Showing papers on "Antibody published in 1983"

Isolation of a T-lymphotropic retrovirus from a patient at risk for acquired immune deficiency syndrome (AIDS).

Abstract: A retrovirus belonging to the family of recently discovered human T-cell leukemia viruses (HTLV), but clearly distinct from each previous isolate, has been isolated from a Caucasian patient with signs and symptoms that often precede the acquired immune deficiency syndrome (AIDS). This virus is a typical type-C RNA tumor virus, buds from the cell membrane, prefers magnesium for reverse transcriptase activity, and has an internal antigen (p25) similar to HTLV p24. Antibodies from serum of this patient react with proteins from viruses of the HTLV-I subgroup, but type-specific antisera to HTLV-I do not precipitate proteins of the new isolate. The virus from this patient has been transmitted into cord blood lymphocytes, and the virus produced by these cells is similar to the original isolate. From these studies it is concluded that this virus as well as the previous HTLV isolates belong to a general family of T-lymphotropic retroviruses that are horizontally transmitted in humans and may be involved in several pathological syndromes, including AIDS.

Recombinant immunoglobin preparations

Abstract: Altered and native immunoglobulins, including constant-variable region chimeras, are prepared in recombinant cell culture. The immunoglobulins contain variable regions which are immunologically capable of binding predetermined antigens. Methods are provided for refolding directly expressed immunoglobulins into immunologically active form.

Production of a mouse monoclonal antibody reactive with a human nuclear antigen associated with cell proliferation

Abstract: The production of a mouse monoclonal antibody, Ki-67, is described. The Ki-67 antibody recognized a nuclear antigen present in proliferating cells, but absent in resting cells. Immunostainings with Ki-67 revealed nuclear reactivity in cells of germinal centres of cortical follicles, cortical thymocytes, neck cells of gastrointestinal mucosa, undifferentiated spermatogonia and cells of a number of human cell lines. The Ki-67 antibody did not react with cells known to be in a resting stage, such as lymphocytes, monocytes, parietal cells and Paneth's cells of gastrointestinal mucosa, hepatocytes, renal cells, mature sperm cells, brain cells, etc. Expression of the antigen recognized by Ki-67 could be induced in peripheral blood lymphocytes after stimulation with phytohaemagglutinin, whereas it disappeared from HL-60 cells stimulated with phorbol esters to differentiate into mature macrophages in a resting stage. These findings suggest that Ki-67 is directed against a nuclear antigen associated with cell proliferation. A first series of immunostainings of tumour biopsies indicated that Ki-67 may be a potent tool for easy and quick evaluation of the proportion of proliferating cells in a tumour.

Abnormalities of B-cell activation and immunoregulation in patients with the acquired immunodeficiency syndrome.

Abstract: We studied B-lymphocyte function in 12 homosexual male patients with the acquired immunodeficiency syndrome, 5 healthy homosexual men, and 12 heterosexual controls. In comparison with the heterosexual controls, the patients were found to have elevated numbers of cells spontaneously secreting immunoglobulin, decreased B-cell proliferative responses to T-cell-independent B-cell mitogens, and qualitatively deficient helper T cells. The hyperactive spontaneous B-cell responses as well as the refractoriness to signals for T-cell-independent B-cell activation were highly suggestive of an in vivo polyclonal activation of B cells and may have been responsible for the manifestations of B-cell hyperreactivity, such as hypergammaglobulinemia, seen in these patients. We conclude that the scope of immune dysfunction in the acquired immunodeficiency syndrome involves B cells as well as T cells.

Isolation of human T-cell leukemia virus in acquired immune deficiency syndrome (AIDS)

Abstract: Several isolates of a human type-C retrovirus belonging to one group, known as human T-cell leukemia virus (HTLV), have previously been obtained from patients with adult T-cell leukemia or lymphoma. The T-cell tropism of HTLV and its prevalence in the Caribbean basin prompted a search for it in patients with the epidemic T-cell immune deficiency disorder known as AIDS. Peripheral blood lymphocytes from one patient in the United States and two in France were cultured with T-cell growth factor (TCGF) an shown to express HTLV antigens. Virus from the U.S. patient was isolated and characterized and shown to be related to HTLV subgroup I. The virus was also transmitted into normal human T cells from umbilical cord blood of a newborn. Whether or not HTLV-I or other retroviruses of this family with T-cell tropism cause AIDS, it is possible that patients from whom the virus can be isolated can also transmit it to others. If the target cell of AIDS is the mature T cell as suspected, the methods used in these studies may prove useful for the long-term growth of these cells and for the identification of antigens specific for the etiological agent of AIDS.

Monoclonal antibodies: Principles and practice : production and application of monoclonal antibodies in cell biology, biochemistry, and immunology

Abstract: Introduction. The Antibody Response. Cellular Basis of the Immune System. Nature of Antigens. Antibody Structure and Function. Genetics of Antibodies. Introduction to Monoclonal Antibodies. Production of Monoclonal Antibodies. Purification. Fragmentation and Isotopic Labeling of Monoclonal Antibodies. Analysis of Antigens Recognized By Monoclonal Antibodies. Affinity Chromatography Immunofluorescence. Immunohistology. Construction, Screening and Expression of Recombinant Antibodies. Generation of Conventional Antibodies.

Characterization of the Murine Antigenic Determinant, Designated L3T4a, Recognized by Monoclonal Antibody GK 1.5: Expression of L3T4a by Functional T Cell Clones Appears to Correlate Primarily with Class II MHC Antigen‐Reactivity

Abstract: We describe here the properties of mAb GK1.5, which recognizes a cell surface molecule designated L3T4; the determinant on L3T4 recognized by mAb GK1.5 is designated L3T4a. We present evidence here that: i) the expression of L3T4a by murine T cell clones correlates primarily with class II MHC antigen-reactivity; ii) mAb GK1.5 blocks all class II MHC antigen-specific functions (cytolysis, proliferation, release of lymphokines) by murine class II MHC antigen-reactive T cell clones, although there appears to be clonal heterogeneity in the degree to which these functions are blocked by mAb GK1.5; iii) mAb GK1.5 blocks class II MHC antigen-specific release of IL-2 from cloned T cell hybridomas by blocking class II MHC antigen-specific binding; and iv) L3T4 is very similar to the human Leu3/T4 antigen. The properties of mAb GK1.5 (complement fixation, reactivity with all mouse strains tested, profound blocking of all class II MHC antigen-specific functions by murine T cells, usefulness for FACS analyses, and usefulness for immuno-precipitation/SDS-PAGE analyses) make it suitable for investigating both the role of class II MHC antigen-reactive T cells in various immunological phenomena and the mechanistic basis, at the molecular level, of class II MHC antigen-reactivity by murine T cells.

Cytotoxic T-cell immunity to influenza.

Abstract: In a study designed to determine whether cytotoxic T lymphocytes contribute to immunity against influenza virus infection, we inoculated 63 volunteers intranasally with live unattenuated influenza A/Munich/1/79 virus. Over the next seven days clinical observations were made, and the amount of virus shed was measured. The protective effects of preinfection serum antibody and of cytotoxic T-cell immunity against influenza A virus were assessed for each participant. All subjects with demonstrable T-cell responses cleared virus effectively. This response was observed in volunteers in all age groups, including those born after 1956, who did not have specific antibody and hence had probably not been exposed to this subtype of influenza A virus before. Cytotoxic T cells show cross-reactivity in their recognition of the different subtypes of influenza A virus, in contrast to the antibody response that is specific for each virus subtype. We conclude that cytotoxic T cells play a part in recovery from influenza virus infection.

Hybrid hybridomas and their use in immunohistochemistry

Abstract: A normal antibody-producing cell only expresses one antibody, resulting in the well-known phenomenon of allelic exclusion. When two myeloma cells are fused, the derived hybrids are capable of co-dominantly expressing the antibody genes of both parents. Although the respective variable (V) and constant (C) region genes remain expressed in the same cis configuration, heavy and light chains of both parents are scrambled, and hybrid molecules are formed. The same is true when a myeloma and an antibody-producing cell are fused to produce a hybrid myeloma (hybridoma). Fusion therefore allows the production of hybrid immunoglobulin molecules containing two different combining sites. Hybrid molecules of this type retain antigen-binding activity and specificity. Bispecific monoclonal antibodies secreted by hybridomas may have a variety of uses in biology and in medicine. Here we have focused on their application in histochemistry. As an example, we have prepared and tested an anti-somatostatin-anti-peroxidase bispecific antibody. This way of producing hybrid molecules is superior to the production of hybrid antibodies by chemical reconstitution methods because the drastic treatment required for chain separation in the latter is likely to lead to some protein denaturation and loss of antibody activity. Intracellularly synthesized and assembled hybrids do not suffer from this disadvantage. In addition, the recombination of heavy and light chains from different antibody molecules is likely to lead to considerable waste.

Immunoglobulin heavy chain binding protein.

Abstract: Pre-B lymphocytes, and hybridomas derived from them, synthesize immunoglobulin heavy (IgH) chain in the absence of light (L) chain1. In the Abelson virus transformed line 18-81, which is representative of the pre-B cell stage, we observed that at least some of the H-chains are bound to a protein other than L-chain. Here we show that the protein (which we term immunoglobulin heavy-chain binding protein, BiP) binds non-covalently to free IgH, but not to IgH associated with IgL.

Clonotypic structures involved in antigen-specific human T cell function. Relationship to the T3 molecular complex.

Abstract: Monoclonal antibodies were produced against a human cytotoxic T cell clone, CT8III (specificity: HLA-A3), with the view of defining clonally restricted (clonotypic) surface molecules involved in its antigen recognition function. Two individual antibodies, termed anti-Ti1A and anti-Ti1B, reacted exclusively with the CT8III clone when tested on a panel of 80 additional clones from the same donor, resting or activated T cells, B cells, macrophages, thymocytes, or other hematopoietic cells. More importantly, the two antibodies inhibited cell-mediated killing and antigen-specific proliferation of the CT8III clone but did not affect the functions of any other clone tested. This inhibition was not secondary to generalized abrogation of the CT8III clone's function, because interleukin 2 responsiveness was enhanced. To examine the relationship of the structures defined by anti-clonotypic antibodies with known T cell surface molecules, antibody-induced modulation studies and competitive binding assays were performed. The results indicated that the clonotypic structures were associated with, but distinct from, the 20,000-mol wt T3 molecule expressed on all mature T lymphocytes. Moreover, in contrast to anti-T3, anti-Ti1A and anti-Ti1B each immunoprecipitated two molecules of 49,000 and 43,000-mol wt from 131I-labeled CT8III cells under reducing conditions. The development of monoclonal antibodies to such polymorphic T cell surface structures should provide important probes to further define the surface receptor for antigen.

Identification and preparation of epitopes on antigens and allergens on the basis of hydrophilicity

Abstract: An immunoglobulin is provided which consists essentially of a mono-specific, hetero-molecular antibody which is mono-specific to a single antigenic or allergenic determinant. The antibody is specific to the H-epitope of a protein antigen or allergen. The H-epitope is defined by a sequence of at least six amino acids corresponding to the sequence of such amino acids in the protein antigen or allergen where the greatest local average hydrophilicity of the protein antigen or allergen is found.

Latrunculins: novel marine toxins that disrupt microfilament organization in cultured cells

Abstract: Two toxins, latrunculins A and B, which contain a new class of 16- and 14-membered marine macrolides attached to the rare 2-thiazolidinone moiety, were purified recently from the Red Sea sponge Latrunculia magnifica. The effects of these toxins on cultured mouse neuroblastoma and fibroblast cells have been evaluated. In both types of cells, submicromolar toxin concentrations rapidly induce striking changes in cell morphology that are reversible upon removal of the toxin. Immunofluorescence studies with antibodies specific for cytoskeletal proteins reveal that the toxins cause major alterations in the organization of microfilaments without obvious effects on the organization of the microtubular system.

Serologic aspects of IgG4 antibodies. I. Prolonged immunization results in an IgG4-restricted response.

Abstract: Labeled antigen-binding tests were used to determine quantitatively the contribution of IgG4 antibodies to the total IgG antibody response in humans In agreement with literature, we found no IgG4-restricted antibody responses with tetanus toxoid or streptococcal carbohydrate In the serum of individuals immunized for several years with phospholipase (PLA) from honey bee venom, grass pollen allergen, or house dust mite allergen, we often found that more than 50% of the total antigen-binding capacity was due to IgG4 antibodies In the case of beekeepers, it could clearly be shown that during prolonged immunization a shift in the IgG4:IgG1 antibody ratio occurs that finally results in an IgG4-dominated antibody response Evidence is provided that antigen-binding assays may even underestimate the contribution of IgG4 antibodies, because in contrast to IgG1 antibodies, IgG4 antibodies act as monovalent antibodies in being unable to cross-link immunosorbent-bound antigen and radiolabeled antigen

Identification of the gastrointestinal and pancreatic cancer-associated antigen detected by monoclonal antibody 19-9 in the sera of patients as a mucin

Abstract: Monoclonal antibody 19-9, produced by a hybridoma prepared from spleen cells of a mouse immunized with a human colon carcinoma cell line, detects an antigen in the serum from most patients with gastrointestinal and pancreatic cancer (M. Herlyn, H.F. Sears, Z. Steplewski, and H. Koprowski, J. Clin. Immunol., 2: 135-140, 1982). The epitope of this antibody is a carbohydrate with the sugar sequence (formula; see text) in which NeuNAc is N-acetylneuraminic acid, Gal is galactose, GlcNAc is N-acetylglucosamine, and Fuc is fucose. In the colon carcinoma cell line and many gastrointestinal and pancreatic cancers, this sequence occurs in a monosialoganglioside containing a sialylated Lea-active pentasaccharide (sialylated lacto-N-fucopentaose II, IV3-alpha-NeuNAc-III4-alpha-Fuc-LcOse4, in which LcOse4 is Gal beta 1-3GlcNAc beta 1-3Gal beta 1-4Glc) (J. L. Magnani et al. J. Biol. Chem., 257: 14365-14369, 1982). However, the antigen in the sera of patients occurs mainly as a mucin, not a ganglioside, based on the following evidence. Little antigen is extracted by organic solvents from sera, and that which is extracted remains at the origin under conditions of thin-layer chromatography where the ganglioside antigen migrates up the plate. Upon gel filtration of serum on Sephacryl S-400, the antigen is eluted in the void volume, indicating a molecular weight of greater than or equal to 5 X 10(6). Incubation for 5 hr at 37 degrees in 0.1 N NaOH destroys the serum antigen but does not affect the ganglioside antigen. The density of the serum antigen as determined in a CsCl gradient is 1.50 g/ml, while in 4 M guanidine. HCl its density is 1.43 g/ml. Finally, antigen affinity purified by antibody 19-9 from the serum of a cancer patient belonging to the Le(a-b+) blood group contains Leb antigen, consistent with the multiple antigenic specificities exhibited by mucins.

Identification of the C3bi receptor of human monocytes and macrophages by using monoclonal antibodies.

Abstract: We have obtained four monoclonal antibodies, IB4, OKM1, OKM9, and OKM10, all directed against the C3bi receptor of human monocytes and macrophages (M phi). Two criteria were used to determine the specificity of these antibodies. First, culture surfaces coated with the antireceptor antibodies caused specific down modulation of C3bi receptor activity on M phi adherent to these substrates. Second, receptor protein purified by using IB4 or OKM1 retained the ability to bind selectively to C3bi-coated erythrocytes. Each of the antibodies recognizes a distinct epitope on the C3bi receptor; they do not compete with one another for binding to monocytes. Further, when immobilized on a solid support, each of the antibodies binds a molecule from M phi lysates that can simultaneously bind one of the other monoclonal anti-C3bi receptor antibodies. OKM10 binds and masks the ligand-binding site of the C3bi receptor, while IB4, OKM1, and OKM9 bind to sites remote from the C3bi binding site. All four antibodies immunoprecipitated polypeptides of Mr 185,000 and 105,000 from 125I-surface-labeled M phi. IB4 also precipitates polypeptides of Mr 185,000, 153,000, and 105,000. We conclude that the C3bi receptor of human M phi is a complex composed of two polypeptides, Mr 185,000 and 105,000. We have identified monoclonal antibodies reacting with four distinct antigenic determinants of this complex. The determinant recognized by antibody OKM10 is at or near the ligand-binding site of the receptor. The determinant recognized by antibody IB4 is shared by at least two other leukocyte surface proteins.

Monoclonal antibodies to mitotic cells.

Abstract: Certain proteins or activities are present in mitotic cells but not in interphase cells. These proteins may be synthesized or activated, or both, just prior to mitosis and are responsible for the breakdown of the nuclear envelope and the condensation of chromosomes. To learn more about the nature of these proteins, we raised monoclonal antibodies to mitotic cells. Spleen cells from mice immunized with a 0.15 M NaCl extract of synchronized mitotic HeLa cells were fused with SP2/0-Ag14 mouse myeloma cells, and hybrids were selected in medium containing hypoxanthine, methotrexate, thymidine, and glycine. Two different hybridoma clones secreting antibodies reactive with mitotic and meiotic cells from every species tested were isolated. Chromosomes as well as cytoplasm in mitotic cells reacted with the antibodies, as detected by indirect immunofluorescence. The proteins from mitotic cells were separated by electrophoresis in NaDodSO4/polyacrylamide slab gels, transferred to nitrocellulose sheets, and stained immunochemically. The two antibodies, designated MPM-1 and MPM-2, recognize a family of polypeptides with apparent molecular masses of 0.40 to greater than 200 kilodaltons (kDa). Both antibodies reacted strongly with three polypeptide bands of 182 kDa, 118 kDa, and 70 kDa. Only mitotic cells exhibited the protein bands that were recognized by the antibodies. All these bands were found to be phosphoproteins as shown by 32P labeling and autoradiography and their removal by alkaline phosphatase treatment.

Immunoglobulin gene rearrangement and cell surface antigen expression in acute lymphocytic leukemias of T cell and B cell precursor origins.

Abstract: We have explored the relationship among immunoglobulin gene rearrangement, cytoplasmic immunoglobulin production, and cell surface antigen expression within 37 cases of acute lymphocytic leukemia. All 12 cases of the T cell type had germ-line kappa and lambda genes and 11 of 12 had germ-line heavy chain genes. In contrast, all 25 cases of the "non-T, non-B" classification, which lacked both definitive T cell markers and surface immunoglobulin, had rearranged immunoglobulin genes, indicating that they represent precursor cells already committed to the B cell lineage at the gene level. 14 had rearranged heavy chain genes, yet retained germ-line light chain genes, whereas 11 cases had both heavy and light chain gene reorganizations. All patterns of immunoglobulin gene rearrangement predicted by a model that proceeds from heavy chain gene recombination to light chain genes were observed. Despite the uniform presence of rearranged immunoglobulin genes, only five cases produced cytoplasmic mu-chain, one exceptional case produced gamma-chain, and another produced only lambda-chain. The cases of B cell precursor type that do not produce immunoglobulin may represent cells that frequently possess ineffectively rearranged immunoglobulin genes. Included in this group may be a set of cells trapped within the B cell precursor series because their ineffective rearrangements have eliminated certain gene subsegments necessary for the assemblage of an effective heavy chain gene. All seven cases of the non-T, non-B subgroup that bore HLA-DR but lacked CALLA (the common acute lymphocytic leukemia-associated antigen) represented the earliest recognizable stage of B cell precursors with rearranged heavy chain genes but germ-line light chain genes. Correlations here suggest that cells entering B cell development express HLA-DR and rearrange heavy chain genes before the expression of a B cell-associated antigen recognized by the antibody BA-1, the antigen CALLA, and any subsequent light chain gene rearrangements.

The functional significance, distribution, and structure of LFA-1, LFA-2, and LFA-3: cell surface antigens associated with CTL-target interactions.

Abstract: Three cell surface antigens associated with the cytolytic T lymphocyte(CTL)-target cell interaction were identified by generation of monoclonal antibodies (MAb) against OKT4+, HLA-DR-specific CTL and selection for inhibition of cytolysis in a 51Cr-release assay. These MAb block cytolysis by both OKT4+ and OKT8+ CTL and the proliferative responses to PHA and the mixed lymphocyte response (MLR). LFA-1 is an antigen widely distributed on lymphoid tissues and is composed of two polypeptides of 177,000 and 95,000 Mr on all cell types studied. Anti-LFA-1 MAb block NK cell-mediated cytolysis in addition to T lymphocyte-mediated cytotoxicity and proliferation. LFA-2 (Mr = 55,000 to 47,000), a determinant on the sheep red blood cell receptor, is expressed by T cells but not B cells and appears specific for T cell functions. LFA-3 (Mr = 60,000) is a widely distributed antigen present on both hematopoietic and nonhematopoietic tissues and appears to only be involved in T cell functions. MAb to LFA-1 and LFA-2 inhibit function by binding to effector cell surface molecules, whereas anti-LFA-3 MAb appear to block by binding to the target cells. Together with previously described molecules, LFA-1, LFA-2, and LFA-3 demonstrate the complexity of CTL-mediated cytotoxicity at the molecular level.

Generation of human monoclonal antibodies reactive with cellular antigens

Abstract: Human lymphocytes from lymph node, peripheral blood, spleen, and tumor specimens have been fused with the LICR-LON-HMy2 (LICR-2) or SKO-007 human cell lines or the NS-1 mouse myeloma line. Over 75 fusions with the three myeloma-lymphoblastoid lines have been performed. Several factors appeared to improve the fusion outcome, including maintenance of the myeloma-lymphoblastoid lines in logarithmic phase growth at greater than or equal to 95% viability, a delay of 24 hr in the introduction of aminopterin to the fused cells, and preselection of the fetal calf serum used in the medium. For a given number of lymphocytes, fusions with NS-1 produced 5-20 times more clones than fusions with LICR-2 or SKO-007, and LICR-2 produced 4 times as many clones as SKO-007. The percentage of clones secreting human immunoglobulin, the range of immunoglobulin production, and the proportion of IgM, IgA, and IgG secretors were comparable for clones derived from the three myeloma-lymphoblastoid lines. Stable Ig-secreting clones were isolated with approximately equal frequency from LICR-2 and NS-1 fusions. A number of stable clones producing human monoclonal antibodies reacting with cell-surface, cytoplasmic, or nuclear antigens have been isolated from tumor-bearing patients and normal individuals. A surface antigenic system present on normal and malignant cells has been defined with a human monoclonal antibody derived from a patient with breast cancer. Techniques for producing human monoclonal antibody now appear to be sufficiently advanced to initiate a serological dissection of the humoral immune response to cancer.

Enzyme-labeling of antibodies and their fragments for enzyme immunoassay and immunohistochemical staining.

Abstract: The use of an enzyme as a label has a number of advantages over the use of other labels in both immunohistochemistry and immunoassay. Immunofluorescence techniques are not suitable for ultrastructural research on cells, and ferritin-labeled antibodies allow only electronmicroscopic studies. By contrast, enzyme-labeled antibodies permit localization of cellular antigens in relation to tissue structures under light microscope and also demonstration of cellular antigens at an ultrastructural level by electronmicroscopy. Antibody or antibody fragments labeled with enzymes of small molecular weight can more readily permeate cells of tissue sections than ferritin-labeled antibodies. The color of tissue sections prepared by immunoenzymatic techniques is stable for years, while immunofluorescence of tissue sections decreases rapidly when exposed to light. Radioisotope-labeled reagents decay with time; there are health hazards due to radio isotopes; and disposal of radioactive waste is becoming increasing...

Biological effects in vitro of monoclonal antibodies to human epidermal growth factor receptors.

Abstract: Four mouse hybridomas secreting monoclonal immunoglobulin G (IgG) antibodies to epidermal growth factor (EGF) receptors of A431 cells were obtained independently from four fusion experiments. Three of the antibodies, 528 IgG, 225 IgG, and 579 IgG, inhibited the binding of [125I]EGF to A431 cells by at least 95%, and they competed with each other for binding to A431 cells. These antibodies bound to A431 cells, HeLa-S cells and human foreskin fibroblasts with dissociation constants in the range of Kd = 0.6 X 10(-9) to 2.5 X 10(-9) M. The fourth monoclonal antibody, 455 IgG, bound to A431 cells with lower affinity (Kd = 2.0 X 10(-8) M), and it had no effect on the binding of either EGF or the other antibodies to A431 cells. All four antibodies immunoprecipitated EGF receptors from Triton X-100 extracts of A431 membranes, but they were unable to bind to three rodent cell lines. In biological assays, none of the antibodies was able to mimic EGF. The antibodies which inhibited the binding of EGF blocked EGF-enhanced phosphorylation of A431 membrane proteins and inhibited EGF induced human fibroblast proliferation. These three antagonistic antibodies also partially reversed the inhibition of A431 growth by EGF. In contrast, 455 IgG had no effect on the early or delayed cellular responses to EGF.

Characterization of an antigenic determinant of the glycoprotein that correlates with pathogenicity of rabies virus

Abstract: The pathogenicity of fixed rabies virus strains for adult mice depends on the presence of an antigenic determinant on the viral glycoprotein. Two virus-neutralizing monoclonal antibodies have been used to identify this determinant. All pathogenic strains of fixed rabies virus bind to these antibodies and are neutralized by them, whereas nonpathogenic strains fail to react with these monoclonal antibodies and are not neutralized by them. Antigenic variants of the rabies virus with altered glycoprotein were selected by growing virus in the presence of one monoclonal antibody, 194-2. All variants that lost their ability to react with this antibody and an additional antibody, 248-8, were found to be nonpathogenic for adult mice. Analysis of tryptic peptides of the glycoproteins of pathogenic parent virus and nonpathogenic variants and the amino acid sequence of a specific variant tryptic peptide revealed that the change in pathogenicity corresponded to an amino acid substitution at position 333 of the glycoprotein molecule. The nucleotide sequence of the nonpathogenic variant glycoprotein gene contained a base change that confirmed the single amino acid substitution in the tryptic peptide replacing arginine-333 in the parental glycoprotein. We conclude that arginine-333 is essential for the integrity of an antigenic determinant and for the ability of rabies viruses to produce lethal infection in adult mice.

Radioactive labeling of antibody: a simple and efficient method

Abstract: A simple and efficient method of covalently coupling the strong chelator diethylenetriaminepentaacetic acid to proteins was developed for radiolabeling immunoglobulin G antibodies. After being coupled and labeled with indium-111, a monoclonal antibody to carcinoembryonic antigen retained its ability to bind to its antigen in vitro and in vivo. In nude mice with a human colorectal xenograft, 41 percent of the injected radioactivity became localized in each gram of xenograft at 24 hours compared with 9 percent for control antibody and 19 percent for radioiodinated antibody to carcinoembryonic antigen.

Immunity to Influenza in Man

Abstract: The observations summarized in this review indicate immunity to infection with type A influenza viruses is subtype specific since little or none is conveyed to subtypes possessing immunologically distinct HA and NA proteins. However, within a subtype, a prior antigenic experience with one variant may prevent or modify illness to another. The resulting degree of subtype immunity depends on the extent of relatedness between variants. Observations with H3N2 viruses indicate that homotypic resistance to subsequent infection and illness with the same virus is potent and of relatively long duration. The long lasting durability of such immunity was indicated by the epidemiologic pattern following the reappearance of H1N1 virus. Knowledge of the duration and specificity of immunity aids considerably in assessing mechanisms that account for host resistance to influenza. Recovery from influenza virus infection must involve a variety of humoral and cell-mediated immune mechanisms, and conclusions regarding the relative importance of each one are not possible at present. To prevent infection, involved immune mechanism(s) must account for: (a) subtype specificity, (b) reduced cross-reactivity of immunity for succeeding antigenic variants, (c) a long duration of immunity, and (d) immunity at the mucosal surface. Only antibody directed toward the HA molecule presently satisfies these properties and thus should be considered the major mediator of resistance to infection. Study of naturally occurring infection is needed for determining the duration and specificity of secretory IgA in nasal and lower respiratory secretions so as to establish its relative importance as a mediator of immunity. However, the described duration, specificity, and consistent relationship to immunity suggest that IgG antibody in respiratory secretions, derived entirely or partly from serum, is the most likely mediator of resistance to natural influenza.

Biological significance of carbohydrate chains on monoclonal antibodies.

Abstract: We have prepared monoclonal hapten-specific mouse IgG2b antibodies depleted of asparagine-linked carbohydrate chains by treating the hybridoma cells with tunicamycin. The carbohydrate-deficient antibodies behaved in an identical manner to the normal antibodies with regard to fine antigen-binding reactivity (a Fab fragment feature) and protein A binding capacity [a feature requiring integrity at the CH2 and CH3 domain-interaction regions in the constant region of the heavy chain (CH)]. However, they lost the ability to activate complement, to bind to Fc receptors on macrophages, and to induce antibody-dependent cellular cytotoxicity. Furthermore, antigen-antibody complexes produced from such carbohydrate-deficient antibodies failed to be eliminated rapidly from the circulation. We conclude that removal of carbohydrate chains from IgG molecules may have a profound and highly select impact on the biological activity to these antibodies.

Recognition of myelin-associated glycoprotein by the monoclonal antibody HNK-1.

Abstract: Myelin-associated glycoprotein (MAG) is a quantitatively minor component in both peripheral and central myelin sheaths that is thought to have a role in cell-cell interactions within the nervous system. We show here that a mouse monoclonal antibody, HNK-1, which is directed against human natural killer cells also recognizes an antigenic determinant of human central and peripheral nervous system white matter by immunoperoxidase staining of tissue sections. Immunoblot analysis of myelin proteins and purified extracted MAG indicates that the antigen recognized is MAG.