Immunocytochemistry/ Immunofluorescence - Anti-6X His tag® antibody [HIS.H8] (ab18184)

LEFT: untransfected control; RIGHT: anti-His (in red) on His-tagged fusion proteins in HEK293 cells. Both counterstained with DAPI (in blue).

Western blot - Anti-6X His tag® antibody [HIS.H8] (ab18184)

ab286909 was shown to bind specifically to SARS-CoV-2 nsp13 protein in western blot. Samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5% milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

Blocking buffer: 5% milk in TBS-0.1% Tween® 20 (TBS-T)

Lanes 1 - 4: Western blot - Anti-6X His tag® antibody [HIS.H8] (ab18184) at 1/1000 dilution

Lanes 5 - 8: Western blot - Anti-SARS-CoV-2 nsp13 antibody [EPR24838-18] (ab286909) at 1/1000 dilution

Lanes 1 and 5: Recombinant SARS-CoV-2 nsp13 protein (His-tagged) at 0.5 µg

Lanes 2 and 6: Recombinant SARS-CoV-2 nsp13 protein (His-tagged) at 0.2 µg

Lanes 3 and 7: Recombinant SARS-CoV2 nsp1 protein (His tagged) at 0.5 µg

Lanes 4 and 8: Recombinant SARS-CoV2 nsp1 protein (His tagged) at 0.2 µg

Secondary

Lanes 1 - 4: Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) at 1/20000 dilution

Lanes 5 - 8: Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/20000 dilution

This data was developed using ab286909, the same antibody clone in a different buffer formulation.