293T cells were transfected by lipofection with 4.0 μg pcACE2-microbody. At 72 hours post-transduction, 0.5 mL of culture supernatant was incubated with nickel-nitrilotriacetic acid-agarose beads (QIAGEN). The beads were washed, and bound protein was eluted with Laemmle loading buffer. The proteins were analyzed on an immunoblot probed with mouse anti-6X His antibody (Invitrogen) and horseradish peroxidase (HRP)-conjugated goat anti-mouse IgG secondary antibody (Sigma-Aldrich). The proteins were visualized and quantified using luminescent substrate on an iBright CL1000 (Invitrogen) gel imager. Ratios were calculated as spike protein divided by P24 band intensities on an identical exposure of the blot.
Horseradish peroxidase hrp conjugated goat anti mouse igg secondary antibody
Manufactured by Merck Group
Horseradish peroxidase (HRP)-conjugated goat anti-mouse IgG secondary antibody is a laboratory reagent used in various immunoassay techniques. It consists of a goat-derived antibody that specifically binds to mouse immunoglobulin G (IgG) molecules, and is conjugated to the enzyme horseradish peroxidase. This conjugate can be used to detect and quantify the presence of mouse IgG in biological samples.
Automatically generated - may contain errors
3 protocols using horseradish peroxidase hrp conjugated goat anti mouse igg secondary antibody
1
Quantification of ACE2 Spike Protein
2
Decoy Protein Binding Assay for SARS-CoV-2 Spike Protein
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Decoy proteins (5, 2, 0.5 and 0.1 μg) were allowed to bind 30 μL of nickel-nitrilotriacetic acid-agarose beads (QIAGEN) for 1 hour after which unbound decoy was removed by washing with PBS. The beads were then incubated with 30 μL (30 μg) of D614G spike protein-pseudotyped virus for 1 hour after which unbound virions were removed by washing with PBS. The bound virions were then eluted from the beads with Laemmle loading buffer containing reducing agent (Invitrogen) and analyzed on an immunoblot probed with anti-p24 monoclinal antibody AG3.0 (Creative Biolabs) and horseradish peroxidase (HRP)-conjugated goat anti-mouse IgG secondary antibody (Sigma-Aldrich). The signals were developed with Luminata Crescendo Western HRP Substrate (Millipore) and membranes were visualized on an iBright imaging system (Invitrogen).35 (link)
3
Quantifying SARS-CoV-2 Spike Protein
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Different amounts (1, 0.5 and 0.1 μg) of sACE2 was incubated with 30 μl of nickel-nitrilotriacetic acid-agarose beads (QIAGEN). After 1 hour of incubation, beads were washed with PBS, and 30µL of virus was added. Post 1 hour of incubation, the beads were washed, and bound protein was eluted with Laemmli loading buffer. The proteins were analyzed on an immunoblot probed with anti-p24 mAb (AG3.0) and horseradish peroxidase (HRP)-conjugated goat anti-mouse IgG secondary antibody (Sigma-Aldrich).
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