You are now leaving the BD Biosciences website. The site you are about to visit is operated by a third party. The link to this site neither makes nor implies any representation or warranty for any products or services offered on a third-party site and is intended only to enable convenient access to the third-party site and for no other purpose. Do you want to continue?
For the best web browsing experience, please use Chrome, Safari or Firefox, minimum versions 77.0.3865, 12.1.2 and 68, respectively.
Receptor-Mediated Apoptosis in Mouse Cells Using anti-Fas/CD95 (Clone Jo2)
- A cell line or primary cells that can be induced to undergo apoptosis by mouse Fas mAb. We typically use thymocytes isolated from a 4-6 week old BALB/c mouse.
Note: Not all cell types that express Fas can be induced to undergo apoptosis using this protocol.
- Anti-mouse Fas mAb, Clone Jo2 (Cat. No. 554254).
- Recombinant Protein G (SIGMA, Cat. No. P4689). The addition of Protein G to the tissue culture medium can enhance the efficiency of Jo2 mAb to induce apoptosis in some experimental systems.
- Tissue culture flasks or plates.
- RPMI-1640 medium supplemented with 10% heat-inactivated fetal bovine serum (FBS), 1% L-glutamine and 1% antibiotics (penicillin/streptomycin; 100 U/ml). This supplemented medium is referred to as 'medium' below.
- Isolate BALB/c thymocytes from the thymus of a 4-6 week old mouse.
- Treat cell suspension (2 x 10 6-1 x 10 7 thymocytes/ml) with 2-20 µg/ml Jo2 mAb. Negative controls should consist of cells cultured in medium without the addition of anti-Fas and medium containing a hamster isotype control (clone Ha4/8, Cat. No. 553961) at the same concentration as the anti-Fas mAb. If Protein G (1-2 µg/ml) is added along with Jo2, then add controls containing (1) Protein G alone and (2) Protein G plus the Ha4/8 mAb.
- Perform a time course to obtain optimum results. Incubation time between 2 to 12 hr at 37°C is suggested.
- Proceed with assays designed to evaluate induction of apoptosis.
APO-BRDU and APO-DIRECT are trademarks of Phoenix Flow Systems.