Coding DNA of the recombinant panfilovirus mAb 7C8F1: cloning and expression in CHO mammalian cell lines

Abstract Background: Uganda recently experienced an outbreak of sudan ebolavirus (SUDV) species, for which there are no approved or licensed rapid diagnostic tests (RDTs). Our group has a longstanding history in the research and development (R & D) of panfilovirus RDTs. This paper describes sequencing of the coding DNA of a recombinant monoclonal antibody (7C8F1) targeting the epitope (YEAGEWAENCY, dubbed peptide 2) conserved across all ebolavirus spp glycoproteins (GP). Methods: Mice hybridoma were prepared & cultured on Dulbecco’s modified eagle’s medium (DMEM) Total RNA was extracted using RNA-easy isolation reagent (Vazyme), and reverse transcribed into cDNA using SMARTScribe reverse transcriptase. Antibody fragments of heavy (H) and light (L) chains were amplified by rapid amplification of cDNA ends (RACE) and cloned into standard vectors for screening in 5 colonies. Inserts of matching phenotypic colonies were sequenced by Sanger capillary sequencing and aligned by IMGT analysis of VDJ. Isotyping was done by constant region analysis. Results: Consensus sequences of the coding DNA of the heavy and light chains of an isotype IgG1/kappa recombinant mice mAb (Genbank accession #s OP966768 and OP966769, respectively) are presented. The biophysical profile of the full expressed recombinant mAb demonstrate a single protein of approximately 142kda under non reducing conditions and tetramer of duplexes of 23ka and 48kda, respectively. The average area under sec-HPLC curve at a wavelength of 280nm is 2490.55 Conclusions: We present the coding DNA of the recombinant panfilovirus mAb 7C8F1 and demonstrate cloning and expression of its purified extracts within CHO mammalian cell lines.