RÉSUMÉ
BACKGROUND: More than 3 million infections were attributed to Chikungunya virus (CHIKV) in the 2014-2016 outbreak in Mexico, Central and South America, with over 500 deaths directly or indirectly related to this viral disease. CHIKV outbreaks are recurrent and no vaccine nor approved therapeutics exist to prevent or treat CHIKV infection. Reliable and robust diagnostic methods are thus critical to control future CHIKV outbreaks. Direct CHIKV detection in serum samples via highly specific and high affinity anti-CHIKV antibodies has shown to be an early and effective clinical diagnosis. METHODS: To isolate highly specific and high affinity anti-CHIKV, Chikungunya virions were isolated from serum of a patient in Veracruz, México. After purification and characterization via electron microscopy, SDS-PAGE and binding to well-characterized anti-CHIKV antibodies, UV-inactivated particles were utilized as selector in a solid-phase panning in combination with ALTHEA Gold Libraries™, as source of antibodies. The screening was based on ELISA and Next-Generation Sequencing. RESULTS: The CHIKV isolate showed the typical morphology of the virus. Protein bands in the SDS-PAGE were consistent with the size of CHIKV capsid proteins. UV-inactivated CHIKV particles bound tightly the control antibodies. The lead antibodies here obtained, on the other hand, showed high expression yield, > 95% monomeric content after a single-step Protein A purification, and importantly, had a thermal stability above 75 °C. Most of the antibodies recognized linear epitopes on E2, including the highest affinity antibody called C7. A sandwich ELISA implemented with C7 and a potent neutralizing antibody isolated elsewhere, also specific for E2 but recognizing a discontinuous epitope, showed a dynamic range of 0.2-40.0 mg/mL of UV-inactivated CHIKV purified preparation. The number of CHIKV particles estimated based on the concentration of E2 in the extract suggested that the assay could detect clinically meaningful amounts of CHIKV in serum. CONCLUSIONS: The newly discovered antibodies offer valuable tools for characterization of CHIKV isolates. Therefore, the strategy here followed using whole viral particles and ALTHEA Gold Libraries™ could expedite the discovery and development of antibodies for detection and control of emergent and quickly spreading viral outbreaks.
