Bright Staining for IFNgamma

April 13, 2020

BlueskyReddit
Charite Strasse 1
Deutsches Rheuma-Forschungszentrum
Research Scientist

Overall

Quality of Results

Ease-of-Optimization

What do these ratings mean?
Write a Review

Company:

BD Biosciences

Product Name:

PE-Cy7 anti-IFNgamma antibody

Catalog Number:

557649

Clone Number:

XMG1.2

Browse Similar Products: IFN-Gamma Flow Cytometry Antibodies

In different healthy and pathological contexts CD4 T cells differentiate into a range of specialized interleukin-secreting subsets. In mouse models of aging and autoimmune diseases we study the plasticity and effector functions of CD4 T helper cells. We used anti-IFNgamma to quantify the number of IFNgamma-producing CD4 T cells in old C57Bl/6J mice.

Experimental Design and Results Summary

Applications

Flow Cytometry

Sample

Splenocytes of wild-type 24 months-old C57Bl/6J mouse, incubated in complete culture medium at 37 °C with (Restimulated) or without (unstimulated) PMA/ionomycin for 4h and brefeldin A for the 3 last hours.

Primary Incubation

Surface staining with anti-CD3 Alexa700 and anti-CD4 Pacific Blue in PBS 0.5% BSA incubated 30min at 4°C. Washed twice in PBS and incubated with L/D fixable aqua dye 20 min at 4°C, then washed twice in PBS.

Blocking Agent

Clone 2.4G2 (Fc block)

Secondary Incubation

Cells are then fixed and permeabilized with commercial kit (30 min at 4°C with fixative, and 15 min at 4°C with permeabilization buffer). Cells are finally incubated with anti-IFNgamma at 1/400 in permeabilization buffer for 30 min at 4°C, and washed once with permeabilization buffer and once with PBS BSA 0.5%.

Tertiary Incubation

N/A

Detection

Flow cytometry, yellow green laser

Results Summary

I could quantify IFNgamma-producing CD4 T cells in the spleen

DOI or PMID #

N/A

Additional Notes

N/A

Related Categories

Image Gallery

Summary

The Good

The antibody gives bright signal

The Bad

There is spreading in other channels reading Cy7

The Bottom Line

This antibody gives a very good signal, but it needs a bit of preliminary work to design the panel and avoid too much spill over in detectors reading longest wave lengths (Cy7, and BV786)

Share your experience with other scientists. Write a Review! »

Join the discussion