Upon initial thawing, recombinant mouse TNF (Cat. No. 554589) should be aliquoted into polypropylene microtubes and frozen at -80°C for future use. Alternatively, the product can be diluted in sterile neutral buffer containing not less than 0.5 - 10 mg/mL carrier protein, such as human or bovine serum albumin, aliquoted and stored at -80°C. For in vitro biological assay use, carrier protein concentrations of ≥ 1 mg/mL are recommended. For use as an ELISA standard, carrier protein concentrations of 5 - 10 mg/mL are recommended. Recombinant mouse TNF (Cat. No. 554589) should not be diluted to less than 50 μg/mL for long term storage. Failure to add carrier protein or store at indicated temperatures may result in a loss of activity. Carrier proteins may have an undesired influence on experimental results due to toxicity, high endotoxin levels or possible blocking activity.
ELISA Standard: Recombinant mouse TNF (Cat. No. 554589) can be useful as a quantitative standard for measuring mouse TNF protein levels using sandwich ELISA with purified G281-2626 antibody (Cat. No. 551225) as a capture antibody and biotinylated MP6-XT3 (Cat. No. 554415) as the detection antibody. To obtain linear standard curves, investigators may want to consider using doubling dilutions of recombinant mouse TNF from 2,000 - 15 pg/mL to be included in each ELISA plate. For measuring mouse TNF in serum or plasma, investigators are highly encouraged to use the BD OptEIA™ Mouse TNF ELISA Set (Cat. No. 555268 or 558534) or BD OptEIA™ Mouse TNF ELISA Kit (Cat. No. 560478).
Bioassay: Investigators are advised that the Bioassay application is not routinely tested for this material and are highly encouraged to both titrate this material and include appropriate controls in relevant experiments. An activity range of 0.2 - 5.0 x 10^9 units/mg, encompassing an
ED50= 2 - 50 pg/mL, has previously been reported using L929 indicator cells for proliferation, with a unit defined as the amount of material needed to stimulate a half-maximal response at cytokine saturation.
Blocking: Recombinant mouse TNF (Cat. No. 554589) can be useful as a blocking control for flow cytometric analysis when used with a flurochrome conjugated-MP6-XT22 antibody (such as PE conjugated MP6-XT22 ; Cat. No. 554419). Investigators are advised that the blocking application is not routinely tested for this material. Intracellular cytokine staining techniques and the use of blocking controls are described in detail by C. Prussin and D. Metcalfe.