Consensus DNA and amino acids sequences of the recombinant panfilovirus glycoprotein specific mAb 4B3B5 and its expression in CHO mammalian cell lines

Abstract

Abstract Background: Monoclonal antibodies (mAbs) are a useful resource towards the detection and treatment of dangerous pathogens. The traditional approach to production of mAbs is, however, cumbersome and time consuming. An alternative option, would be to sequence, sub clone & express the genes encoding the mAbs from within a mammalian cell-line. This paper describes the consensus gene and amino acids sequences a recombinant mAb (4B3B5) targeting a conserved epitope (#1) with all filovirus glycoproteins, and its expression in CHO mammalian cells. 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 OP966770 and OP966771, respectively) are presented. The biophysical profile of the full expressed recombinant mAb demonstrates a single protein of approximately 142 kda under non reducing conditions and tetramer of duplexes of 23ka and 48kda, respectively. The average area under sec-HPLC curve at a wavelength of 280 nm is 2490.55 Conclusions: This work summarizes the consensus DNA and amino acids sequences of the recombinant panfilovirus glycoprotein specific mAb 4B3B5 and its expression in CHO mammalian cell lines.