Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0026764 (multiple myeloma)
 36,148 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A hybridoma cell line that secretes monoclonal antibody, MAb-ER-Br-1-15-4-18 is established. The MAb is highly specific for estrogen receptor (ER) from human breast tumor cells. In order to raise the antibody, the ER was first isolated from human breast tumor. Mice were immunized with the partially purified ER and the fusion of the spleen cells from the mouse, showing the highest serum titer, with the cells of the NS-1 mouse myeloma line, produced hybrid cells which continuously secreted antibodies specific for ER. Three of the hybridoma cultures which tested strongly positive were cloned using limiting dilution method and one of the cell lines was selected for further study. The recovery of the MAb from the cell culture was done by ammonium sulfate precipitation followed by dialysis and then hydroxylapatite liquid chromatography using linear gradients. The purity of the antibody was checked by polyacrylamide gel electrophoresis. The MAb was isotyped and found to be IgG1. When checked against other antigens the MAb showed a minimal cross-reactivity to ER from rabbit uterus and none to ovalbumin or rat liver ferritin. Further experiments showed that the MAb recognized the ER bound to the hormone and ER in the nucleus of breast tumor cells.
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PMID:Production and characterization of a monoclonal antibody to partially purified estrogen receptor from human breast tumor. 292 8

Normal human sera contain one or several factors cytotoxic for normal guinea pig thymocytes, and when serum is precipitated with ammonium sulphate (60% saturated) and the precipitate dissolved and dialyzed, the activity is preserved. Gel chromatography with Sephadex G-150 and Sepharose CL-6B indicated a molecular weight of approximately 900,000 daltons. The active fractions contained a high amount of IgM according to single radial immunodiffusion and two-dimensional gel electrophoresis. Quantitation of the IgM band in one-dimensional gel electrophoresis preparations by gel scanning indicated that IgM accounted for 65% of the eluted proteins in active fractions. Purified human IgM from myeloma patients eluted as the active factor during gel chromatography. Elimination of IgM from serum by affinity chromatography eliminated the cytotoxic activity. The serum could also be inactivated by heating. The mixing of IgM-depleted serum with either polyclonal IgM or heat-inactivated serum restored the activity. Thus, the cytotoxic activity is due to IgM antibodies plus a heat-labile component (presumably complement). The presence of the cytotoxic activity in autologous (guinea pig) serum was recently demonstrated. The possible functional role of these antibodies in the elimination process of a large number of cortical thymocytes is suggested.
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PMID:Purification, identification and elimination of a natural human serum antibody with cytotoxic effect on guinea pig thymocytes. 310 42

A human immunoglobulin M (IgM) antibody secreting hybridoma, HMG1, has been established and studied for its reactivity against human gastric cancer cells. Lymphocytes isolated from a regional lymph node of patient with gastric adenocarcinoma were fused with mouse myeloma cells NS-1. Supernatants from the generated human-mouse hybrids were first screened for immunoglobulin production by ELISA. The identified human IgM-secreting hybridomas were expanded and subcloned for further analysis or cryopreservation. The screening for binding of antibodies to a panel of human cancer cell lines and normal fibroblast was carried out with PAP or indirect immunofluorescence stain. The selected hybridoma, HMG1 after being cloned three times, was stable in secreting IgM (about 4 micrograms/1 x 10(6) cells) for more than 9 months. Large amount of ascites was obtained by injecting this hybrid to BALB/C nude mice pretreated with anti-lymphocyte serum and pristane. The ascitic fluid contained 5-19 mg human Ig/ml. Subsequently this IgM was extracted from ascitic fluid by saturated ammonium sulphate solution. This crude extract was further purified with immuno-affinity chromatography. Both this purified ascite-IgM as well as IgM from HMG1 supernatant would react with gastric cancer cell line BGC-823 but not with human normal fibroblasts 350Q by PAP or immunofluorescence analysis. The human HMG1-IgM reacted with gastric cancer cells on paraffin embedded tissue section but did not react with normal gastric mucosa cells. HMG1-IgM had some complement dependent cytotoxicity against BGC-823. These results suggest that the establishment of anticancer human monoclonal antibodies with human-mouse hybridoma technique be feasible. There is a possibility for clinical applications of this human monoclonal antibody in the future.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:[Study of anticancer human monoclonal antibody--establishment of human monoclonal antibody to gastric cancer by human-mouse hybridoma]. 324 78

A 65-kDa estrogen receptor (ER) protein has been demonstrated both by sucrose gradient analysis and by immunoblot, using anti-ER monoclonal antibodies (MAbs). Since the ER is denatured in many experimental situations, such as formaldehyde fixing of samples for histochemistry and electroimmunoblotting studies, in this work we used a denatured 60-70-kDa ER-rich protein preparation as antigen for mice immunization in order to raise anti-ER MAbs. That material was obtained by affinity purification on an allyl-estradiol matrix of the MCF-7 cytosolic ER, followed by further isolation and enrichment by PAGE. NS-1 myeloma cells and spleen lymphocytes from the immunized mice were fused, and resultant hybridoma colonies were screened by [125I]-estradiol-labelled nuclear ER immunoprecipitation. The isolated MAb, E476, shows a moderate ability to precipitate ER and reacts strongly with a 46-kDa antigen in Western blot assay. The 46-kDa antigen was not detectable in native cytosol but became reactive after 50% ammonium sulfate precipitation of cytosolic proteins. The 46-kDa antigen appeared concentrated in the NaSCN plus estradiol eluate of the affinity column used for cytosolic ER purification. Freshly prepared 60-70-kDa material from the preparative gel electrophoresis did not show any E476 reactivity. However, when the 60-70-kDa proteins were frozen, thawed and speed vacuum concentrated, the 46-kDa antigen became detectable. Storage increased the reactivity of the 60-70-kDa material with the E476 MAb. The 46-kDa antigen was present only in the ER positive cell lines, and was absent in all negative cell lines tested. The 46-kDa protein is also present in the ER positive human breast cancer specimens. We conclude that the 46-kDa protein identified with the E476 MAb in human breast cancer is probably a naturally occurring ER fragment.
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PMID:A 46-kDa antigen associated with estrogen receptor in human breast cancer. 338 60

Human IgE was isolated for the first time from the supernatant of the culture fluid of a human myeloma cell line, U266. The purification procedure consisted of salting out from the supernatant with ammonium sulfate, affinity chromatography on a lysine-Sepharose 4B column, ion exchange chromatography on a DEAE-Sephacel column, gel filtration and recycling chromatography on a Sephacryl S-300 column and removal of bovine proteins on an anti-bovine serum rabbit IgG-Sepharose 4B column. One hundred and twenty eight milligrams of IgE was recovered from 461 of culture fluid. The purification was extremely simplified by the introduction of immunoaffinity chromatography using the monoclonal antibody prepared by immunizing a mouse with an IgE preparation obtained by the above method: about 3.3 mg was recovered from 960 ml of culture fluid. The purified preparation was homogeneous as judged by the double-immunodiffusion test and end group analysis. The amino acid and carbohydrate compositions of the preparation coincided with those reported on other preparations obtained from the sera of myeloma patients. Our preparation, however, showed two bands with apparent mol. wts of 240,000 and 230,000 on sodium dodecyl sulfate polyacrylamide gel electrophoresis. When it was reduced with dithiothreitol and analyzed by electrophoresis, it gave two heavy chains and one light chain with apparent mol. wts of 80,000 and 76,000, and 28,000, respectively. On the other hand, the IgE molecule that was synthesized and secreted into the medium in the presence of tunicamycin (0.5 microgram/ml) gave only one heavy chain and one light chain with apparent mol. wts of 62,000 and 28,000, respectively. These results demonstrated that the two IgE molecular species contained in our preparation differed from each other in the carbohydrate moiety in their heavy chains.
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PMID:Purification and characterization of IgE produced by human myeloma cell line, U266. 370 74

In untreated serum of three patients with multiple myeloma, concentrations of inorganic phosphate ranged from 130 to 270 mg/L as measured with a chromogenic assay based on the interaction of phosphate ion with ammonium molybdate in the presence of ferrous sulfate. There were no clinical features of hyperphosphatemia, and values for total calcium concentration in serum remained within normal limits throughout. Subsequent investigations demonstrated that this hyperphosphatemia was spurious and was caused by high concentrations of the paraprotein interfering with the chromogenic assay. Because this type of assay, adapted for automated systems, is now widely used in clinical laboratories, we call attention to this limitation to avoid confusion in clinical evaluations of patients with multiple myeloma.
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PMID:Paraprotein interference with colorimetry of phosphate in serum of some patients with multiple myeloma. 373 47

Ovine secretory IgA (sIgA) has been purified to relative homogeneity by ammonium sulphate precipitation (to 40 per cent w/v) of lung lavage fluid from 3- or 12-month-old lambs, followed by molecular sieve chromatography on a Sephacryl S300 matrix. Three peaks A, B and C with molecular sizes corresponding to 550,000, 400,000 and 165,000 respectively were eluted from the column. Immunoelectrophoresis, radial immunodiffusion and enzyme-linked immunosorbent assays (ELISAs) with class specific antiserum confirmed that peak B contained only IgA. Polyacrylamide gel electrophoresis of peak B under reducing conditions resolved three subunits corresponding to secretory component, heavy and light chains. Hybridomas generated by fusing spleen cells from IgA-primed Balb/C mice with murine myeloma (Sp2/0) were screened for IgA-specific monoclonal antibodies against a panel of ovine IgG2, IgG1, IgA and IgM. One particular clone, F3-4B4, identified as murine IgG1, was monospecific against ovine IgA with no cross reactivity against bovine immunoglobulins. This hybridoma was successfully tested as a serological probe by ELISA profiling to locate IgA containing fractions in the course of immunoglobulin purification from biological fluids.
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PMID:Monoclonal antibodies against ovine IgA purified from lung lavage fluid. 376 94

Four monoclonal antibodies were obtained to rat brain choline acetyltransferase (CAT). The enzyme was purified 95,000-fold from rat brain by precipitation with acetic acid at pH 4.5, fractionation with 40 to 60% (NH4)2SO4, CM-Sephadex chromatography, and affinity column chromatography on agarose-hexane-coenzyme A. The enzyme preparation was applied to the affinity column in the presence of 10 mM acetylcholine to increase the affinity of CAT for coenzyme A; the enzyme then was eluted with 10 mM acetyl coenzyme A. Fusion of P3X63 Ag8 myeloma cells with spleen cells isolated from a BALB/c mouse that had been immunized with affinity-purified CAT with a specific activity of 29.4 mumol of ACh synthesized/min/mg of protein resulted in the isolation of four hybridomas synthesizing antibodies to CAT that inhibit the activity of the enzyme. Anti-CAT 1 or 2 inhibits CAT activity 100%. At the highest antibody concentration tested, anti-CAT 3 inhibited acetylcholine synthesis 80%. Hybridoma antibody-dependent inhibition of CAT activity was reversed by dissociation of immune complexes via dilution, demonstrating that antibody binding does not irreversibly alter the structure of the enzyme. When bound to [rabbit anti-mouse IgG . protein A Staphylococcus aureus] complexes, anti-CAT 1, 3, and 4 each were effective reagents for the precipitation of CAT activity from solution. Thirty-one to 53% of the precipitated enzyme was recovered following the dissociation of immune complexes. Anti-CAT 1, 2, and 3 inhibit CAT from 18-day chick embryo brain, NS20-Y mouse neuroblastoma cells, and rat brain.
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PMID:Inhibition of choline acetyltransferase by monoclonal antibodies. 388 Aug 11

Cultured mouse peritoneal cells from unstimulated mice developed plaque-forming activity against isologous bromelain-treated erythrocytes. Several IgM monoclonal autoantibodies obtained by fusion of peritoneal cells from NZB or CBA origin with BALB/c myeloma cells were purified by affinity chromatography on trimethyl ammonium (TMA) column on the basis of their cross-reactivity with TMA, phosphorylcholine (PC) or choline haptens. Binding affinity for PC hapten was of the order of 10(3) M-1. Idiotypic studies with a polyclonal rabbit anti-idiotypic reagent revealed strong cross-reactions with all hybridoma autoantibodies thus far tested. In addition, the rabbit anti-idiotypic serum detected idiotypes or cross-reactive idiotypes in the sera of NZB and CBA as well as BALB/c mice. N-terminal amino acid sequence analyses of three hybridoma autoantibodies from NZB mice and one from CBA mice were carried out. The sequences of the first 32 residues of the four heavy chains showed that three were identical while one had one amino acid interchange; they belong to the VHIII-subgroup. The light chains were identical in the first 35 residues with the exception of a substitution at position 3 in two light chains and are members of the VK-9-subgroup. These results entirely support the idiotypic data. These monoclonal autoantibodies from NZB and CBA mice although isolated and eluted from PC-related haptens do not have any apparent structural nor idiotypic relationship to PC-specific antibodies. Idiotypic and V-region N-terminal sequence data suggest that these autoantibodies constitute a highly restricted family of molecules likely to be encoded by unique germ-line genes which may be expressed as such or as somatic variants in different mouse strains.
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PMID:Monoclonal autoantibodies against mouse red blood cells: a family of structurally restricted molecules. 389 36

Nonactivated (8.5S) rabbit uterine progestin receptor was enriched 10- to 30-fold by chromatography on columns of spheroidal hydroxylapatite and DEAE-cellulose. A total of approximately 25 micrograms of receptor (purity approximately 1%) was injected at multiple sites into a BALB/c mouse. After several injections, splenic lymphocytes were fused with P3x63Ag8.653 mouse myeloma cells. This fusion produced in excess of 240 hybridomas, which were screened by an enzyme-linked immunosorbent assay (ELISA), solid-phase radioimmunoassay, and sucrose gradient centrifugation. One colony (KN 382/EC1) produced a mouse immunoglobulin G1 which bound rabbit 8.5S uterine progestin receptor. The cell line has been repeatedly cloned under conditions of limiting dilution and expanded in mice as ascitic tumors. Antibody was purified by (NH4)2SO4 precipitation, DEAE-cellulose chromatography, and affinity chromatography with protein A - Sepharose CL-4B. Specificity of the antibody was determined by sucrose gradient centrifugation and solid-phase radioimmunoassay. The antibody bound to progestin receptors from rabbit uterus and MCF-7 breast cancer cells. It did not react with progestin receptors from rat uterus, guinea pig uterus, or chick oviduct, nor did it bind to estrogen receptors from any of the tissues we tested.
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PMID:Development of a monoclonal antibody to the rabbit 8.5S uterine progestin receptor. 398 62


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