BACKGROUND: Conflicting data have been published concerning the effect of calcium on binding of autoantibodies to tissue transglutaminase (tTG) in celiac disease (CD). METHODS: IgA-tTG and IgG-tTG were measured with radioligand binding assays (RBA) using human recombinant (hr) (35)S-tTG produced in lysate of rabbit reticulocytes and with guinea pig (gp) tTG ELISA in 51 CD children (median: 5.7 years) and 35 controls (median: 2.2 years). Assays were performed with and without calcium. RESULTS: In hr-tTG RBA, IgA-tTG levels remained unchanged after calcium detecting 50/51 CD children and 1/35 controls (p<0.0001). IgG-tTG levels decreased with calcium (p<0.0001) in CD children and detected 48/51 with and 49/51 without calcium as compared to 1/35 controls (p<0.0001). In gp-tTG ELISA, levels increased with calcium (p<0.0001) making it possible to detect an additional three to a total of 50/51 with IgA-tTG and 13 to 39/51 CD children with IgG-tTG compared to 4/35 and 8/35 controls (respectively, p<0.0001). Rabbit reticulocytes displayed calcium-dependent tTG activity. CONCLUSIONS: Calcium increased binding of IgA-tTG and IgG-tTG in the ELISA test. The reverse effect observed in RBA may be explained by competitive binding between calcium activated native rabbit reticulocyte tTG and hr (35)S-tTG. tTG autoantibody assays may need taking calcium into account for accurate diagnostic sensitivity and specificity for CD.
BACKGROUND: Conflicting data have been published concerning the effect of calcium on binding of autoantibodies to tissue transglutaminase (tTG) in celiac disease (CD). METHODS: IgA-tTG and IgG-tTG were measured with radioligand binding assays (RBA) using human recombinant (hr) (35)S-tTG produced in lysate of rabbit reticulocytes and with guinea pig (gp) tTG ELISA in 51 CD children (median: 5.7 years) and 35 controls (median: 2.2 years). Assays were performed with and without calcium. RESULTS: In hr-tTG RBA, IgA-tTG levels remained unchanged after calcium detecting 50/51 CD children and 1/35 controls (p<0.0001). IgG-tTG levels decreased with calcium (p<0.0001) in CD children and detected 48/51 with and 49/51 without calcium as compared to 1/35 controls (p<0.0001). In gp-tTG ELISA, levels increased with calcium (p<0.0001) making it possible to detect an additional three to a total of 50/51 with IgA-tTG and 13 to 39/51 CD children with IgG-tTG compared to 4/35 and 8/35 controls (respectively, p<0.0001). Rabbit reticulocytes displayed calcium-dependent tTG activity. CONCLUSIONS:Calcium increased binding of IgA-tTG and IgG-tTG in the ELISA test. The reverse effect observed in RBA may be explained by competitive binding between calcium activated native rabbit reticulocyte tTG and hr (35)S-tTG. tTG autoantibody assays may need taking calcium into account for accurate diagnostic sensitivity and specificity for CD.