Quantitation of β2-Microglobulin and HLA on the Surface of Human Cells

Relative amounts of beta2-microglobulin (beta2m) and of HLA specificities were analysed on the surface of resting unfractionated peripheral human lymphocytes, enriched B and T cells, and on in-vivo- and in-vitro-stimulated lymphoblasts. Single-cell cytofluorometry and a very sensitive radioimmunoassay were used to determine as closely as possible the absolute amounts of membrane-bound beta2m/HLA antigens. B and T "resting" and "stimulated" lymphoid cells express very similar numbers of beta2m and HLA antigenic determinants, respectively, per unit of surface area when compared with each group, although beta2m was found to exist in two- to three-fold excess of HLA.     

Increased expression of mucosal addressin cell adhesion molecule-1 (MAdCAM-1) and lymphocyte recruitment in murine gastritis induced by Helicobacter pylori

Although T cell involvement in Helicobactor pylori-induced gastritis is known, mechanism about T cell recruitment is not understood. In this study we examined how mucosal addressin cell adhesion -molecule-1 (MAdCAM-1) is involved in lymphocyte recruitment in murine chronic gastritis induced by H. pylori. C57 BL/6 mice were infected with Sydney strain (SS1). Six months after infection, the stomach was removed. The expression of adhesion molecules, MAdCAM-1, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), and the cell surface antigens CD4, CD8, CD45R/B220 or beta7-integrin were determined by immunohistochemistry. A significant increase in CD4 lymphocytes was observed in the body portion of stomach in SS1-infected mice and most of these CD4 cells express beta7-integrin, a known counter ligand for MAdCAM-1 molecule. Strong MAdCAM-1 expression was observed adjacent to these cells in the lamina propria as well as in the submucosa of SS1-infected stomach. Quantitative analysis showed that the area of MAdCAM-1 expression well correlated with the infiltration of beta7-integrin positive lymphocytes. On the other hand, expression of ICAM-1 or VCAM-1 in the lamina propria was few even in the SS1-infected stomach. Increased expression of MAdCAM-1 was well correlated to the location of lymphocytes, which express CD4 and beta7-integrin. These results suggest the possibility that MAdCAM-1 may be largely involved in the lymphocyte recruitment in the gastritis mucosa with H. pylori.     

Increased expression of collagen receptors: α1β1 and α2β1 integrins on blood eosinophils in bronchial asthma

BACKGROUND: Eosinophils are one of the major effector cells in bronchial asthma. Their infiltration of airways correlates with the asthma severity. Recruitment and activation of eosinophils are partially mediated by integrins alpha4beta1 and alpha4beta7. Collagens type I and IV constitute important components of extracellular matrix and vascular basement membrane, respectively. Therefore, collagen-binding integrins (alpha1beta1 and alpha2beta1) may also play a role in eosinophil lung infiltration. OBJECTIVE: To evaluate the possible presence of alpha1beta1 and alpha2beta1 integrins on peripheral blood eosinophils from asthmatic subjects. METHODS: Collagen receptors were studied on eosinophils separated by immunomagnetic CD16-negative method from healthy donors (n=13) and patients with moderate persistent atopic bronchial asthma (n=15). Surface receptor identification was performed by flow cytometry and cell adhesion assay. RESULTS: Eosinophils isolated from the patients showed increased expression of both alpha1beta1 and alpha2beta1 integrins as compared with healthy controls. Moreover, adhesive function of eosinophils to collagen type IV was inhibited by snake venom disintegrins: viperistatin and obtustatin. These disintegrins contain KTS active motif and are specific inhibitors of alpha1beta1 integrin. CONCLUSION: We demonstrated for the first time that collagen receptors: alpha1beta1 and alpha2beta1 integrins are overexpressed on the surface of peripheral blood eosinophils of asthmatic subjects. Further studies may reveal potential application of KTS-disintegrins or their structural analogs for therapy of bronchial asthma.     

Mitogenic Effect of α1-Microglobulin on Mouse Lymphocytes

Human alpha 1-m microglobulin (alpha 1-m), a low molecular weight plasma protein, was found to exert mitogenic effects on mouse lymphocytes from lymph nodes and spleen. The stimulatory effects appeared to be strain-restricted: alpha 1-m induced a varying degree of proliferation of lymphocytes from three strains, whereas one strain responded poorly. Experiments with lymphocyte subpopulations showed only weak stimulatory effects of alpha 1-m on purified T and B lymphocytes cultivated alone. The addition of mitomycin-treated cells of the other subpopulation could not restore the proliferative responses in either T or B lymphocytes. Strong stimulations were recorded only when both T and B lymphocytes were present, indicating that the T and B lymphocytes cooperate to achieve the proliferation. However, FACS studies on cultured splenocytes indicated that the proliferating cells are predominantly B lymphocytes. These data extend our earlier findings of a mitogenic effect of alpha 1-m on guinea pig lymphocytes. Furthermore, results were obtained indicating the presence of a receptor on mononuclear cells. Iodine-labelled alpha 1-m was bound to mononuclear cells prepared from spleens, and the binding could be blocked by an excess of non-labelled alpha 1-m. Scatchard plotting of the data gave an equilibrium constant of 0.7 x 10(5)/M for the binding between alpha 1-m and the receptor. Together with the documented inhibitory activity of alpha 1-m on antigen-driven proliferation of lymphocytes, these results suggest an immunoregulatory role for alpha 1-m.     

Costimulation of CD28− T Cells Through CD3 and β1-Integrins Induces a Limited Th1 Cytokine Response

The costimulatory molecule CD28 regulates antigen-specific T-cell proliferation and the synthesis of multiple cytokines. The absence of CD28 on a subset of CD8bright+ T cells suggests that these cells may utilize alternative costimulatory pathways or have a limited cytokine response to presented antigen. We used fibronectin, a ligand for the beta1-integrins alpha4beta1 and alpha5beta1, as an alternate costimulatory ligand to assess the functional phenotype of CD8bright+CD28- T cells. CD25 expression was significantly up-regulated in CD8bright+CD28- T cells by immobilized anti-CD3i with fibronectin. Costimulation with fibronectin also significantly augmented anti-CD3i-induced IFN-gamma production only among CD8bright+CD28- T cells. The CD8bright+CD28- T cells did not produce significant IL-2 and IL-10 even in response to maximal stimulation with phorbol myristate acetate and ionomycin. These data support a costimulatory role for ss1-integrins in CD8bright+CD28- T cells and indicate that CD8bright+ CD28- T cells have a restricted Th1 cytokine repertoire.     

Lymphocyte Associated β2-Microglobulin Studied by Crossed Radioimmunoelectrophoresis

A sensitive radioimmunoelectrophoretic assay was employed in the study of beta 2-microglobulin (beta2m) from human blood lymphocytes, allowing the detection of 2.5 pmol of beta2m corresponding to 3 x 10(6) lymphocytes. The intrinsic (amphiphilic) membrane proteins were solubilized and examined in crossed immunoelectrophoresis against rabbit antisera in the presence of non-ionic detergent. The beta2m associated precipitate was traced by post-electrophoretic incubation with 125I-labelled antibodies to beta2m and autoradiography. A polyspecific rabbit antiserum to human lymphoid cells retained its capacity to precipitate lymphocyte beta2m after absorption with isolated beta2m (on an immunosorbent column), showing that lymphocyte beta2m is complexed to other molecules. No free beta2m was found on lymphocytes when examined against the absorbed antiserum to human lymphoid cells in an intermediate gel and anti-beta2m in a reference gel.     

Participation of CD11a–c/CD18 and RGD-Recognizing Adhesion Molecules in the Binding of LGL to Fibroblasts

We have previously shown that large granular lymphocytes (LGL) are inactivated by contact with natural killer (NK) resistant monolayer target cells. In this work we have analysed which adhesion molecules are involved in the binding of LGL to such targets, as exemplified by fibroblasts, and in the subsequent inhibition of their NK activity. The results indicate that antibodies against CD54 (intercellular adhesion molecule 1, ICAM-1), CD11a (leucocyte function antigen 1, LFA-1, alpha chain), and CD18 (common beta chain of the beta 2-integrin family) significantly (by 50%) reduce the binding of LGL onto inhibitory target cells. The matrix protein-based synthetic peptide RGD and anti-CD29 (the common beta chain of the beta 2-integrin family) antibodies also diminish the binding (by 35%). The effects of the antiadhesion molecule antibodies and the peptide are additive, the combination of both leading to an almost complete block of adhesion. It may be hypothesized that some of the binding-relevant adhesion molecules of the RGD-binding domain on LGL (CD29) may be involved in the delivery of the inactivating signal to the effector cell. Indeed, incubation of LGL with anti-CD11a antibodies, but neither with antibodies against other binding-relevant epitopes nor with RGD, significantly reduced their NK activity. The mechanism of the inactivation was similar to that induced by intact NK-resistant target cells. On the basis of the present results we suggest that the CD11a molecule is involved in the down-regulation of the NK activity of peripheral blood lymphocytes.     

Activated CD3−CD16+ Natural Killer Cells Express a Subset of the Lymphokine Genes Induced in Activated αβ+ and γω+ T cells

In this study we analysed the potential of highly purified polyclonal TcR alpha beta+, TcR gamma delta + and CD3- NK cells, to produce lymphokines in response to mitogenic stimulation. RNA hybridizations were performed to detect with high sensitivity the induction of multiple lymphokine genes. Upon stimulation with lectin and phorbol ester TcR gamma delta + lymphocytes expressed the same set of lymphokine genes as the TcR alpha beta + lymphocytes expressed the same set of lymphokine genes as the TcR alpha beta + lymphocytes, which included IL-2, -3, -4, -5, GM-CSF, TNF alpha and beta, IFN gamma. In contrast, a more limited set of lymphokine genes (GM-CSF, TNF alpha and beta, IFN gamma) was induced in activated CD3- NK cells, thus indicating that this subpopulation of cells may display different regulatory functions, with respect to CD3+ T lymphocytes.     

Studies on the Cytophilic Properties of Human β2-Microglobulin

The mechanism by which exogenously added beta2m binds to lymphoid cells has been explored. In the mouse it has been shown that beta2m remains associated with plasma membrane macromolecules following solubilization with NP-40 and that approximately 25-30% of the binding could be accounted for by binding to H-2 antigens. No binding to mouse immunoglobulin or Ia antigens could be detected. The sites for binding of the remainder of the cell-bound beta2m were not determined. Whereas normal human lymphocytes showed little or no capacity to bind exogenously added beta2m, it was found that phytohaemagglutinin (PHA)-stimulated cells could bind beta2m. This binding occurred optimally 2 days after PHA stimulation. Approximately half of the binding could be accounted for by binding to HLA antigens. The possible significance of these findings with respect to cellular interactions involving major histocompatibility complex gene products in the immune response is discussed.     

Expression of the LFA-1 β2 Integrin (CD11a/CD18) and ICAM-1 (CD54) in Normal and Coeliac Small Bowel Mucosa

The leucocyte adhesion molecules (beta 2 integrins) comprise CD11 alpha-chains and a common beta-chain (CD18). CD11a (leucocyte function-associated antigen 1, LFA-1) is expressed by most T cells, and is involved in antigen presentation by macrophages via its counter-receptor, intercellular adhesion molecule (ICAM-1, CD54). By criteria of double-label immunofluorescence of cryostat tissue sections, virtually all lamina propria T cells of the normal small bowel were found to express LFA-1 strongly. By contrast, only 30-60% of intra-epithelial lymphocytes (IEL) expressed detectable LFA-1, most of which were LFA-1 weak and CD18-. ICAM-1 was expressed strongly only by vascular endothelium. In coeliac disease, there was a modest increase of diffuse ICAM-1 expression in the lamina propria, mainly in the subepithelial zone, where ICAM-1+ macrophages were occasionally seen. There was also a slight overall increase in CD11a expression by IEL, seen predominantly in surface epithelium and mainly by the CD4+ minority subset, but not by CD4-CD8- (TcR gamma delta +) cells. These data suggest that the LFA-1/ICAM-1-dependent antigen presentation pathway is of minor importance to IEL in the normal small bowel, and does not assume a major role in coeliac disease.     

Analysis and Characterization of the Interleukin 2 Receptor α and β Chain Expression inCD4−CD8− Cells

The differential effects that the binding of interleukin 2 (IL-2) to its beta or alpha beta receptors might induce in two different CD4-CD8- T-cell lines were analysed. While LD1.T3b, a double-negative T cell derived from MRL/lpr mice, constitutively expressed high levels of the IL-2R beta chain, YAC-1, a Moloney sarcoma virus-transformed CD4-CD8- T cell, expressed (as an activated T cell) the beta and alpha chains. The presence of IL-2 in the culture medium was lethal for LD1.T3b cells, while it had no effect on the growth of YAC-1 cells. IL-2 increased the expression of the beta chain and, to a lesser extent, of the alpha chain in YAC-1 cells. In addition, other markers such as CD4 and CD5 were induced by IL-2 in this cell line.     

Rabbit Anti-Human β2 Microglobulin Antibodies: A T-Cell Mitogen

Rabbit anti-beta 2 microglobulin antibodies have been described as a potent mitogen for human B lymphocytes. However, when fractionated after activation, only the T-cell enriched population is actively dividing. The induction of proliferation in purified T cells requires the presence of non-T cells. Daudi cells (which do not express beta 2 microglobulin on their cell surface) were shown to produce 'macrophage replacing factors' which supported anti-beta 2-induced T cell division. Non-T cells could be effectively replaced by addition of interleukin 2 containing cell supernatants, 12-O-tetradecanoyl-phorbol-13-acetate (TPA), or teleocidin B. Thus, anti-beta 2 appears to selectively activate T cells. In contrast to other T cell mitogens, anti-beta 2 microglobulin antibodies did not induce secondary immunoglobulin production in B cells from peripheral blood or spleen.     

β2-Microglobulin from Normal and Leukaemic Guinea-Pig Lymphocytes

beta2-Microglobulin has been isolated in useful quantities from the urine of strain-2 guinea-pigs after either treatment with sodium chromate or induction of the L2C leukaemia. Antibodies raised against the beta2-microglobulin were used to set up a radioimmunoassay which measured its export into culture fluid by normal and leukaemic lymphocytes. Material containing beta2-microglobulin was also obtained by digestion of the lymphocytic surfaces with papain; fractionation demonstrated both free and combined forms, with no qualitative difference between those from normal and those from leukaemic cells.     

Altered β2-adrenergic regulation of T cell activity after allergen challenge in asthma

BACKGROUND: Airway inflammation in asthma is orchestrated by recruitment of T helper (Th)2 lymphocytes to the lung and subsequent production of Th2-like cytokines upon allergen challenge. OBJECTIVE: To examine whether allergen-induced dysfunction of the beta2-adrenergic receptor (beta2-AR) contributes to the enhanced T(h2) cell activity in asthma. METHODS: Beta2-adrenergic regulation of cytokine mRNA expression was studied in alpha-CD3/alpha-CD28-activated peripheral blood lymphocytes from seven asthma patients before and 6 h after allergen challenge, in conjunction with the effects of beta2-agonist fenoterol on T cell chemotaxis and signalling pathways. RESULTS: A complete loss of beta2-AR control over expression of the Th2 cytokines IL-4, IL-5 and IL-13, but not of the Th1 cytokine IFN-gamma, was observed after allergen challenge. Furthermore, we found impaired beta2-AR regulation of T cell migration as well as signal transduction pathways, i.e. the phosphorylation of cyclic adenosine monophosphate-responsive element binding protein and the inhibition of the mitogen-activated protein kinase pathway. The loss of beta2-AR control was associated with increased beta-adrenergic receptor kinase expression, which might be involved in beta2-AR desensitization. In addition, we demonstrate for the first time that T cells exposed to the chemokine thymus and activation-regulated chemokine show hyporesponsiveness to fenoterol. CONCLUSION: Our results suggest that allergen-induced loss of beta2-AR control, possibly mediated by chemokine release, plays an important role in enhanced Th2-like activity in asthma.     

Activation of Human T and B Cells by Rabbit Anti-Human β2-Microglobulin

Rabbit anti-human β₂-microglobulin (anti-β₂m) increased the highest DNA synthesis in un-separated lymphocytes or artificially composed mixtures enriched in T and B cells. In enriched T and B cells no or low stimulation was seen. The maximal response by IgG anti-β₂m was seen in proportions of enriched T/B cells, being 3:1 for blood lymphocytes and 1:1 for spleen cells, which are the same as the physiological proportions of T and B cells in these lymphoid organs. Whereas unseparated lymphocytes gave a peat response on day 3, enriched B cells had a peak response on day 6. Fab anti-β₂m did not activate enriched T cells but increased DNA synthesis in enriched B cells to about the same extent as unseparated lymphocytes and mixtures of enriched T and B cells. The proportion of sheep erythrocyte rosette-forming cells (E-RFC) decreased after stimulation by anti-β₂m and increased after stimulation by phytohaemagglutinin. However, as revealed by autoradiography, a proportion of lymphocytes activated by anti-β₂m were E-RFC and this proportion increased with increasing stimulation by anti-β₂m. DNA synthesis induced by anti-β₂m was unchanged for spleen cells and slightly decreased for blood lymphocytes when phagocytic cells were removed by iron treatment. Supernatants from lymphocytes activated by anti-β₂m only induced low DNA synthesis in enriched T or B cells. Experiments with mitomycin treated cells indicate that cooperation and close contact between T and B cells are needed for activation by IgG anti-β₂m. T cells are needed for B cell activation by anti-β₂m and B cells are required for T cell activation to occur.     

Differential Expression of T-Cell Adhesion Molecules and LFA-1-Dependent Intercellular Adhesion in HgCl2-Induced Autoimmunity and Immune Suppression

Exposure of Brown Norway (BN) rats to HgCl₂ induces Th2-mediated systemic autoimmunity. In contrast, in Lewis rats, HgCl₂ induces immune suppression, mediated by CD8⁺ T cells. HgCl₂ was previously found to enhance expression of LFA-1, ICAM-1 and CD134 (OX40) on T cells in BN rats. In the present study, T cells from Lewis rats were studied at day 4 after injection of HgCl₂. CD8⁺ T lymphoblasts were significantly increased, which were predominantly CD45RC^hi, and which showed enhanced LFA-1 expression. Furthermore, CD4⁺CD45RC^hi T cells showed increased numbers of ICAM-1⁺ cells, whereas expression of CD134 and CD26 was relatively decreased in CD4⁺ T lymphoblasts. Ex vivo experiments demonstrated that HgCl₂- exposure of BN rats, but not of Lewis rats, significantly enhances PMA [phorbol 12-myristate 13-acetate]-induced lymphocyte aggregation, mediated by LFA-1 and ICAM-1. In conclusion, HgCl₂-injected Lewis rats show early signs of T-lymphocyte activation, predominantly on CD8⁺ cells. Strain-dependent effects of HgCl₂ on cell adhesion molecules and expression of CD134 may play an important role in development of either autoimmunity or immune suppression.     

Roles of alpha(4) integrins/VCAM-1 and LFA-1/ICAM-1 in the binding and transendothelial migration of T lymphocytes and T lymphoblasts across high endothelial venules

Several cell adhesion molecules that mediate the binding of lymphocytes to high endothelial venules (HEV) from flowing blood have been identified but the regulation of lymphocyte migration across the HEV wall into the lymph node (LN) is far from understood. In this study we have used an in vitro model of lymphocyte migration across HEV, and analysed the roles of two integrins in the binding and transendothelial migration of T lymphocytes and T lymphoblasts. The adhesion of T lymphocytes to high endothelial cells (HEC) cultured from rat LN HEV differed from that of T lymphoblasts since the percentage of T lymphoblasts that adhered and transmigrated was higher and was not increased by IFN-gamma pretreatment of HEC. Antibodies to alpha(4) integrins, VCAM-1 or LFA-1 maximally inhibited T lymphocyte adhesion by 40-50%, whereas antibodies to ICAM-1 were less effective (<20% inhibition). The effects of alpha(4) integrin and LFA-1 antibodies were additive, giving >90% inhibition. T lymphocytes which adhered in the presence of LFA-1 antibody showed reduced levels of transmigration and, in the presence of alpha(4) integrin antibody, slightly increased transmigration. Antibodies to alpha(4) integrins, VCAM-1, LFA-1 or ICAM-1 had little effect on T lymphoblast adhesion (maxima of 10-30% inhibition) and T lymphoblasts transmigrated normally in the presence of either alpha(4) integrin or LFA-1 antibodies. However, the effects of alpha(4) integrin and LFA-1 antibodies on T lymphoblast adhesion were synergistic, giving >90% inhibition of adhesion. These results suggest that the majority of T lymphoblasts use either alpha(4) integrins or LFA-1 to bind and transmigrate HEV, and the roles of these integrins on activated T cells are overlapping and redundant. In contrast, either integrin supports half-maximal binding of unactivated T lymphocytes to the surface of HEV and LFA-1 makes a larger contribution than alpha(4) integrins to transendothelial migration.     

ICAM-1/LFA-1 interactions in T-lymphocyte activation and adhesion to cells of the blood-retina barrier in the rat.

To identify the signals involved in the adhesion and subsequent migration of lymphocytes across the endothelium (REC) and pigment epithelium (RPE) of the blood-retina barrier we have studied the effects of monoclonal antibodies (mAb) to rat adhesion/accessory molecules on the binding of normal and concanavalin A (Con A)-activated rat spleen lymphocytes to cultured unstimulated and interferon-gamma (IFN-gamma)-stimulated RPE and REC. Forty to 48% of unactivated T cells were found to bind to normal REC or RPE by leucocyte function-associated antigen-1/intercellular adhesion molecule-1 (LFA-1/ICAM-1)-independent mechanisms, despite constitutive expression of ICAM-1 by the RPE cells and LFA-1 by the T cells. Con A-activated lymphocytes showed an enhanced adhesion to both RPE and REC. However, IFN-gamma-stimulated RPE and REC did not demonstrate a significant increase in adhesiveness for normal lymphocytes highlighting the importance of lymphocyte integrin activation from low-affinity to high-affinity state. Activated lymphocyte adhesion to unstimulated RPE and REC was significantly blocked by LFA-1 mAb (35%, P < 0.0001) and ICAM-1 mAb (20%, P < 0.001). Inhibition of adhesion by antibody to CD2 was not significant. Both ICAM-1 and LFA-1 mAb also significantly (P < 0.05) blocked antigen presentation following retinal extract stimulation of lymphocytes from immunized rats in proliferation assay. These data suggest that the ICAM-1/LFA-1 system is important in lymphocyte trafficking into the eye only after lymphocyte activation.     

Target Cell Lysis by Natural Killer Cells is Influenced by β2-Microglobulin Expression

Natural killer (NK) cells form part of the vertebrate defence against viruses and tumours, but show only limited specificity. The molecule(s) recognized by NK cells on target cells are at present unknown. Major histocompatibility complex (MHC) class I antigen concentration on target cells is inversely correlated with NK cell lysis. Here we show that MHC class I-unassociated beta 2-microglobulin (beta 2-m) expression is involved in NK cell-target cell interaction. Two human MHC class I negative cell lines, Daudi and K562, are differentially susceptible to NK cell lysis. Daudi cells are beta 2-m-negative and resistant to NK lysis, K562 are beta 2-m-positive and highly susceptible to lysis by NK cells. Interferon (IFN) treatment augments beta 2-m expression and NK lysis of K562, but not in Daudi cells. NK cell lysis of K562, but not YAC-1 cells, can be inhibited by monoclonal anti-human beta 2-m antibody. Furthermore, susceptibility of mouse embryo fibroblasts (MEF) to NK lysis can be increased by infection with recombinant vaccinia virus expressing the human beta 2-m gene.     

Mitogenic Properties of Fab and F(ab'')2 Fragments of Rabbit Anti-Human β2-Microglobulin for Human Lymphocytes

Bivalent F(ab')2 fragments and monovalent Fab fragments of rabbit anti-human beta2-microglobulin (anti-beta2m) stimulated DNA synthesis in human lymphocytes. Mitogenicity of anti-beta2m antibodies can therefore be ascribed to the antigen-binding site and not to the Fc portion of the molecule. The mitogenic response to F(ab')2, and sometimes Fab, fragments of anti-beta2m IgG was comparable to that obtained with original IgG antibodies when tested at the same protein concentration. Since Fab monomers of anti-beta2m can cause lymphocyte activation, 'cross-linking' of hypothetical beta2-microblobulin-containing lymphocyte receptors does not seem necessary for activation. F(ab')2, as well as Fab, fragments of anti-beta2m blocked the cytotoxic effect of anti-beta2m IgG, showing that the fragments did indeed react with beta2-microblobulin on the cell surface. F(ab')2 dimers, but not Fab monomers, of anti-beta2m were capable of inhibiting the cytotoxic effect of an anti-HLA-A2 antiserum. The mitogenic activity of both anti-beta2m IgG and Fab monomers of such antibodies disappeared after absorption with highly purified beta2-microblobulin. The mitogenic effect of anti-beta2m IgG was inhibited to a minor extent by exposure of cells to high concentrations of pooled multispecific anti-HLA antibodies. This effect was probably nonspecific.