M Sárdy1, U Odenthal, S Kárpáti, M Paulsson, N Smyth. 1. Department of Dermato-Venereology, Semmelweis University of Medicine, H-1085 Budapest, Mária u. 41, Hungary. Miklos.Sardy@Uni-Koeln.DE
Abstract
BACKGROUND: Tissue transglutaminase (TGc) has recently been identified as the major, if not the sole, autoantigen of gluten-sensitive enteropathy (GSE). We developed and validated an ELISA based on the human recombinant antigen and compared it to existing serological tests for GSE [guinea pig TGc ELISA and endomysium antibody (EMA) test]. METHODS: Human TGc was expressed in the human embryonic kidney cell line 293-EBNA as a C-terminal fusion protein with the eight-amino acid Strep-tag II allowing one-step purification via streptavidin affinity chromatography. We carried out ELISA assays for IgA antibodies against TGc using calcium-activated human and guinea pig TGc. The sera were also tested on monkey esophagus sections by indirect immunofluorescence for IgA EMA. We examined 71 serum samples from patients with GSE (38 with celiac disease, 33 with dermatitis herpetiformis), including 16 on therapy, and 53 controls. RESULTS: The human TGc could be expressed and purified as an active enzyme giving a single band on a Coomassie-stained gel. The mean intra- and interassay CVs for the human TGc ELISA were 3.2% and 9.2%, respectively. The area under the ROC curve was 0.999. The specificity and sensitivity were 98.1% (95% confidence interval, 95.7-100%) and 98.2% (95.9-100%), respectively. CONCLUSIONS: The human TGc ELISA was somewhat superior to the guinea pig TGc ELISA, and was as specific and sensitive as the EMA test. The human TGc-based ELISA is the method of choice for easy and noninvasive screening and diagnosis of GSE.
BACKGROUND:Tissue transglutaminase (TGc) has recently been identified as the major, if not the sole, autoantigen of gluten-sensitive enteropathy (GSE). We developed and validated an ELISA based on the human recombinant antigen and compared it to existing serological tests for GSE [guinea pigTGc ELISA and endomysium antibody (EMA) test]. METHODS:HumanTGc was expressed in the humanembryonic kidney cell line 293-EBNA as a C-terminal fusion protein with the eight-amino acid Strep-tag II allowing one-step purification via streptavidin affinity chromatography. We carried out ELISA assays for IgA antibodies against TGc using calcium-activated human and guinea pigTGc. The sera were also tested on monkey esophagus sections by indirect immunofluorescence for IgA EMA. We examined 71 serum samples from patients with GSE (38 with celiac disease, 33 with dermatitis herpetiformis), including 16 on therapy, and 53 controls. RESULTS: The humanTGc could be expressed and purified as an active enzyme giving a single band on a Coomassie-stained gel. The mean intra- and interassay CVs for the humanTGc ELISA were 3.2% and 9.2%, respectively. The area under the ROC curve was 0.999. The specificity and sensitivity were 98.1% (95% confidence interval, 95.7-100%) and 98.2% (95.9-100%), respectively. CONCLUSIONS: The humanTGc ELISA was somewhat superior to the guinea pigTGc ELISA, and was as specific and sensitive as the EMA test. The humanTGc-based ELISA is the method of choice for easy and noninvasive screening and diagnosis of GSE.
Authors: P D Thomas; A Forbes; J Green; P Howdle; R Long; R Playford; M Sheridan; R Stevens; R Valori; J Walters; G M Addison; P Hill; G Brydon Journal: Gut Date: 2003-07 Impact factor: 23.059
Authors: Johannes Wolf; Norman Haendel; Johannes Remmler; Carl Elias Kutzner; Thorsten Kaiser; Thomas Mothes Journal: J Clin Lab Anal Date: 2017-11-23 Impact factor: 2.352