Development of an alternative method to determine the presence of antibodies to deamidated gliadin by SPR

DE DIEGO, G.A.; CERNY, N.; DÍAZ, M.E.; MARTÍNEZ RUIZ, B.L.; IACONO, R.F.; FERNÁNDEZ, M.M.; DE MARZI, M.C.

Mar del Plata

Congreso; LXVIII Reunión Anual de la Sociedad Argentina de Investigación Clínica (SAIC); 2023

Sociedad Argentina de Investigación Clínica (SAIC)

Celiac disease (CD) is a chronic autoimmune enteropathy with areported prevalence of approximately 1%. The associated symp-toms and the severity of the immune response triggered by glu-ten exposure vary between patients. Several antibodies are usedas biomarkers for the diagnosis and monitoring of this pathology.The kinetic interaction and the affinity of autoantibodies for auto-antigens could influence the evolution and clinical presentation ofthe pathology. In recent years, anti-deamidated gliadin antibodies(a-DGP) have become more relevant. Therefore, this work aims todevelop surface plasmon resonance (SPR) assays to determinethe presence of a-DGP and their avidity characteristics in patientsera. DGP was obtained by deamination of gliadin in acidic mediumand separation of the products using an anion exchange column(Mono Q 5/50 GL) in FPLC. Sera samples were obtained from CDpatients who completed a questionnaire with symptom and clinicaldata. Antibodies levels anti-transglutaminase, gliadin and DGP weredetermined by ELISA and expressed as a positivity index (mean ofsample / (mean + 3 SD of controls)). In a preliminary qualitativeassay a-DGP by SPR, DGP (1200 RU, active cell) were immobilizedon a CM5 chip. Different dilutions of positive and negative sera inPBS (1:100, 1:200 and 1:400) were injected over the active surfaceand the binding compare with the reference surface. The resultsobtained were analyzed with BIAevaluation software. The 1:100and 1:200 dilutions of positive sera showed a differential bindingcompare with the negative sera. These promising results aim us todevelop an alternative method to determine the seroprevalence ofa-DGP antibodies. In addition, it allows us to analyze the kinetic pa-rameters of the antibodies and the humoral response, which wouldcorrelate with the severity of the pathology.