Scavenger Receptor BI (SR-BI) Antibody (for Functional Assays) from Novus Biologicals

March 28, 2013

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Quality of Results

Ease-of-Optimization

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Medical Biochemistry
University of Cape Town, South Africa
Research Officer
R6F cells stably transfected with either SR-B1 or SIGNR1 were incubated with either BCG-lux (left panels) or Mtb-lux (right panels) in the presence or absence of an SR-B1 antibody, and quantified for binding by measuring luciferase activity. The white bars show % of luciferase activity relative to binding in the absence of antibody (black bars) (from: Schäfer et al., PLoS one 4 (12): e8448).

Company:

Novus Biologicals

Product Name:

Scavenger Receptor BI (SR-BI) antibody

Catalog Number:

NB400-134

Image

This antibody was used in a study that investigated the role of Scavenger Receptor BI (SR-BI) for infection with Mycobacterium tuberculosis. It has been published in PLoS One in 2009 (Schäfer et al., The Role of Scavenger Receptor B1 in Infection with Mycobacterium tuberculosis in a Murine Model. PLoS one 4 (12): e8448). Several SR-BI antibodies were tested in this study for various applications; the antibody reviewed here (NB 400-134) worked best for functional ligand blocking experiments.

Experimental Design and Results Summary

Applications

Blocking of ligand binding to SR-B1 on live cells (functional assays)

Sample

Live R6F cells (Rat 6 fibroblast, SR-BI negative cell line) stably transfected with murine SR-BI

Primary Incubation

SR-BI antibody ( Novus Biologicals, cat.no. NB 400-134), 1h at 4°C, 1:100 dilution in DMEM (culture medium)

Blocking Agent

N/A

Secondary Incubation

N/A

Tertiary Incubation

N/A

Detection

After receptor blocking with the antibody, the ligand was added and cells were incubated for 1h at 37°C. The ligands used in our study were live strains of Mycobacterium bovis bacilli Calmette-Guérin (BCG) as well as Mycobacterium tuberculosis H37Rv expressing bacterial luciferase. Quantification of ligands was performed by lysis of the cells in triton X-100 and addition of 1% n-decyl-aldehyde to initiate luciferase reaction. Light emission was measured using a Fluoroscan Ascent FL (Thermo Scientific).

Results Summary

This antibody worked well for the described functional blocking experiments. It was specific for blocking binding of mycobacteria to SR-BI compared to control receptors.

Additional Notes

Of all the SR-BI antibodies tested (Novus Biologicals NB 400-104 and NB 400-113) only this one (Novus Biologicals NB 400-134) reviewed here gave satisfying results in the functional receptor blocking experiments. However, this antibody could not be successfully used for other applications such as Western blotting or FACS analysis. Moreover, we never tested blocking of other known ligands to SR-BI, but blocking of mycobacteria binding to SR-BI worked well with this antibody.

Related Categories

Image Gallery

R6F cells stably transfected with either SR-B1 or SIGNR1 were incubated with either BCG-lux (left panels) or Mtb-lux (right panels) in the presence or absence of an SR-B1 antibody, and quantified for binding by measuring luciferase activity. The white bars show % of luciferase activity relative to binding in the absence of antibody (black bars) (from: Schäfer et al., PLoS one 4 (12): e8448).

Summary

The Good

Suitable for functional receptor blocking experiments.

The Bad

Not suitable for Western blotting or FACS.

The Bottom Line

This is a good antibody to block SR-BI functionally on live cells, i.e. ligand (mycobacteria) binding and uptake could be specifically prevented in the presence of this antibody. However, we never tested antibody-mediated SR-BI blocking with other known ligands.

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