Expression of the PD-1 antigen on the surface of stimulated mouse T and B lymphocytes

A mAb J43 has been produced against the product of the mousePD-1 gene, a member of the Ig gene superfamily, which was previouslyisolated from an apoptosis-induced T cell hybridoma (2B4.11)by using subtractive hybridization. Analyses by flow cytometryand immunoprecipitation using the J43 mAb revealed that thePD-1 gene product is a 50–55 kDa membrane protein expressedon the cell surface of several PD-1 cDNA transfectants and 2B4.11cells. Since the molecular weight calculated from the aminoacid sequence is 29,310, the PD-1 protein appears to be heavilyglycosylated. Normal murine lymphoid tissues such as thymus,spleen, lymph node and bone marrow contained very small numbersof PD-1⁺ cells. However, a significant PD-1⁺ population appearedin the thymocytes as well as T cells in spleen and lymph nodesby the in vivo anti-CD3 mAb treatment. Furthermore, the PD-1antigen expression was strongly induced in distinct subsetsof thymocytes and spleen T cells by in vitro stimulation witheither anti-CD3 mAb or concanavalin A (Con A) which could leadT cells to both activation and cell death. Similarly, PD-1 expressionwas induced on spleen B cells by in vitro stimulation with anti-IgMantibody. By contrast, PD-1 was not significantly expressedon lymphocytes by treatment with growth factor deprivation,dexamethasone or lipopolysaccharide. These results suggest thatthe expression of the PD-1 antigen is tightly regulated andinduced by signal transduction through the antigen receptorand do not exclude the possibility that the PD-1 antigen mayplay a role in clonal selection of lymphocytes although PD-1expression is not required for the common pathway of apoptosis.     

Detection of capsid antigen of human papillomavirus (HPV) in benign lesions of female genital tract using anti-HPV monoclonal antibody.

We established a murine monoclonal antibody (K1H8) to human papillomavirus (HPV) using alkaline-disrupted virions of HPV type 1 (HPV-1) as the immunogen. K1H8 recognized a 57 kD capsid protein of HPV-1 and detected the antigen in paraffin sections of formalin-fixed tissue. With K1H8, we examined immunohistochemically 68 biopsy specimens obtained from the female genital tract. The specimens were histologically condyloma acuminatum or koilocytotic lesions with or without dysplasia and each specimen was found to harbour a single type of genital HPV, such as types 6, 11, 16, 18, 31, 33, 42, 51, 52, 56, and 58, by Southern blot hybridization analysis. The antigen was localized in the nuclei and occasionally in the cytoplasm of squamous cells showing koilocytotic changes. Eighty-four per cent of the specimens (57 cases) showed positivity for the antigen, indicating that K1H8 is a broadly-reactive antibody to various genital HPVs. The results suggest that benign mucosal lesions of the female genital tract are more frequently associated with viral production and are a potential source of transmission.     

Rabies virus binding at neuromuscular junctions

Morphological, immunocytochemical, biochemical, and immunological techniques have been used to describe rabies virus binding to a sub-cellular unit and molecular complex at the neuromuscular junction (NMJ). Early after infection in vivo, virus antigen and virus particles were found by immunofluorescence, electron microscopy and immunoelectron microscopy in regions of high density acetylcholine receptors (AChR) at NMJs. One monoclonal antibody (alpha-Mab) to the alpha subunit of the AChR blocked attachment of radio-labeled rabies virus to cultured muscle cells bearing high density patches of AChR. A sub-cellular structure, resembling an array of AChR monomers, bound both rabies virus antigens and alpha-Mab. By immunoblotting with electrophoretically transferred motor endplate proteins, rabies virus proteins and alpha-Mab bound to two proteins of 43 000 and 110 000 daltons. A rabies virus glycoprotein antibody detected virus antigen bound to the 110 000 dalton protein. An auto-immune (anti-idiotypic) response followed immunization of mice with rabies virus glycoprotein antigen; the antibody was directed to the 110 000 dalton protein. This auto-antibody altered the kinetics of neutralization by rabies virus antibody and induced the formation of rabies virus antibody after inoculation of mice. These results define, at the neuromuscular junction, a rabies virus receptor which may be part of the acetylcholine receptor complex.     

Suppressive effects of Sairei-to on monoclonal antibody 1–22–3-induced glomerulonephritis: Analysis of effective components

The effects of treditional Chinese medlclne (Salrel-to) on experimental glomerulonephritls Induced In rats by monodonal antibody (mAb) 1–22–3 lnjectlon was examined. The level of proteinuria in the Sairel-to-treated group was significantly lower than that In the PBS treated group. This suppressive effect was caused by the major component of Sealer-to, Syo-salko-to but not by another component, Gorel-san. The suppressive effect of Syo-salko-to was Identified In Its components ( Bupleuri radix, Pindilae tuber and Zingibers rhizoma ), but not In the other combined components ( Ginseng radix and Zizyphl fructus ). Further study weeled that the suppressive effects of the combined components were mainly derived from Bupleuri radix . It was demonstrated that the actual active Ingredient is probably Salkosaponin-d. Light microscopy revealed that Sairel-to and Its effective components suppressed the proliferation of mesanglal cells and mesanglal matrix expansion. Semi quantitative morphological studies of glomerular lesions on the eighth day showed that Syo-salko-to and Its combined components ( Bupleuri radix, Zinglberis rhizoma and Pinelliae tuber ) suppressed mesanglal matrix expansion significantly compared with phosphate-buffered saline control groups (matrix score: 28.0±19.1 vs 102.3±14.1; 30.9±30.1 vs 102.3±14.1, p<0.005, respectively). It was concluded that Salkosaponln-d, as well as Bupleuri radix , Syo-salko-to and Sairel-to can suppress proteinurla and morphological changes In the rat glomerulonephritls model Induced by mAb 1–22–3.     

HLA-DR residues accessible under the peptide-binding groove contribute to polymorphic antibody epitopes

Many residues involved in polymorphic antibody-binding epitopes on class II molecules are located on the -helix of DRβ chains. Although they have received less attention, residues in the peptide-binding groove and second domain of the DRβ chain may also be critical for polymorphic anti-DR antibody epitopes. In this study, we used transfectants expressing site-directed mutations at positions in the HLA-DR β₁ and β₂ domains and flow cytometry to define the epitopes of several polymorphic anti-DR antibodies. Both DR(β 1^*0403) residues 14 and 25 were shown to be involved in the epitopes of mAbs DA6. 164, HU-20, Q5/6, and 50D6, and DR(β 1^*0701) residue 14 was shown to be critical for the epitopes of two DR7-specific mAbs, SFR16-DR7M and TAL 13. 1. Unlike most other residues shown to be important in antibody-binding epitopes, residue 14 is located in the floor of the peptide-binding groove and residue 25 is in an outer loop, each with their side chains pointing down, such that antibodies may directly contact these residues from below the binding groove. Two residues in the β₂ domain, β180 and β181, were also shown to be involved in the epitopes of three polymorphic anti-DR mAbs, NFLD.D1, NFLD.M1, and LY9. Although these two residues are close to the transmembrane domain in the linear sequence, their solvent accessibility in the DR1 structures is quite impressive. Our data provide new evidence that residues accessible under the peptidebinding groove contribute to polymorphic antibody-binding epitopes.     

Immunohistochemical characterization of an anti-epithelial monoclonal antibody (mAB lu-5)

Summary A mouse monoclonal antibody (mAB lu-5) was prepared using a lung cancer cell line as an antigen. The selected clone produces an IgG with a gamma-1 heavy chain and a kappa-light-chain. Immunohistochemical testing of mAB lu-5 on 117 normal tissue biopsies and 474 tumours revealed reactivity with an intracytoplasmic, formaldehyderesistant antigen present in most epithelial and mesothelial cells, but absent in mesenchymal cells. The antibody can therefore be used as a first order, pan-epithelial marker. It proved also useful for fast tumour diagnosis on frozen sections.     

Immune response of adults to sequential influenza vaccination

Annual immunization against influenza is recommended for numerous individuals, but the antibody response to sequential vaccination has not been well characterized. Levels of hemagglutination-inhibition antibody were measured in adults given either two or three doses of trivalent influenza vaccine at six-month intervals. A significant rise in the number of individuals with antibody titers of greater than or equal to 40 was seen for all three antigens only after initial vaccination. Repeated vaccination was necessary to maintain adequate antibody levels only to the A/Brazil (H1N1) antigen; it did not significantly affect the proportions of individuals with protective levels of antibody to either the A/Bangkok (H3N2) or the B/Singapore 222/79 antigens. These findings do not support the current recommendation for annual immunization when the vaccine formulation has not changed.     

Correlation Between Regional Specificity of Antisperm Antibodies to the Spermatozoan Surface and Complement-Mediated Sperm Immobilization

ABSTRACT: Sera from men at risk for immunity to spermatozoa were screened for antisperm antibodies by immunobead binding following passive antibody transfer to antibody-free sperm of fertile donors. The percent motile sperm after incubation in diluted antibody positive serum in the presence of complement was compared with the regional distribution of immunoglobulins bound to the sperm surface. The extent of complement-mediated sperm immobilization varied with immunoglobulin class and with the location of antibody bound to the sperm surface. Tests utilizing complement-mediated immobilization of sperm are insensitive to the presence of antibodies of IgG and IgA classes that are directed against the head, the distal one-fifth of the sperm tail principal piece, or the tail end piece. A high degree of immobilization was found only when IgG binding occurred on the distal two-fifths to three-fifths of the principal piece of the tail or when IgM bound to the sperm tail end piece.     

Reduction of latex-allergen content in Swedish medical catheter balloons – a survey of 3 years' production

Anaphylactic reactions after intravascular exposure to natural rubber latex (NRL) have been reported. Thus, there is an urgent need to produce medical devices with the lowest possible latex-allergen content. The latex-allergen concentration in extracts prepared from 92 lots of medical catheter (MC) balloons, manufactured by Nolato Polymer AB, Torekov, Sweden, from April 1993 to March 1996, was measured with an EAI (IgE antibody inhibition) assay. Inhibitory capacity was expressed in arbitrary units/ml (U/ml) in relation to reference NRL sap, given an arbitrary value of 1000 U. Extracts from randomly selected lots were measured for protein by the modified Lowry method. Water leaching, chlorination, and treatment with savinase were used experimentally to study reduction of the latex-allergen content. The latex-allergen content in extract from the regular MC balloons varied from 0.1 to 2.9 U/ml. All the methods used to reduce the allergen content were effective, and increased leaching stabilized the allergen content at a low level. The protein concentration of the extracts varied between 9 and 100 mg/1. No correlation was found between protein and allergen content. As a result of this study, the manufacturer has extended the stage of water leaching in the production process. This study shows that cooperation between immunologists and manufacturers may result in product development and improvement.