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Anti-CD3 (OKT3) MoAb is a mitogenic agent which activates lymphocytes. We have studied the effects of murine anti-human OKT3 MoAb (IgGI) alone or in combination with IL-2, human thyroglobulin (Tg) and thyroperoxidase (TPO) antigens on the proliferation of whole peripheral blood mononuclear cells (PBMC) (including monocytes) or subtypes (T, CD4+, CD8+, B) as measured by tritiated thymidine (³H-TdR) incorporation. B cell differentiation was studied by measuring numbers of IgG-secreting cells and specific anti-TPO/anti-Tg-secreting cells by SPOT ELISA. PBMC or lymphocyte subtypes, obtained from 45 patients with Hashimoto's thyroiditis (HT), 40 Graves' disease (GD) and 51 normal controls were cultured in 96 microtitre plates for 6 days in the presence of OKT3 MoAb at final concentrations 25-250 ng/ml, IL-2-15 U/ml, Tg and TPO (1 μg/ml). Then cultures were pulsed with 0.2 μCi³H-TdR/well and incorporation was measured after 18 h. IgG and anti-TPO/Tg-secreting cells were detected at 7 days. Higher proliferative responses from whole PBMC preparations in response to any of the combinations including 0KT3 MoAb were observed in the HT preparations, while the basal values were the lowest. IL-2 alone increased these responses markedly, but equally in all groups. IL-2 in combination with OKT3 had an additive effect on proliferation, with higher responses in HT, Tg and TPO antigens did not change these responses. Most HT preparations responded with their maximum proliferation to the lowest concentration of OKT3 MoAb (25 ng/ml). whereas in GD and control preparations of PBMC these responses were shifted to higher concentrations (250 ng/ml); even with those, proliferation was not so enhanced in controls when compared with HT and GD preparations. In contrast, the proliferative responses of T cells alone and subpopulations of CD8+ suppressor/cytotoxic cells were decreased in HT preparations compared with controls. Monocytes were necessary for proliferation. In the subpopulation of B cells (>95% pure) and CD4+ helper/inducer cells, differences did not reach significance. In spite of the effect on proliferation, OKT3 MoAbonly mildly but significantly increased the numbers of IgG-secreting cells in HT and GD preparations and did not stimulate synthesis of specific antibodies. Our data suggest that the increased proliferative responses of whole PBMC to OKT3 MoAb in HT preparations might be due to insufficient activation of T suppressor/cytotoxic cells. [ABSTRACT FROM AUTHOR]

A MoAb 1-22-3 (IgG3) was produced in mice immunized with rat glomeruli. A blotting study indicated the antigen molecule recognized by this MoAb has a molecular weight of about 25 kD, which is the same as that of the Thy 1.1 molecule. This MoAb is capable of inducing the morphological changes similar to those induced by anti-thymocyte serum or the anti-Thy 1.1 MoAb, ER4 and massive proteinuria in rats by a single i.v. injection. Proteinuria started immediately after MoAb 1-22-3 injection and peaked on day 5. Reaction products were detected by immunoelectron microscopy in vitro on the limited mesangial cell surface facing endothelial cells and in vivo in partially lysed mesangial cells 30 min alter injection. Unlike the proteinuria-inducing MoAb ER4, reactivity of MoAb 1-22-3 was detected neither on endothelial cells, epithelial cells, nor along the glomerular basement membrane in vivo and in vitro. There is a difference in reactivity toward guinea-pig and rabbit materials between MoAb I-22-3 and the commercial anti-Thy 1.1 MoAb (OX7). The antigenic determinant of MoAb 1-22-3 is concluded to be a new epitope and only the binding of MoAb 1-22-3 to this epitope proved to lead to an abnonnal increase of glomerular capillary wall permeability. [ABSTRACT FROM AUTHOR]

We treated 2-week-old and 8-week-old non-obese diabetic (NOD) mice with 1 mg of anti-L3T4 MoAb weekly for 4 weeks. This short-term treatment of anti-L3T4 MoAb prevented the development of overt diabetes in NOD mice, in both groups, even after cessation of the therapy. However, there were overt mononuclear cell infiltrations in the majority of islets, and no appreciable differences in the degree of insulitis between treated and control mice. There were also no significant differences in the percentage of L3T4+ T cells expressing Vβ5. Vβ8 and Vβ11 antigens between the treated and the control group. In contrast, most of male NOD mice injected with 200 mg/kg of cyclophosphamide did not become diabetic when the spleen cells from the MoAb-treated female NOD mice were transferred to these animals 48 h before the cyclophosphamide injection. Thus, the tolerance induced by the short-term administration of anti-L3T4 MoAb to NOD mice may not be due to clonal deletion, but rather to newly generated suppressor cells in the animals. [ABSTRACT FROM AUTHOR]

A collection of new and established FDC-reactive MoAb has been used in an immunohistological study designed to throw light on (a) the nature of FDC (which are of unknown lineage) as judged by their sharing of antigens with other cell types and (b) the reason for the strong expression of some B cell antigens on FDC. The MoAb were tested on: (I) sections of tonsil, (2) sections of lymph nodes from four cases of non-Hodgkin's lymphoma. (3) peripheral blood cells and (4) cells of cultured haemopoietic cell lines. Only one of the ten new MoAb bound to FDC and no other component of the tissues screened. It resembled R4/23, a MoAb known to be specific for FDC. The other nine antibodies showed a range of cross-reactivity patterns involving one or more of the following: monocytes, macrophages, platelets, epithelium, endothelium and connective tissue fibres. Some of the MoAb reacted with B lymphocytes and cells of B lymphoblastive lines but none showed the restricted FDC-staining pattern associated with MoAb which detect the CD23, P45 antigen. The findings are discussed in terms of the intrinsic or extrinsic nature of the antigens detected. [ABSTRACT FROM AUTHOR]

The effect of chlorpromazine one of several calmodulin antagonists that inhibit cytoskeletal movement, on the local kinetics of injected proteinuria-inducing MoAb 5-1-6 was examined to test the hypothesis that proteinuria is inhibited if the antigen recognized by MoAb 5-1-6 or injected MoAb remains on the surface of epithelial foot processes. MoAb 5-1-6 was injected into both chlorpromazine-treated (5 mg/100 g body weight) and untreated rats. As a positive control for the chlorpromazine treatment, anti-Fx1A serum was also injected into other chlorpromazine-treated and untreated rats. Chlorpromazine inhibited neither the change in localization of injected MoAb 5-1-6 nor proteinuria, although it showed an inhibitory effect on redistribution of immune complex and the fixation of complement in passive Heymann glomerulonephritis induced by injection of anti-Fx1A serum. We conclude that the kinetics of bound MoAb 5-1-6 are regulated by a system different from that operating in passive Heymann glomerulonephritis. [ABSTRACT FROM AUTHOR]

Murine monoclonal antibody (MoAb) 5-1-6 was already reported to bind to epithelial cell foot processes and to cause proteinuria in rats. In vivo kinetics of the injected MoAb 5-1-6, relationship between the quantity of kidney-binding antibody and proteinuria, and changes In the amount of antigenic molecule recognized by this MoAb in the proteinuric state were studied. The amount of total kidney-binding antibody (TKAb) as determined 1 h alter a 2 mg administration was 50.8 ± 10.4 μg/2 kidneys, and TKAb declined to 1.9 ± 0.4 at day 15. The minimum dose of MoAb required to induce proteinuria was 125 μg as the injected dose. This dose corresponded to 12.8 μg of TKAb at 1 h and 0.34 μg of TKA bat day 5. The amount of MoAb 5-1-6 binding to isolated normal glomeruli was also shown to exceed 147.7 μg/76000 glomeruli, indicating proteinuria to be induced provided more than 8.7% (= 12.8/147.7) of the critical epitopes is specifically occupied by MoAb. The total amount of MoAb 5-1-6 bound to glomeruli in vivo and in vitro was assayed with [125]-labelled anti-mouse IgG. The amount of [125] anti-mouse IgG bound to glomeruli was 6.93 ± 0.45 μg/10 000 glomeruli from rat 5 days after this MoAb injection and 26.58 ± 0.66 μg/10 000 control glomeruli. indicating the decrease in the number of MoAb 5-1-6-rccognized antigen molecules in glomeruli isolated from the rat in proteinuric state induced by this MoAb. Thus, the MoAb 5-1-6-recognized molecule itself may principally function to regulate the permeability of the glomerular capillary wall and the decrease of the molecule may lead to proteinuria. [ABSTRACT FROM AUTHOR]

The mechanisms by which anti-DNA MoAbs derived from MRL-lpr/lpr mice, bind to human umbilical vein endothelial cells (HUVEC) and glomerular mesangial cells were studied using a cellular ELISA. DNAse-treatment of either the MoAb or HUVEC followed by reconstitution with DNA and/or histones was performed to determine whether DNA and histones mediated such binding. It was found that MoAb 410 bound to HUVEC and mesangial cells in the form of preformed DNA/anti-DNA immune complex, and such binding was facilitated by histones. In contrast. MoAb 152 bound directly to cell membrane-associated DNA, and adding DNA to MoAb 152 reduced its cellular binding. DNA binds endothelial cell, surface and histones enhance the binding of both MoAb 410 and MoAb 152 to HUVEC by increasing cell membrane-associated DNA. Finally, the degree of MoAb binding to HUVEC is critically influenced by the relative concentrations of antibody. DNA, and histones. [ABSTRACT FROM AUTHOR]

CD4 monoclonal antibody (MoAb) was able to inhibit T cell proliferation induced in an autologous mixed lymphocyte reaction (AMLR). The effect of CD4 MoAb on cellular proliferation appears to be directly exerted on CD4+ T lymphocytes, and to be due to inhibition of a post-activation event, since the CD4+ T cell proliferation that occurs after an activation pulse of 24 h with autologous non-T cells could be inhibited when CD4 MoAb was added after, but not during, the pulse period, and the inhibition of autologous MLR-induced CD4+ T cell proliferation by CD4 MoAb was observed even if the Moab was added as late as 72 h after the initiation of culture. The presence of CD4 MoAb did not affect the production of interleukin 2 (IL-2). CD4 MoAb had, however, an inhibitory effect on the expression of IL-2 receptors, such that addition of exogenous IL-2 at the initiation of culture did not restore the AMLR-induced CD4+ T cell proliferation. These results indicate that the hindrance of the recognition of H LA class II products is not the only target of the CD4 MoAb effect in the autologous MLR. Rather, the binding of CD4 MoAb to CD4! T cells interferes with a late event because it is capable of abolishing the proliferative activity of fully activated CD4+ T cells. The data are compatible with the idea that perturbation of the CD4 molecules can transmit a negative signal to CD4+ T cells. [ABSTRACT FROM AUTHOR]

Anti-CD2 MoAbs have previously been shown to induce tolerance and to block B cell differentiation. T cell and monocyte activation. Since these immune functions are important in joint inflammation, we asked whether administration of the anti-CD2 MoAb OX34 has a beneficial effect on established rat adjuvant arthritis, a model of human rheumatoid arthritis, and how it affects CD2-bearing leucocyte subsets. Female Lewis rats with established adjuvant arthritis received a total of 5 mg OX34 or isotypematched control MoAb starting on day 15 after adjuvant injection. Weight and arthritis score (AS) were measured in a blinded fashion. Peripheral blood cells were analysed for numbers of leucocyte subsets at various time points. Animals were killed on day 30 and lymphatic organs were processed for immunohistology. Clinically, OX34 treatment led lo increased body weight and reduced AS. Although OX34 binds to CD4 + and CD8 + T cells in a comparable fashion. OX34 treatment reduced CD4 + T cells, bul not CD8 + T cells. Among CD4 + T cells CD45RC + ('naive') T cells virtually disappeared; CD45RC- ('recently activated') T cells were slightly reduced. A reduction of CD4 + T cells was also found in the lung, liver, bone marrow, spleen and lymph nodes. Down-modulation of the CD2 molecule by OX34, again, affected CD4 + T cells, suggesting a specific signal for CD4 + but not CD8 + T cells. In conclusion, the anti-CD2 MoAb OX34 attenuates established rat adjuvant arthritis. In spite of similar binding to CD4 + and CD8 + T cells. OX34 depletes only CD4 + T cells and down-modulates the CD2 molecule on these cells. These results suggest a therapeutic benefit from CD2-directed therapy for chronic types of arthritis. [ABSTRACT FROM AUTHOR]

Anti-CD3 MoAb treatment is widely used as an immunosuppressive therapy. In the present study we examined the in vitro T cell response in mice having received 24 h before it single i.v. injection of 10 μg of anti-CD3 MoAb. We found that splenocytes from these mice displayed a dramatically decreased proliferative response to the T cell mitogens concanavalin A (Con A), anti-CD3, phytohaemagglutinin (PHA) and phorbol myristate acetate (PMA) + calcium ionophore, while the effect of lipopolysaccharide (LPS) was not impaired. T cell suppression persisted for about 10 days after anti-CD3 injection, returning to normal within 15 days. The F(ab')2 fragment of anti- CD3 had no such effect, indicating the requirement for in vivo activation. At the dose used, anti- CD3 resulted neither in T cell depletion nor in down-modulation of the CD3/Tcell receptor (TCR) complex. The low proliferation was also not explained by apoptosis, following secondary challenge with Con A. Splenocytes from anti-CD3-injected mice were highly responsive to IL-2, but generated little or no IL-2. IL-3, lL-4 and interferon-gamma (IFN--)) when exposed to Con A. Normal cytokine production could not be restored by the addition of optimal doses of IL-2 during Con A stimulation. Transforming growth factor-beta (TGF-β) was the only cytokine whose mRNA expression was not modified in stimulated splenocytes from anti-CD3-injected mice. Furthermore, anti-TGF-β antibodies increased Con A-induced T cell proliferation, but not cytokine production. [ABSTRACT FROM AUTHOR]

A mouse anti-cholera toxin (CT) MoAb, mAb1, specific for the GM1-binding epitope of CT, was used to raise a syngenic anti-idiotypic MoAb, mAb2. Purified mAb2 was specific for mAb1 as shown by latex particle counting immunoassay and ELISA. Several experiments of competition between mAb2 and CT for binding to mAb1 demonstrated that mAb2 bore an internal image of the GM1-binding epitope of CT. Binding of mAb2 to GM1 unambiguously corroborated the mAb1-paratopic specificity of mAb2. Furthermore, mAb2 acted as a CT-surrogate antigen: rabbits injected with mAb2 produced some anti-CT antibodies, Ab3, which resembled mAb1 in specificity as expected. The potential use of this mAb2 as vaccine or as prophylactic agent to prevent CT from binding to its cellular receptor is discussed. [ABSTRACT FROM AUTHOR]

Cat and dog albumins from commercial extracts were used to produce monoclonal antibodies (MoAb). Anti-cat albumin MoAb recognized both cat and dog albumin equally, as did anti-dog albumin MoAb; this confirms cross-reactivity between cat and dog. The MoAb were separated into two groups according to their epitopic specificity; they recognized two overlapping epitopes of cat albumin. Furthermore, by competitive inhibition of radio-allergosorbent test (RAST). it was shown that one MoAb group inhibited significantly the binding of human IgE antibodies (from a pool of 13 patients allergic to both cats and dogs) to insolubilized cat or dog extracts. These observations suggest that murine anti-cat or anti-dog MoAb and human IgE antibodies recognize identical or closely related determinants on cat and dog albumin. [ABSTRACT FROM AUTHOR]

In a previous investigation, we found that murine MoAb 42C3, raised against human Tg, recognized Tg differently depending upon its level of iodination of Tg. A possible explanation for this finding is that iodine is directly involved with the specific epitope recognized by MoAb 42C3. In the present study, we report that the binding of MoAb 42C3 to iodinated Tg is inhibited by T4, T3, reverse T3 (rT3), triiodothyroacetic acid (triac), diiodothyronine (T2), diiodotyrosine (DIT), but not by thyronine (T0) or tyrosine. The order of inhibition of these iodinated compounds is T4 > T3 > rT3 > triac > T2 > DIT. The MoAb 42C3 does not have the same specificity as the T3, T4-receptor since the order of binding of these iodinated compounds on the receptor differed from the order of their inhibition of this MoAb. Monoclonal antibody 42C3 also recognized non-iodinated Tg that was subsequently iodinated in vitro. It failed to recognize another protein, bovine serum albumin, that was iodinated in vitro by the same method. These results suggest that iodinated tyrosines and thyronines determine the binding specificity of MoAb 42C3. The inhibitory effects of these compounds on MoAb 42C3 depend on their iodine content as well as location of iodine in the aromatic ring. [ABSTRACT FROM AUTHOR]

A cell wall fraction of Mycobacterium leprae has enhanced potency in activating immune T ceils. By using a panel of monoclonal antibodies (MoAb). the dominant immunogen in This preparation was shown to be a complex of proteins of apparent molecular weight (M.) 65 to 50 kD with a major antigen of 65 kD. Antigen capture assays supported the results of immunoblots and ELISA that this protein was concentrated in the cell wall. By varying the MoAb used as capture or tracer antibody, one of [he three MoAb-defined epitopes on the 65 kD protein proved to be unique to M. leprae while the other two were shared by M. bovis (BCG) and M. tuberculosis. The cross-reactive epitope defined by MoAb L22 was present on a protein of Mt 12 kD as well as the 65 kD protein. The 12 kD protein was strongly radiolabelled with 125I and was immunoprecipitated by L22 but not by two other MoAb. LI2 or L14. By contrast the higher molecular weight forms were only weakly precipitated by the three MoAb, Competitive inhibition assays with lepromalous leprosy sera demonstrated that the MoAb-defined epitopes were recognized by human B cells. The proteins bearing one of the cross-reactive determinants was purified from M. bovis (BCG) sonicate by affinity chromatography with MoAb L22 coupled to Sepharose 4B, This antigen fraction stimulated proliferation in peripheral blood mononuclear cells from BCG vaccinated, mantoux positive individuals indicating that the cell wall protein has cellular as well us humoral reactivity. The three MoAb defined epitopes are encoded by the DNA clone Y3I78 recently isolated from M. leprae. [ABSTRACT FROM AUTHOR]

Lipoarabinomannan (LAM) is a major structural carbohydrate antigen of the outer surface ofMycobacterium tuberculosis. High antibody titres against LAM are often seen in active tuberculosis (TB). The role of such LAM-specific antibodies in the immune response against TB is unknown. Here we have investigated a monoclonal antibody (MoAb) SMITB14 of IgG1 subclass and its corresponding F(ab′)2 fragment directed against LAM fromM. tuberculosisstrain H37Rv. MoAb SMITB14 was shown by immunofluorescence to bind to whole cells of the clinical isolateM. tuberculosisstrain Harlingen as well as toM. tuberculosisH37Rv. The binding of MoAb SMITB14 to LAM was inhibited by arabinomannan (AM) and oligosaccharides (5·2 kDa) derived from LAM, showing that the MoAb binds specifically to the AM carbohydrate portion of LAM. In passive protection experiments BALB/c mice were infected intravenously withM. tuberculosisHarlingen. MoAb SMITB14 was added intravenously either prior to, or together with, the bacteria. The antibody proved to be protective against theM. tuberculosisinfection in terms of a dose-dependent reduction in bacterial load in spleens and lungs, reduced weight loss and, most importantly, increased long-term survival. [ABSTRACT FROM AUTHOR]

The hyperthyroidism of Graves’ disease (GD) is caused by TSH-receptor (TSH-R) stimulating autoantibodies (TSAb), leading to overproduction of thyroid hormones. We present evidence for TSAb interaction with three distinct regions of the TSH-R, one in immediate vicinity of the carboxy terminal serpentine. Three murine monoclonal antibodies (MoAbs 28·1, A9 and 31·7) directed to amino acids 36–40, 147–228 and 382–415 were labelled and tested for their binding to human recombinant TSH-R on solid phase. All MoAbs bound to TSH-R with a Kd of 8–12 n m and showed no competition among themselves. We tested 114 sera from euthyroid controls, 118 TBII positive sera from patients with GD (containing TSAb confirmed by bioassays), 16 TBII positive sera from patients with autoimmune thyroid disease (AIT), who were hypothyroid and had TSH blocking antibodies (TBAb), and 20 patients with AIT, who were hypothyroid but negative for all TRAb. Mid-regional MoAb A9 tracer achieved the highest sensitivity in the GD group (72·0%), whereas C-terminal MoAb 31·7 found most sera positive in the AIT group (87·5%). Surprisingly, the N-terminal MoAb 28·1 had the lowest sensitivity in the GD (10·4%) and AIT group (43·8%). Using a mixture of all three tracer MoAbs did not increase the sensitivity in the GD or AIT group, compared to the best single MoAb alone. Median inhibition of MoAb A9 was significantly ( P < 0·001) higher than inhibition of MoAbs 28·1 or 31·7 in the group of GD patients but not in other groups. Almost all patient sera with positive reactivity in the MoAb tracer assays had TBII values in the higher range. However, there were many highly TBII positive sera, which did not show a displacement of the MoAb tracers. We conclude that, contrary to some reports, the binding of TSAb and TBAb to the TSH-R is not restricted to distinct and distant epitopes. The middle part of the TSH-R seems to be more relevant for TSAb binding than the N-terminal part, while a proportion of TSAb autoantibodies also binds to a C-terminal epitope of the TSH-R. The method described here is a TSH independent competitive assay for the detection of TSH-R autoantibodies. [ABSTRACT FROM AUTHOR]

We tested the capacity of three rat monoclonal antibodies (MoAb), recognizing different epitopes on the L chain of mouse interleukin-2 receptor (IL-2R), to block delayed-type hypersensitivity (DTH) and local graft-versus-host reaction (GVHR). Furthermore, we investigated the effect of IL-2R- targeted immunotherapy on serum soluble IL-2R levels by ELISA technique. Following administration of AMT- 13 MoAb, in sufficient amounts to suppress both DTH (60-90% inhibition) and local GVHR (55-70% inhibition), an increase in soluble IL-2R levels up to 12-fold was observed. In contrast, the administration of 7D4 MoAb did not show any immunosuppressive effect in vivo and even decreased the soluble IL-2R levels. The third anti-IL-2R MoAb AMT45-20 had an intermediate effect. AMT4S-20 MoAb marginally suppressed the DTH as well as local GVHR (15-35% inhibition) and induced only a slight increase in soluble IL-2R levels (up to 4-fold). Both cyclosporin A, a conventional immunosuppressive drug, and the anti-L3T4 MoAb, which defines the entire T helper cell subset, suppressed GVH and DTH response but did not increase the soluble IL-2R serum levels. The increased concentration of soluble IL-2R in the serum of successfully treated mice may be due to destruction of IL-2R-positive cells by anti-IL-2R-targeted immunotherapy and seems to be a sensitive indicator for the success of such a therapy. [ABSTRACT FROM AUTHOR]

Using hybridoma technology we produced monoclonal antibodies (MoAb) to idiotypic determinants on human anti-cardiolipin antibodies purified from a patient with SLE. Hybridomas were screened by inhibition of cardiolipin binding activity of sera from patients with SLE. Seven hybridomas were selected, two of which were studied extensively. Sera from a number of patients with SLE were found to share idiotypic determinants. This cross-reacting idiotype was not detectable on anti-cardiolipin antibodies in syphilis sera. The cross-reacting idiotype was present in sera with anti-ssDNA antibodies even though some of these sera had no anti-cardiolipin antibodies. We propose that these MoAb may recognize a regulatory idiotype. [ABSTRACT FROM AUTHOR]

The injection of the 145-2C11 anti-CD3 MoAb in mice induces a polyclonal T cell activation resulting in the release of several cytokines, including interferon-gamma (IFN-γ) and tumour necrosis factor-alpha (TNF-α). As these cytokines are known to be involved in the host defence against Trypanosoma cruzi, we measured serum levels of IFN-γ and TNF-α after injection of the 145-2C11 MoAb in the course of experimental murine Chagas' disease. Compared with control mice, T. cruzi-infected BALB/c mice were found to be primed to secrete very high levels of IFN-γ and TNF-α from the second and the first week of infection, respectively, up to the chronic phase. In vivo cell depletion experiments indicated that CD8+ T cells were responsible for these dramatic hyperproductions of IFN-γ and TNF-α. While all control mice survived anti-CD3 MoAb injection, a high lethality rate was observed in T. cruzi-infected mice within 24 h after anti-CD3 MoAb challenge. Pretreatment with neutralizing anti-IFN-γ MoAb or depletion of CD8+ T cell population dramatically decreased the mortality induced by anti-CD3 MoAb in T. cruzi-infected mice. Finally, we showed that anti-CD3 MoAb injection in T. cruzi-infected mice was followed by a massive release of nitric oxide (NO) metabolites, which was partially reduced by IFN-γ or TNF-α neutralization. The administration of the NO synthase inhibitor N-nitro-L-arginine methyl ester (L-NAME) before anti-CD 3 MoAb challenge did not prevent and even enhanced lethality in T. cruzi-infected mice, suggesting that NO overproduction and lethal shock are not causally related. We conclude that injection of anti-CD3 MoAb in the course of experimental Chagas' disease induces a CD8+ cell-dependent shock mediated by IFN-γ and TNF-α. [ABSTRACT FROM AUTHOR]

Susceptibility to the development of MoAb 5-1-6-induced proteinuria was investigated in four different rat strains, i.e. Brown-Norway (BN), Lewis (LEW). Sprague-Dawley (SD) and Wistar. An intravenous injection of 5 mg of MoAb 5-1-6 to female 7-week-old rats of a given strain induced massive proteinuria in BN, LEW and Wistar rats. However, SD rats developed almost no proteinuria. A similar tendency was observed in the second experiment, in which the injected dose of MoAb was adjusted according to the body weight of each rat (3 mg/l00 g body weight). Immunofluorescence (IF) and immunoelectron microscopy (IEM) revealed no differences between the binding patterns of the MoAbs to normal rat kidneys derived from each strain. Quantitative study using 1251-labelled MoAb showed that there was no significant difference in the amount of antibody bound to the kidney I h and 5 days after injection between two rat strains, LEW and SD, Localization of 5-1-6 in vivo and its kinetics were investigated. In IF a linear-like pattern along capillary walls was observed 2 h after injection in both LEW and SD strains. This linear-like pattern was shifted to a granular pattern in proteinuric LEW rats 6 days after injection, whereas it remained linear-like in non-proteinuric SD rats. IEM confirmed this difference in the localization of injected MoAb 6 days after injection to LEW and SD rats also at the ultrastructural level. We conclude that there is a clear-cut strain difference in the development of proteinuria induced by MoAb 5-1-6. SD rats were less susceptible to MoAb-induced glomerular injury than BN, LEW and Wistar rats. Although the exact reason for strain variation in susceptibility to MoAb-induced proteinuria remains to be clarified, the movement of bound MoAb, presumably together with corresponding antigenic molecule along the glomerular epithelial cell surface followed by endocytosis into the epithelial cell, seems to be closely related to the induction of proteinuria. [ABSTRACT FROM AUTHOR]

Anti-CD3 MoAbs arc widely used in T cell activation studies, and are effective in immunosuppressive therapy. We used a panel of mouse (m) anti-CD3 switch variant MoAbs of five different isotypes to study IL-6 release from accessory cells. Incubation of human (h) mononuclear cells with anti-CD3 MoAbs resulted in increased lL-6 levels with MoAbs of mIgGI and mIgG2a isotypes. with no effect of mIgG2b or mIgA. This suggested involvement of IgG Fc receptors (FcγR) in triggering IL-6 production. To evaluate the role of different FcγR molecules individually we used a panel of hFcγR-transfected mouse fibroblasts. and Jurkat T cells as a model. IL-6 secretion by CD32 transfectants expressing the hFcγRHa high-responder (HR) allelic form was triggered by mIgGl anti-CD3 MoAb, with no effect of four other isotypes. None of theanti-CD3 MoAbs induced IL-6 secretion by CD32 transfectants expressing either a variant of this receptor, containing only a single intracellular amino acid (CT-). the hFcγRIIa low-responder (LR) allelic form, or hFcγRIIb1. hFcγRI (CD64) transfectants exhibited IL-6 production after incubation with mIgG2a anti-CD3 MoAb, and to a lesser extent with mlgG2b. and mIgGI MoAb. Indirect involvement of T cells in triggering IL-6 secretion could be excluded by experiments in which transfectants were cultured with immobilized anti-CD3 MoAb. These data indicate that cross-linking of either hFcγRI, or hFcγRIIaHR by appropriate anti-CD3 MoAbs triggers IL-6 production of accessory cells, and not T cells. This may also take place in vivo during immunosuppressive therapy with anti-CD3 MoAbs. and related antibody-mediated immune responses. [ABSTRACT FROM AUTHOR]

A new monoclonal antibody (MoAb) HA58 (IgG1) was prepared, which recognizes the binding site on the intercellular adhesion molecule-1 (ICAM-1) antigen to the lymphocyte function-associated antigen-1 (LFA-1). The double-determinant immunoassay (DDIA) was established with use of MoAb HASS and another anti-JCAM-1, MoAb CL207, to detect the soluble, shedding ICAM-1 antigen. Human recombinant interferon-gamma (IFN-γ) induced not only the expression of cell surface ICAM-1, but also the shedding ICAM-1 antigen in an IFN-γ concentration-dependent and incubation-time-dependent manner. DDIA was applied to detect the shedding ICAM-1 antigen in the sera of patients with malignant or benign diseases. The incidence of positivity for ICAM-1 antigen in malignant diseases was higher than that in benign diseases or in healthy controls. Furthermore, the sera of cancer patients with liver metastasis showed higher levels of the shedding ICAM-1 antigen. These findings suggest that serum ICAM-1 antigen may be a useful marker to monitor tumor burden in cancer patients. [ABSTRACT FROM AUTHOR]

The effects of a monoclonal antibody (MoAb DB-1), which neutralized rat interferon-gamma (IFN-γ), on the induction and progression of adjuvant arthritis in Lewis rats induced by intraplantar injection of Freund's complete adjuvant was studied. The animals were treated intraperitoneally with MoAb DB-1 (0.3-5 mg) for various times. Prophylactic treatment with MoAb DB-1, starting 2 days prior to arthritis induction, inhibited oedema in both the injected and non-injected hind paws and delayed joint destruction as shown by radiography. However, despite continued MoAb treatment, the disease progressed. High doses of MoAb DB-1 exacerbated the disease. A control MoAb of the same isotype did not have significant effects on adjuvant arthritis. Therapeutic treatment with the MoAb DB-1 starting 8 days after arthritis induction caused only slight and short-lived inhibitory effects. IFN-γ appears to be a critical lymphokine for the development of adjuvant arthritis. [ABSTRACT FROM AUTHOR]

Local oral passive immunization in human subjects with a monoclonal antibody (MoAb) raised against the 185-kD antigen I/II from S. mutans significantly reduced or prevented oral colonization of an exogenous strain of the organism. In subjects sham-immunized with either saline or an unrelated MoAb, however, significantly greater proportions of S. mutans persisted for a longer duration than in those immunized with the specific anti-streptococcal MoAb. Recolonization of indigenous. S. mutans after this organism was reduced to undetectable levels by an antimicrobial agent has also been completely prevented with specific MoAb. Indeed. S. mutans was not detected for a period of over I year, as compared with recolonization within 10-82 days in the control subjects. The specificity of MoAb in preventing colonization of the streptococci was studied with four MoAb. This revealed that: (1) the sub-class of antibody is not an essential factor, as both MoAb Guy's 1 and 13 prevented colonization, although Guy's 1 is an lgG2a and Guy's 13 is an IgG1 class of antibody; (2) serotype specificity is important, as MoAb Guy's 9, which only recognizes S. sobrinus (serotypes d and g), does not prevent colonisation by S. mutans (serotype c); (3) neither protein nor carbohydrate nature of the putative adhesin was a determining factor, because MoAb Guy's 1 recognizes a carbohydrate and Guy's 13 a protein determinant and both MoAb prevented adherence of S. mutans, and (4) epitope specificity appears to be the most important factor in preventing adherence of S. mutans, as MoAb Guy's 11 and 13 share the same serotype specificity and both recognize a protein determinant, yet only Guy's 13 prevents colonisation. The long duration of protection from re-colonization by indigenous S. mutans, lasting about I year after application of the specific MoAb was stopped, cannot be accounted for by functional MoAb remaining on the teeth. We suggest that initially the MoAb prevents colonization by S. mutans and that the ecological niche vacated by this streptococcus is filled by other organisms from the oral flora, thereby discouraging re-colonization by S. mutans. [ABSTRACT FROM AUTHOR]

The role of Class I major histocompatibility complex (MHC) molecules in the autologous (AMLR) and allogeneic mixed lymphocyte reactions was investigated by using monoclonal antibodies (MoAb) directed to polymorphic MHC determinants. The AMLR from subjects with the HLA-A2 phenotype was consistently inhibited by the anti-HLA-A2 MoAb, CR11-351, and the inhibition was dose-dependent and complete even at low antibody concentrations. The allogeneic MLR was inhibited by CR11-351 less than 30% when HLA-A2-bearing cells were used either as stimulator or responder cells. Addition of interleukins 1 and/or 2 to the AMLR in the presence of the inhibiting MoAbs did not restore the proliferative response. These studies suggest that Class 1 MHC polymorphic determinants, or closely related structures, participate in the induction of the AMLR. [ABSTRACT FROM AUTHOR]

Monoclonal antibodies (MoAb) produced against a sonicated antigen extract of Mycobacterium bovis were examined for their ability to bind to 10 common tissue antigens. The binding patterns of MoAb MB2, MB3, MB7. MBI I and MB13 indicated that each recognized different epitopes. MoAb M B5 and MB17 failed to bind to any of the tissue antigens. MB3 reacted strongly with normal tissue from M. bovis infected guinea pigs and ferrets but there was no reaction with MB5 and MBI7. These specificities were confirmed by reciprocal absorption experiments, it is concluded that Mycobacterium species contain epitopes that are also present in host tissue antigens. [ABSTRACT FROM AUTHOR]

A mouse hybridoma (IGI2) producing sperm-immobilizing MoAb to human .sperm was established and characterized in order to study the antigens relevant to sperm immobilization by antibodies. MoAb 1G12 had strong sperin-immobilizing and agglutinating activities and also showed a fertilization blocking activity on in vitro fertilization tests. The antibody absorption experiments showed that MoAb 1G12 reacted not only to ejaculated sperm but also human seminal plasma. suggesting that the corresponding antigen might be a sperm coating antigen. The MoAb also reacted with peripheral blood lymphocytes. In histochemical studies, the epithelia of corpus epididymis were most strongly stained. Ejaculated sperm were stained with a granular pattern for their entire surface by immunofluorescence. MoAb 1G12 recognized polymotphic glycoproteins of 15-23 kD in the ejaculated sperm extract in Western blot analysis. After deglycosilation of the sperm extract, only a single staining band of under 15 kD was detected by MoAb IG12. This suggests that the antigen epitope recognized by MoAb IGI2 might be a peptide of the core portion of the glycoprotein. MoAb 1G12 might be a useful tool for studying the mechanism of egg-sperm interaction, and also be applied to identifying the corresponding antigen by using gene technology. [ABSTRACT FROM AUTHOR]

To study the role of anti-idiotypic antibodies in the regulation of the immune response to Lol p I (the major allergenic component of rye grass pollen), we have recently generated a panel of three MoAbs directed against distinct epitopes of Lol p I and an anti-idiotypic MoAb directed against the idiotype borne by one of the anti-Lol p I MoAbs (290A-I67). The effects of pretreatment with this anti-idiotypic MoAb in BALB/c mice before immunization with the antigen have been examined. The anti-idiotypic MoAb or unrelated MoAb were given weekly for 8 weeks intraperitoneally. Mice then received the antigen (2 μg) adsorbed with alum (2 mg) at weeks 9, 11 and 13. Serum anti-Lol p I antibodies (IgG or IgE) and specific idiotypic responses were measured. Anti-Lol p I IgG antibodies could be detected before immunization with Lol p I only in mice pretreated with anti-idiotypic MoAb. Immunization with Lol p I induced an anti-Lol p I IgG response in both groups, but this response was higher in mice that received anti-idiotypic MoAb. Similar profiles were seen for specific IgE antibodies and idiotypic responses. Surprisingly, idiotypes borne by other anti-Lol p I MoAbs (539A-6 and 348A-6) had also been enhanced after pretreatment with the anti-290A-167 MoAb. These observations suggested that the pretreatment with this anti-idiotypic MoAb modulates not only the expression of the respective idiotype, but also affects other idiotype responses. [ABSTRACT FROM AUTHOR]

Both murine CD4+ and CD8+ cells are found in the thyroid infiltrate in experimental autoimmune thyroiditis (EAT) induced with mouse thyroglobulin (MTg). MTg activation of immune cells in vitro enables CD4+ cells to transfer thyroiditis adoptively and to aid the cytotoxic capacity of CD8+ cells for thyroid monolayers. To dissect their relative contribution to pathogenesis in vivo, depleting doses of paired rat monoclonal antibodies (MoAb) recognizing two distinct CD4 or CD8 epitopes were injected alone or in combination. Early treatment with CD4 MoAb interfered with the induction and development of EAT, whereas similar treatment with CD8 MoAb reduced infiltration moderately and did not enhance antibody response. To examine the long-term effect of therapy on advancing EAT, administration of MoAb was delayed to days 21 and 25, and thyroids were analysed immunohistochemically on days 28 and 70. Whereas control mice showed about 30% CD4+ and CD8+ cells at a 2:1 ratio (the remainder being mostly macrophages) on both days 28 and 70, the CD4 therapy regime led to reduced severity and the lesions on day 70 contained very low percentage of CD4+ cells, but elevated percentage of CD8+ cells (ratio 1:3-5). The CD8 therapy regime led to reduced CD8+ cells without changing the range of CD4+ cells (ratio 4:1). Thus, subset involvement may be influenced by the MoAb used. When CD4 and CD8 MoAb were combined, > 50% of the thyroids were cleared of all inflammatory cells; lesions when found were very small and contained < 10% T cells (ratio 1:1). Since emerging T cells were not retained in the thyroid despite ongoing antigenic stimulus leading to increased antibody titres, the therapeutic effect of MoAb, even at an advanced stage of disease, was long lasting. [ABSTRACT FROM AUTHOR]

Seventeen patients with Dupuytren's contracture underwent partial fascicetomy, and frozen tissue sections from the involved palmar fascia were prepared for binding studies with hybridoma-derived murine monoclonal antibodies (MoAb) recognizing connective tissue differentiation antigens. The two MoAb used were both generated using human sarcomas as immunizing agents, 23H7 known to bind to an antigen shared by selected sarcomas and carcinomas but not normal adult tissues except a subset of granulocytes,. and 12C9 shown to recognize a common fibroblastic marker. MoAb 23H7 was discovered to bind to a subset of fibroblasts within the lesions of six of 17 patients with Dupuytren's disease. Occasionally it immunostained a single cell population associated with tissue granulocytes dispersed in the surroundings of the lesions. MoAb 12C9 was found to be expressed in only 12 of 17 specimens prepared from involved lesions from Dupuytren's disease. It is suggested that fibroblasts from selected patients with Dupuytren's contracture express a novel antigen, defined by MoAb 23H7 previously shown to be associated with human sarcomas and other neoplasia. The other fibroblast marker which is defined by MoAb 12C9 and known to be a common connective tissue antigen, is only occasionally expressed in lesions involved with this disease. Though additional markers associated with Dupuytren's contracture remain to be delined, the MoAb, capable of defining connective tissue differentiation markers, reported in this study may serve as new immunological probes for immunodissecting this syndrome into subsets of diseases which may better define the variety of clinical patterns presented by patients. [ABSTRACT FROM AUTHOR]

As part of a study of the characteristics of the synovial membrane which make it susceptible to inflammatory reactions, we tested a number of monoclonal antibodies (MoAb) which revealed novel features of the synovium using tissue from rheumatoid, osteoarthritic and traumatized (mechanically deranged) joints. In a previous study we detected macrophages (Mph) lining the synovial membrane by means of Mph specific and HLA-DR specific MoAb. These may account for the type A synoviocytes. Type B synoviocytes are thought to be fibroblastic and we used an anti-Thy1 MoAb to identify these cells. Many fibroblasts were seen in a subintimal position but only few in the lining layer, not in sufficient numbers to account for the type-B-category of synoviocyte. Staining with a new MoAb, 67. was found to precisely delineate the lining layer. This MoAb was previously seen to react with dendritic reticulum cells (DRC) of germinal centres, cells involved in B lymphocyte activation. However, other MoAb which react with germinal centre DRC did not label the synovial lining layer. Several MoAb revealed features of the vasculature not previously recognized. In rheumatoid samples. MoAb10 labelled small capillaries near the synovial surface and larger vessels deeper in the intima were labelled with a second MoAb. SMØ. This dichotomy of staining was not so apparent in synovium from control osteoarthritis and trauma (OA/T) samples. In addition, the Thy1 epitope, identified previously on a variety of human cells, was strongly expressed on all vascular endothelium. Finally a new vessel or duct like structure was identified in OA/T samples. located subintimally. These ducts contained keratin, detected with MoAb LE61 and may be the normal counterpart for the rare malignancy, biphasic synovial sarcoma. [ABSTRACT FROM AUTHOR]

The importance of the idiotypic network is represented in experimental SLF induced by active immunization of naive mice with an anti-DNA idiotype (Abl) emulsified in adjuvant. The mice after 4 months of incubation generate Ab3 having anti-DNA activity. In addition, the mice develop other serological markers for SLE associated with clinical and histopathological manifestations characteristic of the disease. To confirm further the etiological role of the idiotype in this experimental model, the mice were treated with specific anti-idiotypic antibodies (anti-id) which were also conjugated to a toxin-saporin (Immunotoxin (IT)). Pretreatment of hybridoma cell line producing the anti-anti-Id (anti-DNA = (Ab3)) for 48 h with the anti-Id MoAb (Ab2) reduced the production of anti-DNA by 58%, while pretreatment with the IT resulted in 86% decrease in anti-DNA secretion (saporin alone had only 12% effect). The anti-Id MoAb had no effect on the production of immunoglobutin by an unrelated cell line. In vivo treatment of mice with experimental SLE led to a significant decrease in titres of serum autoantibodies, with diminished clinical manifestations. The results were more remarkable when the IT was employed. These suppressive effects were specific, since an anti-Id treatment of experimental anti-phospholipid syndrome was of no avail. The anti-Id effect was mediated via a reduction in specific anti-DNA anti body-forming cells, and lasted only while anti-Id injections were given. Discontinuation of the anti-Id injection was followed by a rise in titres of anti-DNA antibodies. No immunological escape of new anti-DNA Ids was noted. Our results point to the importance of pathogenic idiotypes in SLF and to the specific potential of implementing anti-idiotypic therapy, enhanced by the conjugation of the anti-Id to an immunotoxin, in particular one with low spontaneous toxicity. [ABSTRACT FROM AUTHOR]

A single i.v. injection of 3 mg of the F(ab')2 fragment of MoAb 5-1-6 into rats induced immediate proteinuria (128.1±80.7 mg/24 h on day 1) which lasted 1-2 days. In contrast, rats administered 10 mg of the corresponding Fab fragment did not develop abnormal proteinuria even though an equivalent dose of the intact MoAb 5-1-6 far exceeded the nephritogenic dose. The total kidney binding of 125-Fab fragment was 209.5 ± 34.3 μg/2 kidneys. This exceeded that obtained by injection of 3 mg MoAb 5-1-6 IgG1 (58.9 ± 12.5 μg/2 kidneys at 1 h) and was similar to that obtained following injection of 3 mg F(ab)2 fragment (235.3 ± l6.9 μg/2 kidneys). Immunofluorescence (IF) showed a linear pattern along the glomerular capillary wall at 1h after the administration of MoAb 5-1-6 IgG 1, F(ab )2 or Fab fragment. On day 5, fine to coarse granules were observed scattered in F(ab')2-injected rat glomeruli, whereas granules were densely localized in Fab-injected rat glomeruli. Complement-depleted rats injected with 3 mg of MoAb 5-1-6 IgG1 developed proteinuria with the same time course as non-depleted rats. This observation, together with the ability of F(ab')2 to induce proteinuria, indicates that proteinuria induced by MoAb 5-1-6 is complement-independent. This study suggests that MoAb 5-l-6-induced proteinuria is initiated by cross-linking of the epitopes by divalent MoAb 5-1-6 and is independent of complement activity. [ABSTRACT FROM AUTHOR]

An anti-idiotypic antiserum was prepared in a rabbit immunized against a pool of six monoclonal anti-DNA antibodies generated in B/W mice. This antiserum detected idiotypic determinants in four out of the six monoclonal anti-DNA antibodies but also in the serum of several non autoimmune strains (BALB/e. NZB × BALB/e) F1 hybrids & CBA/LH), The antiserum also reacted, but only to a weak degree, with B/W mouse sera. These results indicate that some idiotypes of anti-DNA antibodies produced by autoimmune R W mice are present in normal mouse sera. [ABSTRACT FROM AUTHOR]

MRL/Mp-lpr/lpr (MRL/l) mice spontaneously produce antibodies to poly (ADP-ribose) that are cross-reactive with single stranded DNA (ssDNA). Spleen cells from these animals were used for fusion with murine plasmacytoma cells to prepare hybridomas that produce autoantibodies to poly (ADP-ribose). Monoclonal antibodies (MoAb) produced by the selected hybridomas not only preferred ssDNA to poly (ADP-ribose). hut also reacted with left handed Z-DNA; the MoAb reflected the nature of serum antibodies to poly (ADP-ribose) in MRL/l mice. These results suggest that similar antigenic determinants exist in poly (ADP-ribose). ssDNA and left handed Z-DNA and that the cross-reactive nature of autoantibodies to poly (ADP-ribose) in MRL/1 mice may be the results of expansion of such clones as selected in this experiment. [ABSTRACT FROM AUTHOR]

Phagocytosis of apoptotic neutrophils by macrophages is required for resolution of an inflammatory response. Removal of intact apoptotic neutrophils prevents the release of cytotoxic granules that would otherwise cause tissue damage and may lead to development of fibrosis. Importantly, macrophage phagocytosis of apoptotic neutrophils fails to induce release of proinflammatory mediators, consistent with a `safe' pathway for disposal of potentially harmful inflammatory cells. One pathway for increasing phagocytosis of apoptotic cells to allow matching of tissue phagocyte capacity to apoptotic cell load in vitro is via antibody-mediated cross-linking of CD 44, providing a mechanism for limiting tissue damage during resolution of inflammation. In this study, we have defined divalent cation-dependent and -independent actions of the CD44 antibody. For the divalent cation-independent CD44 antibody effect, we provide evidence that `enabled' CD32 on the apoptotic neutrophil binds to intact CD44 antibody on the macrophage surface. One implication is that macrophages can phagocytose apoptotic neutrophils that are `tethered' to the macrophage surface in a manner that is independent of defined apoptotic mechanisms. These data also provide an explanation for the greater efficacy of intact CD44 antibody when compared with F(ab')2 fragments. [ABSTRACT FROM AUTHOR]

It is generally believed that neutrophils from HIV-infected patients are functionally competent, but several studies have shown impairment in neutrophil fungal killing and cytokine production. In this study we evaluated the ability of neutrophils from healthy donors and HIV-infected patients to produce IL-12 in response to stimulation with Candida albicans, lipopolysaccharide (LPS) and Cryptococcus neoformans (acapsular and encapsulated), with and without MoAb opsonization. Neutrophils from healthy donors secreted IL-12 in response to LPS or C. albicans but not in response to encapsulated or acapsular C. neoformans, regardless of MoAb opsonization. Surprisingly, neutrophils from HIV-infected patients demonstrated constitutive IL-12 production, although these cells were not responsive to LPS stimulation. The inability of MoAb to C. neoformans capsular polysaccharide to promote IL-12 production by neutrophils excludes phagocytosis and/or CD16 cross-linking in this process, and distinguishes neutrophils from monocytes. Our results provide additional evidence for cytokine dysregulation in neutrophils from HIV-infected patients. Furthermore, the IL-12 response of neutrophils and monocytes to CD16 stimulation appears to be different, suggesting differences in the role of these phagocytic cells during the inflammatory response. [ABSTRACT FROM AUTHOR]

Small cytoplasmic ribonucleoproteins (scRNPs) are important autoantigens in patients with systemic lupus erythematosus and Sjögren's syndrome, MoAbs against these proteins were made by immunization of BALB/c mice with purified human recombinant 60-kD Ro/SS-A or 50-kD La/SS-B proteins. Five stable hybridoma cell lines were obtained, of which four secreted anti-Ro/SS-A antibodies (clones ID8. 1D11, 2G10 and 6G8) and one produced anti-La/SS-B antibodies (clone 7E6). The MoAbs were further characterized using four different immunoassays: immunofluorescence. inimunoblotting. RNA precipitation combined with Northern blotting, and recombinant protein precipitation. All four MoAbs against Ro/SS-A recognized the native protein and one of them (2G10) recognized also intact scRNP particles. Interestingly, hY3-RNA was reproducibly not efficiently precipitated by MoAb 2G10. Epitope mapping using deletion mutants of the 60-kD Ro/SS-A antigen showed that MoAb 1D8 recognized the C-terminal part of this protein, while 1D11 and 2G10 recognized distinct epitopes in the region between the RNP motif and the putative zinc finger domain. The epitopes recognized by these MoAbs are highly conserved among species, and the epitope recognized by MoAb 2G10 may be identical to an autoepitope recognized by sera of patients. This is the first report describing the isolation and characterization of MoAbs of the IgG class against the 60-kD Ro/SS-A and La/SS-B autoantigens obtained by immunization with purified human recombinant proteins. These MoAbs can be of great use in studying the cellular processes in which scRNPs are involved, and may help to determine why these scRNPs become autoantigenic in autoimmune diseases. [ABSTRACT FROM AUTHOR]

Monoclonal antibody (MoAb) KT38 raised against a human T leukaemic cell line TALL-1, reacted with another T leukaemic cell line Jurkat, but not with any other cell lines tested. The co-modulation or CD3 and KT38 antigen was observed with stimulations of either MoAb T3 or KT38 on both TALL-1 and Jurkat. Upon radioimmunoprecipitation and SDS-PAGE analysis. MoAb KT38 precipitated the heterodimer of 40-60 kD from Jurkat and TALL-1 under reducing conditions. Thus. MoAbKT38 is considered to bean anti-T idiotype (Ti) antibody to TALL-1 and Jurkat cells. MoAb KT38 was also shown to react with a minor population of peripheral blood lymphocytes (PBL) and with very few cells (0.5-2.0%) in the paracortical area of the lymph node. When PBL were stimulated in a KT38-coated culture flask for 5 days, the percentage of KT38-positive PBL was markedly increased. The CD3 antigen on these cultured PBL in the flask was modulated by the stimulation with MoAb KT38. Thus, it is suggested that a common idiotope exists on the T cell receptor of Jurkat, TALL-1 and a small percentage (1.9-6.1%) of PBL. [ABSTRACT FROM AUTHOR]

An IgG2a monoclonal antibody (MoAb) reacting with the brush border of the renal proximal tubule and glomerular capillary wall was produced by fusion of NSl myeloma cells with spleen cells from BALB/c mice immunized with renal brush border preparations from rat kidney cortex. This antibody reacts with a 90,000 mol. wt protein which can be isolated by immunoprecipitation of radiolabelled brush border or glomerular preparations and localized on these structures by immunoperoxidase electron microscopy, thus demonstrating the presence of common antigenic determinants. Survey of various organs showed that the MoAb reacted with the brush border of the gut, but also with antigens associated with the distal vascular system, in the liver antigenic determinants were located along the sinusoid walls but mainly on bile canaliculi. Specific glomerular binding could be demonstrated in vivo by immunofluorescence after an intravenous injection of 2 mg of antibody or by paired label methodology using tracer amounts. Kinetics however were dramatically different from those observed in classical passive Heymann nephritis since glomerular binding was transient during the first hours after injection. Binding was also found in tubular structures, as well as in lung, liver, spleen and heart. These results identify a well defined antigen antibody system responsible for the formation of transient extramembranous glomerular deposits and may be relevant to some human cases of glomerulonephritis. They may also provide new models to study glomerular and tubular transfer of membrane bound antibodies. [ABSTRACT FROM AUTHOR]

Measurement of erythrocyte [red blood cells (RBC)] complement receptor type 1 (CR1, CD35) has the potential to serve as a sensitive assessment of complement activation and immune complex clearance. All previously reported monoclonal antibodies (MoAb) to the extracellular region of CR1 recognize epitopes within the long homologous repeats (LHR) of CR1 and the epitopes for the most frequently used MoAbs are repeated at least twice per CR1 molecule. Furthermore, CR1 exhibits structural polymorphism characterized by a variable number of LHR per molecule. Thus, accurate enumeration of cell surface CR1 using currently available MoAb would require that the results be corrected for the number of antibody epitopes per CR1 molecule encoded by each individual's alleles. To obtain a MoAb to a non-polymorphic epitope on human CR1, hybridomas were generated from mice immunized with recombinant soluble CR1 (sCR1) and MoAb were screened for those that recognized the full-length extracellular domain but failed to bind to all four recombinant LHR fragments. A single antibody, CR1-2B11, was identified and was found to recognize an epitope located wholly within SCR29-30 of CR1, NH2-terminal to an elastase cleavage site. Like other CR1 MoAb, the CR1-2B11 epitope expression decreased on old erythrocytes compared to younger cells and CR1-2B11 did not identify a CR1 ‘stump’ on RBC. Importantly, CR1-2B11 immunofluorescence did not change with storage or handling of RBC, unlike the apparent decrease in immunofluorescence observed with other MoAb. CR1-2B11 should be useful for the accurate enumeration of RBC CR1. [ABSTRACT FROM AUTHOR]

Production of IgM, IgG and IgA was induced from human blood B lymphocytes by culturing with a CD40 MoAb and IL-2 for 9 days. Replacement of IL-2 by IL-10 markedly enhanced production of all three isotypes. High levels of immunoglobulin production also occurred when activated irradiated autologous T cells replaced the CD40 MoAb, and when IL-10 replaced IL-2 in these cultures a spectacular increase in IgG production occurred. The effectiveness of the T cell stimulus depended on the mode of purification of the T cells and the nature of the stimulant used to activate them. Differences in the kinetics and level of expression of CD40L on the various T cell preparations were observed, but did not account for variations in immunoglobulin-inducing efficiency. Immunoglobulin production from sIgD+ and sIgD- B cells was investigated. IgG and IgA were found in sIgD+ cultures, indicating that some isotype switching had occurred, but the major part of the IgG and IgA secreted was from cells already committed to these isotypes. Anti- IgD or anti-IgM MoAbs enhanced the proliferation of B cells induced by anti-CD40 antibody, but immunoglobulin production was not enhanced. Factors affecting the balance of proliferation and differentiation are discussed. [ABSTRACT FROM AUTHOR]

We analysed human anti-mouse antibodies (HAMA) in 12 patients (six with multiple myeloma (MM) and six with metastatic renal cell carcinoma (MRCC)) who were treated with B-E8, an IgG1 MoAb against IL-6. Efficiency of the treatment was evidenced by the drop in the scrum levels of Creative protein (CRP), the in vivo production of which is under the control of IL-6. Three patients with MM and the six patients with MRCC became immunized to the injected MoAb. HAMA appeared between days 7 and 15 after the beginning of the treatment. The nine patients made IgG antibodies: four also made IgM. All immunized patients made anti-idiotype antibodies specific to B-E8. Two of them also developed HAMA directed to murine IgGI isotype; in these two patients B-E8 MoAb cleared rapidly from the circulation with loss of treatment efficiency. In the patients who developed only anti-idiotype antibodies, serum levels of B-E8 remained unchanged and CRP production remained inhibited, indicating that treatment remained efficient in the presence of HAMA. Circulating B-FS MoAbs were still able to bind to IL-6 and to inhibit IL-6-dependeni proliferation despite the presence of anti-idiotypic HAMA. Therefore, in contrast to HAMA produced against MoAb directed against cellular targets, HAMA against anii-IL-6 MoAb idioiopes led neither to clearance nor to functional in activalion of the injecied MoAb. This was further shown by resuming the B-E8 treatment with success in a patient who still had anti-idiolypic HAMA. [ABSTRACT FROM AUTHOR]

Murine MoAb 1-22-3 has already been reported lo bind lo the mesangial cell surface and to cause transient proteinuria and mesangial morphological changes characterized by mesangiolysis. Subsequent mesangial cell proliferation and mesangial matrix increase by a single i.v. injection. In this study, MoAb-induced glomerulopathy was quantitatively analysed. No correlation between the severity of mesangial morphological changes and the degree of proteinuria was detected (r = 0.190). The minimum dose injected to induce abnormal proteinuria was 25 μg. This dose corresponded to 1.79μg/2 kidneys 30 min after MoAb injection. The highest average level of proteinuria was observed in rats injected with 500 μg of MoAb, and less proteinuria was observed in rats injected with 10.0, 5.0 and 2.0 mg. Although the amounts of kidney-fixing MoAb and the subsequent deposition of rat C3 in the high-dose-injected group were larger than in the 500 μg injected group, the numbers of infiltrating inflammatory cells were the same in both groups. No correlations between the degrees of such mediators and proteinuria were observed. [ABSTRACT FROM AUTHOR]

Quantitative measurements of cultured human and murine thymic, and human thymoma reticulo-epithelial cells (REC), immunolabelled by different antibodies (Ab) (TE3, TE4, anti-HTLV p19(p19), lu5, K11 and Aks) and by thymic hormones (thymulin and thymosin alpha 1(Ta1)) within these cells, were performed using a flow cytometric technique. The anti-keratin polyclonal Ab labelled nearly the whole human or murine population. The p19 monoclonal Ab (MoAb), specific for the subcortical/medullary thymic regions, labelled 37-77% of the human REC. The TE3 MoAb, specific for the cortical region, labelled 54-83% of the REC. These percentages suggest that the cultured thymic REC (TREC) had markers of both regions together and therefore that these markers are not absolutely specific to determine their subcortical/medullary or cortical thymic origin. For the three populations there were more cells containing Ta1 than thymulin. The overlap of the percentage of labelled cells suggests that the same cell could synthesize the two hormones and that these hormones could be localized within the TE3 positive cells. [ABSTRACT FROM AUTHOR]