Selection of human antibody fragments – scFv – against immunomodulatory receptors for inflammation control in In Vitro models of viral infection

Introduction: This study investigates the efficacy of a novel approach toidentifying antibodies targeting cellular receptors with a regulatory function in viral infections.It is suggested thatmodulating these receptors can control excessive production ofpro-inflammatory cytokines, potentially serving as a strategictreatment for patients with severe conditions, such as severedengue.In this context, monoclonal antibodies or antibodyfragments emerge as an important strategy for modulating thesereceptors, consequently increasing survival and reducinglethality in critically ill patients.Objectives: To select human antibody fragments (scFv) using thephage display technique against modulatory receptors involvedin the inflammatory process.Methodology: A human scFv library was used for the biopanning ofscFvs against immunomodulatory receptors, followed by scFvexpression, CDR identification, and evaluation throughimmunoenzymatic assays.Results: Three selection cycles (rounds) were performed, optimizedto increase the affinity and specificity of the scFvs.The firstcycle resulted in a 10-fold enrichment, followed by a 22-fold enrichment in the second cycle, while no enrichment wasobserved in the third cycle.Thus, the second selection cyclewas chosen to initiate the screening of scFv-producing andspecific clones.A total of 96 individual clones were screenedusing an ELISA assay to identify specific clones.Among them, 26 clones tested positive, demonstrating significant and specificinteraction with the target antigen (10x cut-off).These 26 clones were sequenced, and their respective CDRs wereidentified, revealing nine different antibody fragments.Basedon screening criteria (predicted CDRs associated with ELISA four fragments were selected for furtherimmunoenzymatic evaluations. Conclusion:The phage display technology successfully identified andselected nine distinct antibody fragments (scFv) againstimmunomodulatory receptors capable of specifically binding toth in viral infections.