertension in spontaneously hypertensive rats.

Immunochemical and immunohistochemical studies of cadmium associated proteins in Rana tadpoles

Previous observations suggested that Rana tadpoles treated with aqueous cadmium (Cd) accumulate Cd in their liver and mesonephros. In order to study the response to Cd in these tissues we (a) exposed tadpoles in mid-limb bud stages to sublethal quantities of Cd, (b) isolated Cd-associated protein (CAP) from a liver cytosol fraction, (c) prepared a heterologous rabbit antiserum against glutaraldehyde-treated CAP (G-CAP), (d) used the rabbit anti-G-CAP antiserum in order to assess the tissue distribution of CAP in Cd-treated and untreated tadpoles, and (e) assessed species crossreactivities of our anti-G-CAP with CAPs and metallothioneins (MTs) isolated from Cd-treated vertebrate liver cytosol fractions. We found that (a) CAP was present in higher quantities in liver cytosol obtained from Cd-treated tadpoles compared to liver cytosol obtained from untreated control tadpoles, (b) indirect immunofluorescent analysis revealed that CAP was localized in liver hepatocytes and kidney tubule epithelial cells in Cd-treated tadpoles, and (c) the anti-G-CAP crossreacted with rodent and fish CAP. These observations suggest that the developing liver and mesonephros are involved in responses to toxic metals and that our anti G-CAP antiserum may be used to gauge exposure to environmental Cd.

Immunohistochemistry of the pituitary pars distalis of the musk shrew, Suncus murinus

Immunocharacteristics of the pituitary pars distalis cell types of the musk shrew, Suncus murinus, were studied by the unlabeled antibody enzyme technique, using peroxidaseantiperoxidase or avidin-biotin-peroxidase complex. The thyrotropin (TSH)-, gonadotropin (GTH)-, corticotropin (ACTH)-, prolactin (PRL)-, and growth hormone (GH)-secreting cells of the PD were identified on the basis of their immunoreactivity with different heterologous antisera. The TSH cells showed specific immunoreactivity with antisera against human (h) TSHβ and rat (r) TSHβ. Cells showing immunoreactivity with the antisera against hLHβ and ovine (o) LHβ were designated as GTH cells as no immunoreactivity was observed with antisera against hFSHβ and oFSHβ. The ACTH cells as well as the cells of the pars intermedia were revealed by anti-ACTH 1–24 and anti-ACTH 1–10 sera. Whereas the PRL cells were recognized by their immunoreactivity with antisera against hPRL and oPRL, the GH cells were identified with anti-hGH, anti-oGH, and anti-bovine (b) GH sera. TSH and GTH, TSH and ACTH, GTH and ACTH, ACTH and GH, ACTH and PRL, and GH and PRL cells were visualized in the same section using the dual immunoperoxidase technique. Comparison of the immunohistochemically identified cells with those described histochemically reveals several discrepancies, which expose the limitations of the latter techniques identifying adenohypophysial cells.

Discrimination of cytoskeletal elements of Paramecium by heterologous antisera: A preliminary investigation on the presence of intermediate filament-related protein

The present study of the cytoskeleton of Paramecium employed a number of heterologous monoclonal antisera raised against intermediate filament proteins of higher organisms, and also an antiserum against a 49 K protein from Tetrahymena. Anti-49 K, and also monoclonal anti-bovine cytokeratin K 8.13 which has a broad cross-reaction spectrum for epithelial intermediate filaments, differentiated the microtubular systems of Paramecium into two categories. Those closely associated with surface membranes, such as basal bodies, contractile vacuole pores and the cytospindle, and the gullet also, were labeled by these antibodies. Others internal to the epiplasm are not labeled, such as transverse and postciliary microtubular bands associating with basal bodies, contractile vacuole canals, cytopharyngeal ribbons and postoral fibers. None of the heterologous anti-tubulins tested in this and previous studies generated such a pattern of discrimination of the microtubular systems of Paramecium, indicating that the discrimination by anti-49 K and by anti-cytokeratin K 8.13 was not based on the molecular composition (α- or β-tubulin) of the microtubular systems, their state of tyrosination, or acetylation. These two antibodies thus reveal a determinant that is associated exclusively with certain membrane-bound microtubular systems of Paramecium. This determinant appears to participate in the development of nascent structures like the cytospindle and the oral apparatus, since it co-distributed with these structures in their different stages of development. In addition, two monoclonal antibodies were superimposable specifically on certain cytoskeletal elements also residing close to surface membranes: anti-pig vimentin V 9 labeled the epiplasm underlying the inner alveolar membrane, while anti-human cytokeratin K 4.62 labeled the outer lattice lining the cortical ridges. On the other hand, anti-49 K labeled the micronucleus and its derivatives in all stages of sexual nuclear reorganization in Paramecium, unlike the specific labeling of postmeiotic and zygotic nuclei in Tetrahymena. The presence of intermediate filament-related protein in Paramecium is discussed.

Antigenic shifts in serotype determinants of Campylobacter coli are accompanied by changes in the chromosomal DNA restriction endonuclease digestion pattern.

Changes in somatic (O) lipopolysaccharide (LPS) antigenic specificities of Campylobacter coli serostrains were observed after continuous laboratory subculture. Two serostrains (C. coli O34 and C. coli O48) lost O specificity and did not react with homologous or any of the available heterologous antisera. The C. coli serostrain for serogroup O5, after subculture, yielded a variant that had acquired a new specificity which was detectable with a heterologous antiserum. In a repeat experiment with the original isolate of the O5 strain, a second variant was obtained which had not only acquired the same new determinant but had, unlike the first variant, lost reactivity with the homologous antiserum. Immunoblot experiments with homologous and heterologous antisera indicated that changes in antigenic specificity were associated with the O side chains of the LPS molecules. Results of restriction endonuclease analysis of chromosomal DNA of the variants and their parents revealed minor differences in restriction patterns which suggested that C. coli is capable of undergoing genomic re-arrangements that lead to changes in LPS specificity and structure.

Homologous amino-terminal radioimmunoassay for rat parathyroid hormone

Existing radioimmunoassays for parathyroid hormone (PTH) in rat plasma are based on cross-reactivity of rat PTH (rPTH) with heterologous antisera. We used the synthetic NH2-terminal fragment of rPTH [rPTH-(1-34)] to develop a homologous radioimmunoassay for circulating PTH. An antiserum to rPTH-(1-34) was raised in a goat (G-813), and the same peptide was used as radioligand (125I) and standard. Purification of the label by high-performance liquid chromatography (HPLC) increased specific binding greater than twofold and sensitivity by 50-100%. With a final antiserum dilution of 1:70,000, maximum specific binding of 30-33%, nonspecific binding of 1-5%, and 50-microliters sample additions, the assay detection limit was 1.8-2.5 pmol/l. A midregional fragment of human PTH did not displace 125I-labeled rPTH-(1-34). HPLC of extracts of rat parathyroid glands and hyperparathyroid plasma showed only a single peak of immunoreactivity that eluted 2 min after rPTH-(1-34). Dose dilution curves for rat parathyroid gland extracts, rPTH-(1-34) added to rat plasma, and endogenous rat plasma PTH all paralleled the standard curve. Immunoreactive PTH (irPTH) was detectable in greater than 90% of fasting normal rat plasma and changed appropriately in response to hyper- and hypocalcemia induced by low-calcium and vitamin D-deficient diets, injections of calcium and EDTA, and after thyroparathyroidectomy. The normal range for rat plasma irPTH was less than 2.0-12 pmol/l, in general agreement with bioassay results of others. Thus rPTH-(1-34) is an excellent immunogen for raising antisera to rPTH, and assays incorporating it may be of great value in studying rat parathyroid physiology.

Clinical complications of heterologous antisera administration

Clinical complications of heterologous antisera administration

Cross-reacting antigens between Mycoplasma ovipneumoniae and other spices of mycoplasma of animal origin, shown by ELISA and immunoblotting with reference antisera

Using enzyme-linked immunosorbent assays with Mycoplasma ovipneumoniae as antigen, the cross-reactivity of antigens between this species and 22 other mycoplasma species was examined using reference polyclonal antisera. Significant cross-reactivity with M. ovipneumoniae was demonstrated by five species, only, viz. M. bovoculi, M. dispar, M. flocculare, M. hyopneumoniae and M. hyorhinis. Using one-dimensional SDS-PAGE and immunoblotting techniques with homologous and heterologous antisera, cross-reacting antigens of M. dispar, M. flocculare, M. hyopneumoniae and M. ovipneumoniae were further investigated. Cross-reacting antigens with apparent molecular weights of 64, 44 and 32 kDa were common to all and a 184 kDa cross-reacting antigen occured in all except M. ovipneumoniae. Further cross-reacting antigens (one-way and two-way) between two of the four species are reported. Four monoclonal antibodies against different antigens of M. ovipneumoniae did not recognise any antigen in the other three species examined.

Cross-neutralizations of phospholipase A 2 neurotoxins from snake venoms

Rabbit antisera were produced against several purified phospholipase A 2 neurotoxins from snake venoms. The neutralizing and cross-neutralizing capacities of these antisera were evaluated using mice. In all except one case, homologous antisera neutralized the lethal effects of the neurotoxins. In several instances, antisera that exhibited strong ELISA cross-reactivity ( Middlebrook and Kaiser, Toxicon, 27, 965–977, 1989) were also capable of cross-neutralizations. Homologous antisera neutralized elapid neurotoxins much more effectively than did heterologous antisera. With the crotalid neurotoxins, homologous and heterologous antisera had similar neutralization potentials. These observations further define the immunological relatedness of the phospholipase A 2 neurotoxins and suggest that common neutralizing epitopes exist within subgroups of these toxins.

Phenotypic features and proliferative activity of B cell progenitors in X-linked agammaglobulinemia.

In this study, we applied mAb and heterologous antisera in double marker combinations to investigate the phenotype and the proliferative activity of immature B lineage cells in XLA. Bone marrow (BM) samples from eight male adult patients with no circulating B lymphocytes were studied. The proportions and the phenotype of the earliest identifiable B cell progenitors, expressing nuclear terminal deoxynucleotidyl transferase (TdT), cytoplasmic CD22, and membrane CD19 and CD10 were identical to those observed in normal BM. In XLA these cells represented 1.2% to 22% of BM mononuclear cells; 5% to 42% and 1% to 45% of such cells weakly expressed CD20 and CD37, respectively, and invariably lacked CD13 and CD33. Cytoplasmic mu+ sIg- pre-B cells were seen in low numbers (0.1% to 0.3%) in four samples and were undetectable in the remaining four. Consequently, the ratio TdT+/c mu+ was greater than 100 in five out of eight samples studied in contrast to the less than 10 values seen in normal individuals. The proliferative activity of B lineage progenitor cells was studied by using Ki67 and anti-bromodeoxyuridine mAb. Although the proliferation of TdT+ cells in XLA was comparable with that seen in normal BM samples (24% to 59% of TdT+ were Ki67+ and 11% to 27% incorporated bromodeoxyuridine), this was dramatically reduced in the c mu+ cells (no c mu+, Ki67+ seen in three samples where pre-B cells were observed). Thus, the abnormalities of B cell differentiation in XLA are first seen at the c mu+ pre-B stage and suggest a maturation block in the transition between TdT+, c mu- pre-pre-B cells and c mu+ pre-B cells. The severity of this block may be variable, allowing the generation of a near normal number of pre-B cells in some patients, which nevertheless have a defective proliferative activity. Finally, our study further supports the concept that the effects of the "XLA gene" are confined within the B lineage by demonstrating that the proportions of T cells bearing TCR-alpha beta and TCR-gamma delta in XLA are similar to those seen in normal individuals.

Immunodiffusion test for serodiagnosing subcutaneous zygomycosis

Culture filtrate antigens of Basidiobolus ranarum and Conidiobolus coronatus were analyzed by immunodiffusion (ID) with homologous rabbit antisera. B. ranarum and C. coronatus were each found to have five specific antigens. Results of tests with heterologous antisera indicated that all of the species shared at least one antigen. ID tests incorporating the specific precipitin bands as references were developed for detection of basidiobolomycosis and conidiobolomycosis. These tests were performed with sera from humans and horses with proven basidiobolomycosis and conidiobolomycosis as well as with control sera from humans and animals with and without heterologous mycotic and oomycotic infections. Only sera from cases of basidiobolomycosis and conidiobolomycosis produced lines of identity with the reference precipitates of B. ranarum and C. coronatus, respectively. The ID tests were found to be completely sensitive and specific for determining the etiology of zygomycosis caused by these two species. In addition they appeared useful for monitoring resolution of the infections.