Production of Biopharmaceutical Proteins from Scallop Hemocytes as a Bioreactor Using Improved Transfection Conditions.

Despite the widespread use of protein-based biopharmaceuticals, conventional production systems using mammalian cells and E.coli have some limitations, including the absence of post-translational modifications and potential risk of viral contamination. To address this, we investigated a novel platform using primary hemocyte culture from Yesso scallop (Mizuhopecten yessoensis) as a bioreactor, leveraging the strong activity of the OsHV-1 promoter in bivalve cells. The optimal condition for cell culture and transfection in a 12-well plate were established by adjusting the Penicillin-Streptomycin concentration, seeding cell number, hemolymph-to-medium ratio, and the volume of transfection reagent to 20 U/mL, 2 × 106 cells/well, 600 µL of diluted hemolymph to 300 µL of L-15 × 2, and 200 µL of X-tremeGENE transfection component into 900 µL of culture medium, respectively. Under these optimized conditions, recombinant human growth hormone (hGH) was successfully expressed in scallop hemocyte culture and its expression was confirmed at both mRNA and protein levels. The yield of hGH extracted from hemocyte cultures was up to 13.92 ± 4.77 pg/1 × 106 cells in a 12-well plate. We also achieved expression and secretion of human erythropoietin (hEPO) into the culture medium. This study is the first to demonstrate not only human gene expression in molluscan cells but also propose a new platform using molluscan cells for in vitro biopharmaceutical production.