Concentração de partículas pseudovirais de Zika vírus por ultrafiltração

Abstract

This work assessed the separation of virus-like particles (VLPs) through ultrafiltration operation. The VLPs were constitutively expressed by HEK293 - 3F6 cell line (Human Embryonic Kidney cells) transfected with commercial plasmidial vector PCINEO® (Promega) encoding the genetic sequences of premembrane (pr-M) and envelope (E) structural proteins of Zika virus. The assessment techniques were based on qualitative analysis, such as the Immunoblot test, SDS-PAGE and Western blot; protein quantification, through densitometry and Bradford assays; and the particle diameter distribution analysis by using HPLC/SEC-UV. Sequential batches of 14 mL in Vivaspin® devices embedded with polyethersulfone of 300 kDa and 1000 kDa membranes were analyzed and better performance was achieved with the lowest cut-off. Therefore, batch operations in agitated Amicon® cell filled with 200 mL of clarified supernant were conducted with 500 kDa polyethersulfone and 100 kDa regenerated cellulose membranes. Despite both membranes showing concentration capacity to Zika virus-like particles, the PES membrane displayed higher efficiency.