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Zhong Nan Da Xue Xue Bao. Yi Xue Ban =... Mar 2011To obtain I50 anti-idiotype antibody and identify its activity in vitro.
OBJECTIVE
To obtain I50 anti-idiotype antibody and identify its activity in vitro.
METHODS
I50 anti-idiotype (Id) antibody gene was amplified from the template of fuse 5-I50 by PCR to construct a prokaryotic expression vector pET25b-I50. The expression of pET25b-I50 in E. coli BL21(DE3) was induced by isopropylthio-β-D-galactopyranoside (IPTG) and was confirmed by SDS-PAGE and Western blot with Ab1(FC2) monoclonal antibody and an anti-hexahistidine tag antibody. The method of dialysis refolding was used to restore the activity of I50 anti-Id antibody, which was measured by Dot-ELISA and lymphocyte proliferation assay.
RESULTS
The recombinant vector was successfully constructed and the recombinant protein was successfully expressed and purified with 90% purity. The relative molecular weight of the expressed protein was 15 kD, which was in accordance with expectation. The activity of I50 anti-Id antibody could be restored and could promote the proliferation of lymphocyte in a dose-dependent manner.
CONCLUSION
These results suggested that I50 anti-Id protein vaccine is likely an option in the therapy against nasopharyngeal carcinoma in vivo.
Topics: Antibodies, Anti-Idiotypic; Antigens, Neoplasm; Cancer Vaccines; Cell Line, Tumor; Escherichia coli; Genetic Vectors; Humans; Nasopharyngeal Neoplasms; Recombinant Proteins
PubMed: 21464538
DOI: 10.3969/j.issn.1672-7347.2011.03.001 -
Immunology Jan 1983A method is described for using mini-Marbrook chambers for culturing spleen cells together with anti-idiotype antibody (anti-Id) to induce the appearance of suppressor T...
A method is described for using mini-Marbrook chambers for culturing spleen cells together with anti-idiotype antibody (anti-Id) to induce the appearance of suppressor T cells (Ts). Spleen cells that have been cultured with affinity prepared anti-Id (IgG) but not those cultured with normal IgG, suppress a secondary IgE response to timothy grass pollen antigen B (AgB) when injected intravenously into AGB-primed and boosted syngeneic recipient mice. Suppressor T cells are not induced if the spleen cells cultured with anti-Id are depleted of B cells of if the cells are cultured with the F(ab)2 fragment of anti-Id: both of these results are compatible with Fc+ cells playing a role in the induction of Ts cells by anti-Id. Analysis of soluble suppressor factors in an ELISA test suggests that both TS1 and TS2 cells may be induced by anti-Id.
Topics: Animals; Antibodies, Anti-Idiotypic; Cells, Cultured; Enzyme-Linked Immunosorbent Assay; Immunoglobulin E; Immunoglobulin G; Immunoglobulin Idiotypes; Lymphocyte Activation; Mice; Mice, Inbred Strains; Pollen; Rats; Rats, Inbred Strains; Spleen; T-Lymphocytes, Regulatory
PubMed: 6217153
DOI: No ID Found -
Journal of Autoimmunity Apr 1993The presence of anti-idiotype antibody (anti-id) activity to anti-myeloperoxidase autoantibodies (anti-MPO) was measured by assessing binding of IgG in patients' sera to... (Comparative Study)
Comparative Study
The presence of anti-idiotype antibody (anti-id) activity to anti-myeloperoxidase autoantibodies (anti-MPO) was measured by assessing binding of IgG in patients' sera to F(ab')2 fragments of polyclonal heterologous anti-MPO antiserum. Thirteen paired acute and remission sera and three groups of sequential sera from patients with systemic vasculitis were studied. The levels of anti-MPO fell as disease activity subsided with treatment. However, levels of anti-id activity against these anti-MPO rose as patients entered remission. This effect was still seen after controlling for binding to F(ab')2 constant regions and after polyethylene glycol precipitation of immune complexes. In the sequential studies, levels of anti-MPO and anti-MPO anti-id tended towards a reciprocal relationship. These observations point to dynamic interactions between autoantibody and complementary anti-idiotype antibodies in this system and a role for idiotypic networks in the regulation of anti-neutrophil cytoplasm antibodies.
Topics: Acute Disease; Antibodies, Anti-Idiotypic; Antibodies, Antineutrophil Cytoplasmic; Autoantibodies; Autoimmune Diseases; Convalescence; Cytoplasm; Enzyme-Linked Immunosorbent Assay; Humans; Immunoglobulin Fab Fragments; Immunoglobulin G; Neutrophils; Peroxidase; Vasculitis
PubMed: 8388692
DOI: 10.1006/jaut.1993.1019 -
Proceedings of the National Academy of... Nov 1980Hapten-augmentable plaque-forming cells (PFC) are cells whose secretion of antibody is specifically inhibited by surface-bound auto-anti-iodotype antibody that can be...
Production of auto-anti-idiotypic antibody during the normal immune response: changes in the auto-anti-idiotypic antibody response and the idiotype repertoire associated with aging.
Hapten-augmentable plaque-forming cells (PFC) are cells whose secretion of antibody is specifically inhibited by surface-bound auto-anti-iodotype antibody that can be displaced by hapten. This study showed that the percentage of hapten-augmentable PFC present in mice during the primary response to trinitrophenylated Ficoll (TNP-F) increases with age. The data suggest that there is a relative increase in the auto-anti-idiotypic antibody response with age and therefore a greater down-regulation of antibody production. The effect of age on idiotype expression was also studied. Hapten-reversible inhibition of plaque formation was used as an assay for anti-inhibition of plaque formation was used as an assay for anti-idiotype antibody and idiotype-bearing antibody-secreting cells. Sera from aged (21- to 22-month-old) C57BL/6 mice immunized with TNP-F significantly inhibited plaque formation, in a hapten-reversible manner, by spleen cells from 81% of TNP-F-immunized aged mice. However, these sera inhibited plaque formation by cells from only 50% of similarly immunized young adult (6- to 8-week-old) mice and 20% of immature (3- to 4-week-old) syngeneic mice. Similarly, sera from TNP-F-immunized young adult or immature mice inhibited formation of plaques by cells from immunized donors of the same age as the mice from whom the serum was obtained, but only rarely inhibited plaque formation by cells from mice of other age groups. The data thus suggest that the repertoire of TNP-specific idiotypes that are produced in response to TNP-F varies with age in syngeneic mice.
Topics: Aging; Animals; Antibodies, Anti-Idiotypic; Antibody Formation; Autoantibodies; B-Lymphocytes; Immunoglobulin Idiotypes; Mice; Receptors, Antigen, B-Cell
PubMed: 6969889
DOI: 10.1073/pnas.77.11.6788 -
Proceedings of the National Academy of... Nov 1997The anti-idiotype approach is based on the assumption that an antibody specific for a receptor-binding domain of a ligand could be structurally related to the receptor....
The anti-idiotype approach is based on the assumption that an antibody specific for a receptor-binding domain of a ligand could be structurally related to the receptor. Therefore, a structural mimic of a receptor-binding domain, selected with an anti-ligand antibody, might be a functional substrate for the receptor. This hypothesis was addressed here by generating antibodies recognizing the Rev-nuclear export signal (NES). A functional NES is required for active export, presumably by interacting directly or indirectly with the nuclear pore complex. Anti-NES antibodies were used to isolate RNA mimics of the NES peptide from combinatorial RNA libraries. The RNA-mimics are exported actively, block Rev-dependent export of a reporter RNA, and inhibit cap-dependent U1 snRNA export in Xenopus oocytes, properties previously reported for NES-peptide conjugates.
Topics: Animals; Antibodies, Anti-Idiotypic; Binding Sites, Antibody; Biological Transport; Cell Nucleus; Gene Products, rev; Nucleic Acid Conformation; Oocytes; RNA; RNA Caps; Temperature; Transcription, Genetic; Xenopus
PubMed: 9371762
DOI: 10.1073/pnas.94.24.12839 -
Journal of Immunology (Baltimore, Md. :... Jul 1986Five murine monoclonal antibody (mAb) anti-idiotypes (id), shown in the accompanying report by binding studies to be reactive with five different id on a single member...
Five murine monoclonal antibody (mAb) anti-idiotypes (id), shown in the accompanying report by binding studies to be reactive with five different id on a single member of the 5AF6 family of BALB/c antibodies against the p-azophenylarsonate (Ar) hapten, were used to examine the distribution of their recognized id among anti-Ar in BALB/c and other mouse strains immunized with keyhole limpet hemocyanin-Ar (KLH-Ar). Differences in id expression in BALB/c and other strains substantiate that all five monoclonal anti-id reacted with different id. This suggests that the anti-id repertoire for a single antibody molecule may be extensive. Two of the anti-id reacted with id that were found in virtually all KLH-Ar immunized BALB/c mice, but constituted only a subset (approximately 33%) of the antibodies representing the 5AF6 family. The other three anti-id reacted with id infrequently expressed among BALB/c anti-Ar. Other mouse strains producing 5AF6 family anti-Ar antibodies also produced antibodies recognized by mAb 2CB8 and 6BA1; however, the three id infrequently expressed in BALB/c mice were produced in higher quantities and in a greater percent of mice. Monoclonal anti-id were capable of suppressing a portion but not all of the 5AF6 family of anti-Ar antibodies. Four of the five anti-id suppressed a greater fraction of the 5AF6 family than that id represented in a normal immune response, suggesting that suppression was mediated via an id other than that recognized by these monoclonal anti-id. Overall, the results indicate that an extensive repertoire of anti-id can be produced against a single id antibody, but suppression induced by treatment with these anti-id in this model is presumably mediated via another as yet unidentified id determinant(s).
Topics: Animals; Antibodies, Anti-Idiotypic; Antibodies, Monoclonal; Azo Compounds; Binding Sites, Antibody; Binding, Competitive; Immunoglobulin Idiotypes; Immunosorbent Techniques; Mice; Mice, Inbred A; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Inbred DBA; Species Specificity; p-Azobenzenearsonate
PubMed: 3522733
DOI: No ID Found -
Research in Immunology May 1991
Review
Topics: Animals; Antibodies, Anti-Idiotypic; Antibody-Producing Cells; B-Lymphocytes; Haptens; Immunoenzyme Techniques; Spleen
PubMed: 1925000
DOI: 10.1016/0923-2494(91)90085-w -
Infection and Immunity Feb 1990We produced anti-idiotype antibodies to antibody to lipid A from Eikenella corrodens. The ALA-1 monoclonal antibody (immunoglobulin M [IgM] isotype), which had already...
Protection of mice against the lethal toxicity of a lipopolysaccharide (LPS) by immunization with anti-idiotype antibody to a monoclonal antibody to lipid A from Eikenella corrodens LPS.
We produced anti-idiotype antibodies to antibody to lipid A from Eikenella corrodens. The ALA-1 monoclonal antibody (immunoglobulin M [IgM] isotype), which had already been produced in our laboratory (T. Kato, I. Takazoe, and K. Okuda, Infect. Immun. 57:656-659, 1989), had reacted strongly with lipid A from E. corrodens, Escherichia coli, and Salmonella minnesota. Four anti-idiotype monoclonal antibodies to ALA-1 (Ab1), designated A2LA-1 (IgG1 isotype), A2LA-2 (IgG2a isotype), A2LA-3 (IgG2a isotype), and A2LA-4 (IgG3 isotype), which recognized the idiotype Ab1, were produced. A2LA-1, A2LA-2, and A2LA-3 were capable of over 61% inhibition of ALA-1 reactivity to E. coli J5 lipid A in an enzyme-linked immunosorbent assay system. The sera of mice and rabbits immunized with the anti-idiotype antibodies revealed that the internal image anti-idiotype antibody induced the production of IgG antibodies that cross-reacted with or bound to lipid A. These studies indicate that A2LA-1 and A2LA-2 contained an antigenic epitope that mimicked lipid A. Immunization of mice with A2LA-1 resulted in prevention of lethal toxicity from E. coli J5 lipopolysaccharide.
Topics: Animals; Antibodies, Anti-Idiotypic; Antibodies, Monoclonal; Bacteroides; Eikenella corrodens; Enzyme-Linked Immunosorbent Assay; Escherichia coli; Female; Immunization; Lipid A; Lipopolysaccharides; Mice; Mice, Inbred BALB C; Rabbits
PubMed: 2404870
DOI: 10.1128/iai.58.2.416-420.1990 -
Cancer Immunology, Immunotherapy : CII Sep 2010RP215 monoclonal antibody (Mab) was shown to recognize carbohydrate-associated epitope(s) in the heavy chains of cancer cell-expressed immunoglobulins, designated in...
RP215 monoclonal antibody (Mab) was shown to recognize carbohydrate-associated epitope(s) in the heavy chains of cancer cell-expressed immunoglobulins, designated in general as CA215 pan cancer marker. Growth inhibitions of tumor cells in vitro by RP215 Mab and antibodies against its anti-idiotype (anti-id) antibodies were investigated. Polyclonal rabbit anti-id antibodies and the corresponding rat anti-id Mabs were generated and characterized. Following immunizations in mice, antisera raised against anti-id antibodies were analyzed by typical immunoassays. It was observed that mouse anti-anti-id sera (Ab3) revealed binding affinity and specificity to CA215 that are comparable to those of RP215. Both RP215 and Ab3 were shown to induce apoptosis of cultured cancer cells in vitro by TUNEL and MTT assays. These experimental observations were consistent with that of in vivo tumor growth inhibition by RP215 in previous nude mouse experiments. Therefore, heterologous or homologous anti-id antibodies of RP215 that contain the internal image of its specific epitope in CA215 may serve as effective anti-cancer vaccines for therapeutic treatments of various cancers in humans. The relative stability of RP215-specific carbohydrate-associated epitope was compared to that of human IgG at extreme pH's (
or=12) or following NaIO(4) treatments. The major molecular forms of CA215 were further documented with various enzyme immunoassays and found to have similar secondary structures to those of normal human immunoglobulin G. Topics: Animals; Antibodies, Anti-Idiotypic; Antibodies, Monoclonal; Antibody Affinity; Antigens, Neoplasm; Apoptosis; Cancer Vaccines; Cell Growth Processes; Cell Line, Tumor; Epitopes; Humans; Immunization; Mice; Mice, Inbred BALB C; Neoplasms; Protein Conformation; Protein Stability; Rabbits; Rats
PubMed: 20473495
DOI: 10.1007/s00262-010-0864-7 -
MAbs 2010Recombinant antibodies may be engineered to obtain improved functional properties. Functional mapping of the residues in the binding surfaces is of importance for...
Recombinant antibodies may be engineered to obtain improved functional properties. Functional mapping of the residues in the binding surfaces is of importance for predicting alterations needed to yield the desired properties. In this investigation, 17 single mutation mutant single-chain variable fragments (scFvs) of the anti-idiotypic antibody anti-TS1 were generated in order to functionally map amino acid residues important for the interaction with its idiotype TS1. Residues in anti-TS1 determined to be very important for the interaction were identified, Y32L, K50L, K33H, and Y52H, and they were distributed adjacent to a centrally located hydrophobic area, and contributed extensively to the interaction energy (≥2.5 kcal/mol) in the interaction. Quantitative ELISA assays, BIAcore technologies and three-dimensional surface analysis by modeling were employed to visualize the consequences of the mutations. The expression levels varied between 2 - 1,800 nM as determined by ELISA. All the 17 scFvs displayed higher dissociation rates (60 - 1,300 times) and all but two of them also faster association rates (1.3 - 56 times). The decrease in affinity was determined to be 1.6 - 12,200 times. Two of the mutants displayed almost identical affinity with the wild type anti-TS1, but with a change in both association and dissociation rates. The present investigation demonstrates that it is possible to generate a large panorama of anti-idiotypic antibodies, and single out a few that might be of potential use for future clearing and pre-targeting purposes of idiotypic-anti-idiotypic interactions.
Topics: Amino Acid Sequence; Antibodies, Anti-Idiotypic; Antibody Affinity; Electrophoresis, Polyacrylamide Gel; Enzyme-Linked Immunosorbent Assay; Epitope Mapping; Humans; Hybridomas; Immunoglobulin Variable Region; Keratin-8; Kinetics; Molecular Sequence Data; Mutagenesis, Site-Directed; Protein Structure, Quaternary; Single-Chain Antibodies
PubMed: 21124071
DOI: 10.4161/mabs.2.6.13275