Isolation of a Monoclonal Human scFv Against Cytomegalovirus pp71 Antigen Using Yeast Display

TL;DR

Researchers isolated a new monoclonal antibody targeting a key protein in cytomegalovirus using a yeast-based method.

Contribution

A novel monoclonal antibody against CMV pp71 was developed using yeast display and MACS.

Findings

A monoclonal scFv antibody (ID2) showed high sensitivity and specificity for CMV pp71.

The antibody was validated using flow cytometry, immunofluorescence, ELISA, and BLI.

Abstract

Background: Human cytomegalovirus (CMV) is a major pathogen that poses significant risks to immunocompromised individuals and neonates. The tegument protein pp71, encoded by the UL82 gene, plays a pivotal role in initiating viral lytic replication and evading host immune responses. Despite its clinical relevance, standardized monoclonal antibodies (mAbs) for pp71 remain limited, prompting the need to expand the available repertoire of antibodies targeting this critical protein. Methods: In this study, we constructed a diverse human single-chain variable fragment (scFv) library using RNA derived from the B cells of four healthy donors. The library was expressed in Saccharomyces cerevisiae, and iterative rounds of magnetic-activated cell sorting (MACS) were performed against recombinant pp71. Clonal enrichment was monitored using flow cytometry. Results: Among the isolated clones, one designated ID2 exhibited high sensitivity and specificity for pp71, as demonstrated by flow cytometry, immunofluorescence, an enzyme-linked immunosorbent assay (ELISA), and biolayer interferometry (BLI). Conclusions: Collectively, these findings establish a novel pp71-specific mAb and underscore the utility of yeast surface display combined with MACS for expanding the antibody toolkit available for CMV research and diagnostics.

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