Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0011854 (type 1 diabetes)
 20,749 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Because of the low titer of ICSA, as well as the polyclonality of serum, it has been difficult to determine whether there is a variety of autoantibodies, each reacting with a different organ, or whether the autoimmune response is more restricted, with only a limited number of autoantibodies recognizing common antigenic determinants in the different organs. To study this and other questions, we sought to prepare human monoclonal autoimmune ICSA. Peripheral-blood lymphocytes from patients with freshly onset IDDM or other autoimmune abnormalities were transformed by EB virus. And rapid screening on In 111 cells by an ELISA were used to identify monoclonal ICSA. But, ICSA titer was very low even in freshly onset IDDM patients, and reduced very quickly. ICSA positive wells at cloning were extremely rare. So, the methods that enhance the efficiency of cloning were required. Firstly, ultrasensitive Enzyme Immunoassay for ICSA were developed. Secondly, new preselection method of ICSA producing lymphocyte by Protein A Magnetic Microspheres were employed. Thirdly, in vitro immunization was partly employed to enhance the frequency of ICSA producing lymphocytes. Fourthly, EBV-hybridoma technique was employed to stabilize the ICSA producing clone. The efficiency of cloning was increased from 0.07% to 24% by the new preselection method. The sensitivity of the fluorometric enzyme immunoassay for ICSA was 140pg/ml. 21 times sensitivity to usual ELISA was obtained. We describe sure methods for production of Human Monoclonal Autoantibodies that react with Islet-Cell-Surface Antigens.
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PMID:[Production of human monoclonal autoantibodies that react with islet-cell-surface antigens by EBV-transformed lymphocytes: new method of preselection of ICSA producing lymphocyte by protein A magnetic microspheres]. 285 14

Monoclonal antibody 3A4 to islet cell surface antigen has been previously established in our laboratory, using hybridization of spleen lymphocytes from non-obese diabetic (NOD) mice transferred into immunologically incompetent recipient mice. In the present study, monoclonal islet cell surface antibody 5C12 could be newly obtained in the 10:1 ratio of NOD mice spleen cells and mouse myeloma cells (SP2/0) without any modifications. Protein A radioligand assay and indirect immunofluorescence on living cells showed that 5C12 antibody reacted to normal rat islet cells and cultured rat insulinoma cells (RIN-r), but not to cultured lymphocytes (Bri-7, IM-9) and Chang-liver cells. Analysis of 125I-labeled antibody binding revealed that unlabeled 5C12 effectively inhibited subsequent 125I-5C12 binding to RIN-r cells, whereas unlabeled 3A4 did not. The scatchard plot from these data showed the curvilinearity, and about 150,000 binding sites to antibody per RIN-r cell were counted. The treatment of RIN-r cells with papain and neuraminidase reduced the binding of 5C12 to RIN-r cells, whereas the effect of trypsin was not observed. Immunoprecipitation of 125I-labeled insulinoma cell lysates followed by SDS-PAGE and autoradiography indicated that 5C12 recognized 105K dalton cell surface protein in RIN-r cells. Immunoblotting also showed that 5C12 antibody recognized 105K dalton cell surface protein in RIN-r cells. These results demonstrated that 5C12 was an important tool for clarifying the immunoresponse against certain antigenic determinants on pancreatic B cells. Furthermore, 5C12 has not only qualitatively and quantitatively improved diagnostic methodology, but it may also provide new reagents useful to the treatment and prevention of type 1 diabetes.
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PMID:[An analysis of islet cell surface antigen defined by monoclonal islet cell surface antibody 5C12]. 354 94

The characterization of target antigens in several autoimmune disorders has made it possible to develop antigen-specific immunoassays that are superior in terms of sensitivity, specificity, reproducibility and ease of standardization, compared to immunohistological methods that are highly subjective, rely on skilled technicians and are not applicable to large-scale studies. In the case of celiac disease (CD), tissue transglutaminase (tTGase) has been identified as a major autoantigen, and antibodies against this molecule are present in most CD patients before gluten is removed from diet. In general, anti-tTGase detection assays detect the presence of IgA class antibodies, but these immunoglobulins are absent among patients with IgA deficiency, a frequent condition in which CD is very prevalent. In this report, we have analyzed 64 patients at diagnosis of CD for the presence of antibodies against tTGase of both IgA (TGA) and IgG (TGG) classes, using anti-IgA antibodies or Protein A, respectively, for the immunoprecipitation of 35S labeled, in vitro transcribed and translated human recombinant tTGase. In our hands, the TGG assay matches TGA in terms of sensitivity (97%) and specificity (100%), and besides, the combination of both assays is able to detect antibodies in all patient samples. The method described uses only 6 microl of serum, can be adapted to automated large-scale analysis and, in combination with other antigens, can be used for the simultaneous screening of other autoimmune diseases, like type 1 diabetes mellitus.
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PMID:Immunoglobulin G autoantibodies against tissue-transglutaminase. A sensitive, cost-effective assay for the screening of celiac disease. 1248 93

The GAD65 epitope immunoglobulin binding pattern in cord blood of children (n=37), who later developed type 1 diabetes at 3.2-14.9 years of age, was analyzed. First, the binding at diagnosis was compared with that in the cord blood serum. The next comparison was between the cord blood serum and the mothers' serum taken at delivery. Basal GAD65 binding levels were determined in Protein A Sepharose-based radiobinding assays with (35)S-labeled human and rat GAD65, rat GAD67 and GAD65/67 fusion proteins representing N-terminal (N), middle (M) and C-terminal (C) epitopes. In the first comparison, 28/37 children had GAD65 binding above 2.44 relative units (RU) (upper three quartiles), representing a marked increase from birth in the binding to human GAD65 (p<0.0001), rat GAD65 (p<0.0001), N- (p=0.04), M- (p<0.0001), C- (p=0.001), and M + C-epitopes (p<0.0001), but not to rat GAD67. At birth, 9/37 had GAD65 binding above 1.56 RU (upper quartile) demonstrating that their binding of human (35)S-GAD65 was higher in cord blood than in the mother (p=0.008). Higher cord blood binding was also observed for the N- (p=0.02) terminal epitope but not for rat GAD65, rat GAD67, and the remaining epitopes. These data suggest that differences in the epitope GAD65 binding between mother and child at birth are limited. In contrast, the epitope pattern at diagnosis differed from that at birth, supporting the view that disease-associated epitopes develop between birth and diagnosis.
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PMID:Epitope analysis of GAD65 binding in both cord blood and at the time of clinical diagnosis of childhood type 1 diabetes. 1799 32