Generación de moléculas derivadas de anticuerpos de IgY (Gallus gallus domesticus) para la detección antigénica del Protoparvovirus carnivoro 1

Abstract

Canine parvovirus 2 (CPV-2), a subspecies of carnivore protoparvovirus 1, is one of the main causes of viral gastroenteritis in dogs, generating high morbidity and mortality rates. The objective of this study was to design, express, and validate recombinant proteins based on IgY antibody fragments derived from Gallus gallus domesticus, specifically αVP2scFv and its mimetic variant αVP2scFvM, aimed at the antigenic detection of CPV-2.The genetic constructs were designed in silico and cloned into expression vectors pET28-SP-MCS-APEc and pET15- MCS-PoBP, which incorporated polystyrene-binding peptides (PoBP) and alkaline phosphatase (APEc) as adhesion domains and colorimetric signal generation. The recombinant proteins were heterologously expressed in Escherichia coli Rosetta 2 (DE3) and purified by immobilized metal affinity chromatography. Functional validation was performed using immunoenzymatic assays, capture ELISA, and Dot blot, using antigenic extracts of VP2, commercial vaccines, and biological samples, demonstrating specific antigen recognition and detectable enzymatic activity in the dot blot. The results showed that the recombinant proteins developed constitute a viable platform for the antigenic diagnosis of CPV-2 in the veterinary field.