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)

We used the hybridoma technique to characterize further the platelet glycoprotein abnormality in Glanzmann's thrombasthenia. Spleen cells from Balb/c mice immunized with human platelets were fused to mouse myeloma cell line Sp2/0-Ag14. Hybridoma lines producing a variety of antiplatelet antibodies were isolated by hypoxanthine-aminopterin-thymidine selection and cloned, and purified monoclonal IgG from six lines was prepared. One of these lines, 8aB5-9, produced an antibody, Tab, that binds to a protein on normal but not thrombasthenic platelets. We isolated this protein from Triton X-100 solubilized normal platelet membranes by affinity chromatography on Tab-Sepharose. As determined by SDS polyacrylamide gel electrophoresis, the isolated protein is a complex of glycoproteins IIb and IIIa, because the two subunits comigrate with glycoproteins IIb and IIIa of whole platelets and show identical changes in mobility after disulfide bond reduction. We prepared (125)I-Tab to determine the number of glycoprotein IIb-IIIa complexes on normal and thrombasthenic platelets by a direct binding assay. Platelets from 17 normal donors bound 39,000+/-4,600 (SD) Tab molecules/platelet. Platelets from four patients with thrombasthenia lacked Tab binding sites (<5%). Five obligate and four presumed heterozygotes for thrombasthenia bound 24,500+/-5,800 Tab molecules/platelet. The platelet alloantigen, Pl(Al), is not that recognized by Tab, because platelets from three Pl(Al)-negative subjects bound Tab normally. Studies with the Tab antibody have (a) enabled quantitation of the number of glycoprotein IIb-IIIa complexes on normal platelet membranes, (b) demonstrated that thrombasthenic homozygotes lack and heterozygotes have a partial deficiency of this complex, and (c) made possible the isolation of this membrane protein which may be required for normal platelet aggregation and clot retraction.
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PMID:Isolation and quantitation of the platelet membrane glycoprotein deficient in thrombasthenia using a monoclonal hybridoma antibody. 644 21

To investigate the role of the leader peptide in modulating secretion from living cells, we injected a synthetic peptide into Xenopus oocytes. The peptide consisted of the NH2-terminal leader sequence of mouse immunoglobulin light chain precursor. We found that the leader peptide has two different roles in regulating secretion from the oocytes. First, it competitively inhibits the synthesis of secretory and membrane proteins but not of cytoplasmic proteins. The inhibition occurs both with oocyte proteins and with proteins directed by coinjected myeloma mRNA. The inhibition reaches a maximum 2 hr after injection and decays within 3 hr. It appears to be mediated through the cell membrane, because 125I-labeled leader peptide segregates into the membrane fraction of microinjected oocytes simultaneously with the interference with methionine incorporation. A second role of the microinjected leader peptide is to induce a rapid acceleration in the rate of export of secretory proteins from the oocyte. The maximal enhancement effect is obtained upon injection of 50 ng of leader peptide per oocyte. It is not merely due to the small size, negative charge, or hydrophobicity of the peptide, because enhanced secretion does not occur when glucagon, poly-L-glutamic acid, or Triton X-100 is injected. Furthermore, immunoreaction of the peptide with specific antibodies prior to microinjection prevents the accelerated export. Our observations indicate that in Xenopus oocytes, the leader peptide is involved in both translocation and later step(s) in the secretory pathway.
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PMID:Synthetic leader peptide modulates secretion of proteins from microinjected Xenopus oocytes. 658 Jun 39

We have developed a procedure to purify the recombinant fusion toxin IL6-PE4E from Escherichia coli which results in a high yield of fully active monomeric protein of high purity and very low endotoxin content. The chimeric toxin is composed of human interleukin 6 (IL6) fused to a derivative of Pseudomonas exotoxin (PE) containing mutations in the binding domain which prevent binding to the PE receptor. In a typical preparation, 20 g of E. coli cells expressing the plasmid encoding IL6-PE4E were treated with lysozyme and washed repeatedly with detergent (Triton X-100), to obtain 500 mg of inclusion bodies. The recombinant protein was denatured and reduced in guanidine hydrochloride solution containing dithioerythritol and refolded in a redox buffer containing oxidized glutathione and L-arginine. After purification of the dialyzed protein by anion-exchange, polymyxin B, and sizing chromatography, we obtained 100 mg (20% of recombinant protein) of purified monomer with 0.6-2.5 endotoxin units/mg of protein. Amino terminal sequencing confirmed the first 20 amino acids. IL6-PE4E purified in this manner was fully cytotoxic toward human multiple myeloma, hepatoma, epidermoid carcinoma, and prostate carcinoma cell lines. After intravenous injection into mice, we found the dose-limiting toxicity to be to the liver, by measurement of serum transaminases and histologic evaluation of the liver. The LD50 was 450 micrograms/kg. We conclude that IL6-PE4E can be purified efficiently for preclinical testing.
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PMID:Purification and characterization of IL6-PE4E, a recombinant fusion of interleukin 6 with Pseudomonas exotoxin. 830 30

To develop an enzyme-linked immunosorbent assay (ELISA) for monitoring the toxicity due to polychlorinated dibenzo-p-dioxins and dibenzofurans contaminated in human breast milk, we have generated novel monoclonal antibodies using some haptenic derivatives linked to bovine serum albumin via the C-1 or C-2 position on the dioxin skeleton. BALB/c or A/J mice were repeatedly immunized with the immunogen, and spleen cells were fused with P3/NS1/1-Ag4-1 myeloma cells. After five fusion experiments, a hybridoma clone was established that secretes an antibody D9-36 group specifically recognizing the major toxic congeners, 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TCDD), 1,2,3,7,8-pentachlorodibenzo-p-dioxin, and 2,3,4,7,8-pentachlorodibenzofran. An ELISA is developed on the basis of the competitive and labeled-antigen format. The toxic congeners extracted from butter or milk specimens by a novel extraction cartridge and a peroxidase-labeled dioxin analogue were sequentially reacted with a fixed amount of D9-36 in the presence of Triton X-100. The bound fraction was captured on a microtiter plate, immobilizing a second antibody, and the enzyme activity was colorimetrically determined. This ELISA afforded a practical sensitivity (measurable range, 1-100 pg/assay; detection limit, 1.0 pg/assay as 2,3,7,8-TCDD equivalent). The assay values for milk and butter samples were in reasonable accordance with the sum of the toxicity-equivalent quantity of each congener, which had been determined by a high-resolution gas chromatography/high-resolution mass spectrometry method.
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PMID:Enzyme-linked immunosorbent assay for monitoring toxic dioxin congeners in milk based on a newly generated monoclonal anti-dioxin antibody. 1505 56

Interstitial cells of Cajal (ICC) are pacemaker cells for the spontaneous muscular contractions and neuromodulators that mediate neurotransmission from enteric neurons to smooth muscle cells in the gastrointestinal (GI) tract. They express c-Kit, and the antibody for c-Kit (especially ACK2) has been a useful tool for functional and morphological studies. ACK2, however, does not work on tissues fixed with paraformaldehyde, and not all ICC express c-Kit in human. Therefore, in order to find a new marker of ICC and/or new antibody resisting aldehyde fixation, we produced a new monoclonal antibody that identifies ICC and then investigated the properties of its antigen. Isolated ICC were used for immunization. Hybridomas fused with myeloma SP2 were screened by immunohistochemistry. ACK2 and each antibody were applied on serial sections, and the clone producing anti-ICC antibody (AIC) that stains ICC was established. The distribution of AIC immunopositive cells was examined in other organs and also GI muscles of W/Wv mice. The biochemical properties were studied using dot blot analysis. AIC recognized ICC; however, distribution of immunopositive cells in W/Wv mice and other organs was different from that of c-Kit. The immunoreactivity was stable for paraformaldehyde but was blocked by either Triton X-100 or SDS. In conclusion, new antibody AIC recognized ICC but the antigen was not c-Kit, which confirms the existence of good markers of ICC besides c-Kit. Although the antigen has not been isolated, AIC is suitable for morphological study and is useful for investigation of ICC in c-Kit mutants.
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PMID:New monoclonal antibody (AIC) identifies interstitial cells of Cajal in the musculature of the mouse gastrointestinal tract. 1529 91

Monoclonal antibodies (MAbs) against glycophorin-A (GPA) could be used in identifying MN blood groups, detecting specific markers of erythroid differentiation, and studying parasite interactions. Large-scale production of MAbs in bioreactors demands an efficient and rapid separation technology. The present study describes the production of a human anti-GPA monoclonal antibody and its purification using a pseudo-bioaffinity L-histidine-convective interaction media (CIM) monolithic column. Hybridomas were generated by fusion of mouse myeloma cell line (Sp2/0) and spleen cells from the mouse immunized with Triton X-100 solubilized RBC membrane proteins. Hybridomas producing antibodies specific to commercial glycophorin-A were screened by indirect enzyme-linked immunosorbent assay (ELISA). The antibodies produced by the stable clones were found to be IgG1 with kappa light chain. Purification of IgG1 MAbs from the cell culture supernatant carried out with a CIM-EDA-histidine disk resulted in high specific activity with purification fold of 8.3 in the fraction eluted with MOPS buffer containing 0.2 M NaCl. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and ELISA showed that the antibodies obtained were highly pure, with high antigen-binding efficiency. The results indicate that faster separation and efficient recovery of high-purity anti-GPA MAbs could be achieved by using CIM-EDA-histidine disk.
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PMID:Production of human anti-glycophorin-A monoclonal antibodies and their purification by pseudoaffinity chromatography using a convective interaction media monolithic column. 2250 14


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