Pierce high sensitivity streptavidin hrp conjugate

The capacity of soluble ANDV Gn^H/Gc (sGn^H/Gc) or rVSVs bearing ANDV, SNV, or HTNV Gn/Gc to recognize sEC1-2, and rVSVs bearing ANDV Gn/Gc to recognize sEC1-4 was determined by capture ELISA. High-protein binding 96-well ELISA plates (Corning) were coated with purified sEC1-2 or sEC1-4 (100 ng/well) overnight at 4 °C, washed briefly with PBS, and blocked with 5% nonfat dry milk in PBS (1 h at RT). ANDV sGn^H/Gc was serial diluted twofold with an initial starting dilution of 6.25 µg/well. Equivalent amounts of rVSV-ANDV-Gn/Gc, rVSV-SNV-Gn/Gc, and rVSV-HTNV-Gn/Gc were membrane-labeled with FSL-biotin as described above, and serially diluted (twofold). ANDV sGn^H/Gc and rVSVs were added to either sEC1-2 or sEC1-4 coated plates and incubated for 1 h at 37 °C. Bound ANDV sGn^H/Gc was detected by incubation with Strep-Tactin®-HRP conjugate (1:5,000 dilution, IBA Lifesciences) and rVSVs were detected by incubation with Pierce^TM High Sensitivity Streptavidin-HRP conjugate (1:10,000 dilution, Thermo Fisher). Coated sEC1-4 protein was detected by incubation with an anti-Flag, clone M2 monoclonal antibody (mAb)-HRP conjugate (1:1,000 dilution, catalog No. A8592, Sigma-Aldrich) (1 h at 37 °C). ELISA signal was developed and measured as noted above.

The capacity of soluble ANDV Gn^H/Gc (sGn^H/Gc) or rVSVs bearing ANDV, SNV, or HTNV Gn/Gc to recognize sEC1-2, and rVSVs bearing ANDV Gn/Gc to recognize sEC1-4 was determined by capture ELISA. High-protein binding 96-well ELISA plates (Corning) were coated with purified sEC1-2 or sEC1-4 (100 ng/well) overnight at 4 °C, washed briefly with PBS, and blocked with 5% nonfat dry milk in PBS (1 h at RT). ANDV sGn^H/Gc was serial diluted twofold with an initial starting dilution of 6.25 µg/well. Equivalent amounts of rVSV-ANDV-Gn/Gc, rVSV-SNV-Gn/Gc, and rVSV-HTNV-Gn/Gc were membrane-labeled with FSL-biotin as described above, and serially diluted (twofold). ANDV sGn^H/Gc and rVSVs were added to either sEC1-2 or sEC1-4 coated plates and incubated for 1 h at 37 °C. Bound ANDV sGn^H/Gc was detected by incubation with Strep-Tactin®-HRP conjugate (1:5,000 dilution, IBA Lifesciences) and rVSVs were detected by incubation with Pierce^TM High Sensitivity Streptavidin-HRP conjugate (1:10,000 dilution, Thermo Fisher). Coated sEC1-4 protein was detected by incubation with an anti-Flag, clone M2 monoclonal antibody (mAb)-HRP conjugate (1:1,000 dilution, catalog No. A8592, Sigma-Aldrich) (1 h at 37 °C). ELISA signal was developed and measured as noted above.

The capacity of soluble ANDV Gn^H/Gc (sGn^H/Gc) or rVSVs bearing ANDV, SNV, or HTNV Gn/Gc to recognize sEC1-2, and rVSVs bearing ANDV Gn/Gc to recognize sEC1-4 was determined by capture ELISA. High-protein binding 96-well ELISA plates (Corning) were coated with purified sEC1-2 or sEC1-4 (100 ng/well) overnight at 4 °C, washed briefly with PBS, and blocked with 5% nonfat dry milk in PBS (1 h at RT). ANDV sGn^H/Gc was serial diluted twofold with an initial starting dilution of 6.25 µg/well. Equivalent amounts of rVSV-ANDV-Gn/Gc, rVSV-SNV-Gn/Gc, and rVSV-HTNV-Gn/Gc were membrane-labeled with FSL-biotin as described above, and serially diluted (twofold). ANDV sGn^H/Gc and rVSVs were added to either sEC1-2 or sEC1-4 coated plates and incubated for 1 h at 37 °C. Bound ANDV sGn^H/Gc was detected by incubation with Strep-Tactin®-HRP conjugate (1:5,000 dilution, IBA Lifesciences) and rVSVs were detected by incubation with Pierce^TM High Sensitivity Streptavidin-HRP conjugate (1:10,000 dilution, Thermo Fisher). Coated sEC1-4 protein was detected by incubation with an anti-Flag, clone M2 monoclonal antibody (mAb)-HRP conjugate (1:1,000 dilution, catalog No. A8592, Sigma-Aldrich) (1 h at 37 °C). ELISA signal was developed and measured as noted above.