title: Antibodies to tissue transglutaminase: comparison of ELISA and immunoprecipitation assay in the presence and in the absence of calcium ions.
abstract: Endomysial antibodies are characteristic of coeliac disease and tissue transglutaminase (tTG) has been identified as a major component of the endomysial antigen. tTG autoantibodies were measured in sera from patients with coeliac disease using ELISA based on guinea pig tTG and immunoprecipitation assay (IPA) based on 35S-labelled human tTG produced in an in vitro transcription/translation system. In addition, the effect of calcium ions on the interaction between tTG autoantibodies in the two assays was studied. Under standard (i.e. Ca2+-free) conditions, 36/39 (92%) coeliac sera were positive for IgA tTG antibodies by ELISA and 34/39 (87%) sera were positive by IPA. Comparison of ELISA and IPA results showed three sera positive by ELISA but negative by IPA and one serum which was positive by IPA but negative by ELISA. Bland and Altman analysis of the correlation between the ELISA and IPA showed that the results for 37 out of 39 samples were in the agreement. The results by ELISA carried out without and with Ca2+ were in good agreement (r=0.99; n=39). IPA using Ca2+ containing buffer detected fewer samples compared to IPA using standard assay buffer however the results of the two assays also showed a good agreement (r=0.93; n=39). Our studies confirm that antibodies to tTG are good markers of coeliac disease and indicate that the autoantibody binding sites on tTG are formed in a way which is essentially independent of Ca2+.
authors.affiliation: FIRS Laboratories, RSR Ltd., Parc Ty Glas, Llanishen, Cardiff CF14 5DU, UK.
authors.forename: K
authors.initials: K
authors.lastname: Nakachi
authors.forename: G
authors.initials: G
authors.lastname: Swift
authors.forename: D
authors.initials: D
authors.lastname: Wilmot
authors.forename: C
authors.initials: C
authors.lastname: Chapman
authors.forename: S
authors.initials: S
authors.lastname: Baker
authors.forename: M
authors.initials: M
authors.lastname: Powell
authors.forename: J
authors.initials: J
authors.lastname: Furmaniak
authors.forename: B
authors.initials: B
authors.lastname: Rees Smith
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001323:Autoantibodies / Q000097:blood*; D002118:Calcium / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D011233:Precipitin Tests; D012015:Reference Standards; D015203:Reproducibility of Results; D011503:Transglutaminases / Q000276:immunology*
chemical_list: D001323:Autoantibodies; D011503:Transglutaminases; D002118:Calcium