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BD Pharmingen? Technical Data Sheet Purified NA/LE Hamster Anti-Mouse/Rat TNF Product Information Material Number:

557370 Size: 0.

5 mg Concentration: 1.0 mg/ml Clone: TN3-19.12 Immunogen: Recombinant mouse TNF protein Isotype: Armenian Hamster IgG1, λ1 QC Testing: Mouse Reactivity: Storage Buffer: No azide/low endotoxin: Aqueous buffered solution containing no preservative, 0.2?m sterile filtered. Endotoxin level is ≤0.01 EU/?g (≤0.001 ng/?g) of protein as determined by the LAL assay. Description The TN3-19.12 antibody reacts with rat and mouse tumor-necrosis factor (TNF) proteins (also known as TNF-α) . Moreover, the TN3-19.12 antibody is reported to crossreact with rabbit TNF, but it does not crossreact with mouse lymphotoxin-α (LT-α, also known as TNF-β) nor with human TNF. The immunogen used to generate the TN3-19.12 hybridoma was E. coli-expressed, purified recombinant mouse TNF protein. This monoclonal antibody has been reported to neutralize the bioactivities of mouse, rat and rabbit TNF. Preparation and Storage Store undiluted at 4°C. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. This preparation contains no preservatives, thus it should be handled under aseptic conditions. Application Notes Application ELISA Routinely Tested Neutralization Tested During Development Immunoprecipitation Reported Western blot Reported Neutralization Activity: This antibody has been reported to be useful for the neutralization application. Neutralization of TNF bioactivity may be measured with a cytolysis assay with MTT using

12 ng/mL recombinant mouse TNF (Cat. No. 554589) with L929 indicator cells at

2 x 10^5 cells/mL (i.e. preincubation of the antibody with recombinant mouse TNF for

60 minutes). 50% Neutralization (ND50) at 0.3 - 2.0 μg/mL ≥ 60% Neutralization at

10 -

100 μg/mL Recommended Assay Procedure: ELISA: Purified Hamster Anti-Mouse/Rat TNF mAb (Cat. No 557516) can be used as a capture antibody for a sandwich ELISA that measures TNF protein levels. For measuring Mouse or Rat TNF in complex biological samples, such as serum or plasma, investigators are highly encouraged to use the BD OptEIA? Mouse TNF ELISA Set II (Cat. No 558534) or the BD OptEIA? Rat TNF ELISA Set (Cat. No. 558535). IP/WB: The TN3-19.12 antibody has been reported to be useful for immunoprecipitation and Western blotting. Investigators should note that these are applications not routinely tested. Suggested Companion Products Catalog Number Name Clone Size

554709 Purified NA/LE Hamster IgG1, λ1 Isotype Control 0.5 mg G235-2356

554589 Recombinant Mouse TNF

10 ?g (none)

558535 Rat TNF ELISA Set

20 plates (none)

558534 Mouse TNF ELISA Set II

20 plates (none)

557516 Purified Hamster Anti-Mouse/Rat TNF 0.5 mg TN3-19.12

555268 Mouse TNF (Mono/Mono) ELISA Set

20 plates (none)

562336 Mouse TNF Enhanced Sensitivity Flex Set

100 tests (none)

557370 Rev.

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2 Product Notices Since applications vary, each investigator should titrate the reagent to obtain optimal results. 1. An isotype control should be used at the same concentration as the antibody of interest. 2. Although hamster immunoglobulin isotypes have not been well defined, BD Biosciences Pharmingen has grouped Armenian and Syrian hamster IgG monoclonal antibodies according to their reactivity with a panel of mouse anti-hamster IgG mAbs. A table of the hamster IgG groups, Reactivity of Mouse Anti-Hamster Ig mAbs, may be viewed at http://www.bdbiosciences.com/documents/hamster_chart_11x17.pdf. 3. Please refer to www.bdbiosciences.com/pharmingen/protocols for technical protocols. 4. References Ferran C, Dautry F, Merite S, et al. Anti-tumor necrosis factor modulates anti-CD3-triggered T cell cytokine gene expression in vivo. J Clin Invest. 1994;