Recombinant Immunoglobulin A Specific for Influenza A Virus Hemagglutinin: Production, Functional Analysis, and Formation of Secretory Immunoglobulin A

Abstract

Secretory immunoglobulin (Ig) A (SIgA), comprised of dimeric IgA and secretory component (SC), is believed to provide a defense mechanism on the mucosal surface. Influenza A virus (IAV) hemagglutinin (HA)-specific SIgA is thought to play an important role in the prevention of IAV infection. However, the topical application of preformed IAV-specific SIgA has not been shown to prevent IAV infection. This is due to the difficulty in the production of antigen-specific IgA monoclonal antibodies (mAbs) and monoclonal SIgA. Here, a recombinant hybrid IgA (HIgA) was established that utilizes variable regions of an HA-specific mouse IgG mAb and the heavy chain constant region of a mouse IgA mAb. We expressed the dimeric HIgA in Chinese hamster ovary-K1 (CHO-K1) cells. When in vitro IAV infection of Madin-Darby canine kidney (MDCK) cells was tested, 10 times lower concentrations of HIgA were able to inhibit it as compared with an HA-specific IgG with the same variable regions. A functional hybrid secretory IgA (HSIgA) was also produced through incubation of the dimeric HIgA with recombinant mouse SC in vitro. It was demonstrated that HSIgA could be separated from the dimeric HIgA on size exclusion chromatography. This study provides a basic strategy for investigating the role of SIgA upon IAV infection on the mucosal surface.

Protection of Human Colon Cells from Shiga Toxin by Plant-based Recombinant Secretory IgA

“…The subcloned J chain cDNA was digested with Xba I/ Xho I and ligated into the pGEM5zf vector (Promega, Madison, WI, USA) harbouring chlorophyll a/b -binding protein terminator 2 (T CAB2 )25, resulting in the production of pGEM5zf/Jc-T CAB2 . To subclone mouse SC cDNA, PCR was performed using pcDNA3.1/mouse SC45 as a template, and SC-specific SCF- Xho and SCR- Xba as primers. Following Xba I/ Xho I digestion and generation of blunt ends, SC cDNA was inserted into Eco RV-digested pGEM5zf harbouring T CAB1 (pGEM5zf/T CAB1 ), resulting in the production of pGEM5zf/SC-T CAB1 .…”