FITC Mouse Anti-Rat IFN-γ
Clone DB-1 (RUO)
- Brand BD Pharmingen™
- Vol. Per Test 20 µl
- Isotype Mouse IgG1, κ
- Reactivity Rat (QC Testing)
Intracellular staining (flow cytometry) (Routinely Tested)
- Immunogen Recombinant Rat IFN-γ
- Storage Buffer Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
- Regulatory Status RUO
Regulatory Status Legend
The DB-1 monoclonal antibody specifically binds to rat interferon-γ (IFN-γ). The immunogen used to generate the DB-1 hybridoma was recombinant rat IFN-γ expressed in COS cells. This is a neutralizing antibody.
FITC, fluorescein isothiocyanate, is a fluorochrome with a molecular weight of 389 Da. FITC is sensitive to pH changes and photobleaching. Due to nearly identical excitation and emission properties but different spillover characteristics, FITC and Alexa Fluor® 488 cannot be used simultaneously. FITC is relatively dim and should be reserved for highly expressed markers whenever possible.
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Preparation and Storage
The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.
The antibody was conjugated with FITC under optimum conditions, and unreacted FITC was removed.
Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.
- This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- Please refer to www.bdbiosciences.com/pharmingen/protocols for technical protocols.
- Source of all serum proteins is from USDA inspected abattoirs located in the United States.
- Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
Immunofluorescent Staining and Flow Cytometric Analysis: The DB-1 antibody is useful for immunofluorescent staining and flow cytometric analysis to identify and enumerate IFN-γ producing cells within mixed cell populations. This 100 Test Size formulation of the FITC-conjugated DB-1 antibody has been pre-titrated to assure effective intracellular detection of rat IFN-γ using 20 µl/1 x 10^6 cells in a final volume of 100 µl. For specific methodology, please visit our web site, www.bdbiosciences.com, and go to the protocols section or the chapter on intracellular staining in the Immune Function Handbook. A useful control for demonstrating specificity of staining is to pre-block the fixed/permeabilized cells with unlabeled DB-1 antibody prior to staining. The intracellular staining technique and the use of blocking controls have been described in detail by C. Prussin and D. Metcalfe. A suitable mouse IgG1 isotype control for assessing the level of background staining on paraformaldehyde-fixed/saponinpermeabilized rat cells is also available: FITC-MOPC-21 (Cat. No. 554679).
Important Note: This pre-titered antibody solution does not contain a cell permeabilization agent. It is necessary to include a cell permeabilization agent when using the pre-titered antibody solution to stain fixed and permeabilized cells. Perm/Wash™ Buffer (Cat. No. 554723) contains the permeabiliization agent saponin and is useful for this purpose as described below.
Resuspend one million fixed and permeabilized cells in 20 µl of the pre-titered antibody solution and 30 µl of 1X Perm/Wash™ Buffer (Cat. No. 554723). Incubate the cell suspension for 15 minutes (at RT or 4°C). Wash twice in 100 µl of 1X Perm/Wash™ Buffer (Cat. No. 554723).