Multicolor flow cytometric analysis of IFN-γ expression by stimulated pig lymphocytes. Pig peripheral blood mononuclear cells were stimulated for 20 h with Leukocyte Activation Cocktail, with BD GolgiPlug™ (Cat. No. 550583). The cells were fixed using BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized with BD Perm/Wash™ Buffer (Cat. No. 554723). The cells were stained simultaneously with PE Mouse Anti-Pig CD3ε antibody (Cat. No. 561485) and Alexa Fluor® 647 Mouse Anti-Pig IFN-γ antibody (Cat. No. 561480; Left Panel) or an Alexa Fluor® 647 Mouse IgG1 Isotype Control (Cat No. 557732, Right Panel). Two-color flow cytometric dot plots showing the correlated expression of IFN-γ (or Ig Isotype control staining) versus CD3 were derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
Multicolor flow cytometric analysis of IFN-γ expression by stimulated pig lymphocytes. Pig peripheral blood mononuclear cells were stimulated for 20 h with Leukocyte Activation Cocktail, with BD GolgiPlug™ (Cat. No. 550583). The cells were fixed using BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized with BD Perm/Wash™ Buffer (Cat. No. 554723). The cells were stained simultaneously with PE Mouse Anti-Pig CD3ε antibody (Cat. No. 561485) and Alexa Fluor® 647 Mouse Anti-Pig IFN-γ antibody (Cat. No. 561480; Left Panel) or an Alexa Fluor® 647 Mouse IgG1 Isotype Control (Cat No. 557732, Right Panel). Two-color flow cytometric dot plots showing the correlated expression of IFN-γ (or Ig Isotype control staining) versus CD3 were derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
Product Details
BD Pharmingen™
IFNG; IFN-gamma; Interferon gamma; Interferon-gamma
Pig (QC Testing)
Mouse IgG1, κ
Recombinant pig IFN-γ protein
Intracellular staining (flow cytometry) (Routinely Tested)
0.2 mg/ml
AB_10681056
Aqueous buffered solution containing ≤0.09% sodium azide.
RUO
Preparation And Storage
The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated to Alexa Fluor® 647 under optimum conditions, and unreacted Alexa Fluor® 647 was removed. Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.
Product Notices
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
- Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
- The Alexa Fluor®, Pacific Blue™, and Cascade Blue® dye antibody conjugates in this product are sold under license from Molecular Probes, Inc. for research use only, excluding use in combination with microarrays, or as analyte specific reagents. The Alexa Fluor® dyes (except for Alexa Fluor® 430), Pacific Blue™ dye, and Cascade Blue® dye are covered by pending and issued patents.
- 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.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
Companion Products
Fixation Buffer RUO
Size
100 mL
Cat No.
554655
Perm/Wash Buffer RUO
Size
100 mL
Cat No.
554723
Alexa Fluor® 647 Mouse IgG1 κ Isotype Control RUO
Size
100 Tests
Cat No.
557732
Leukocyte Activation Cocktail, with BD GolgiPlug™ RUO
Size
200 µL
Cat No.
550583
PE Mouse Anti-Pig CD3ε RUO
Size
50 µg
Cat No.
561485
561480 Rev. 1
Antibody Details
P2G10
The P2G10 monoclonal antibody specifically binds to porcine interferon-γ (IFN-γ). The immunogen used to generate the P2G10 hybridoma was recombinant pig IFN-γ protein.
561480 Rev. 1
Format Details
Alexa Fluor™ 647
Alexa Fluor™ 647 Dye is part of the BD red family of dyes. This is a small organic fluorochrome with an excitation maximum (Ex Max) at 653-nm and an emission maximum (Em Max) at 669-nm. Alexa Fluor 647 is designed to be excited by the Red laser (627-640 nm) and detected using an optical filter centered near 520-nm (e.g., a 660/20 nm bandpass filter). Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
Alexa Fluor™ 647
Red 627-640 nm
653 nm
669 nm
561480 Rev.1
Citations & References
Development References (2)
-
Mateu de Antonio E, Husmann RJ, Hansen R, et al. Quantitative detection of porcine interferon-gamma in response to mitogen, superantigen and recall viral antigen. Vet Immunol Immunopathol. 1998; 61(2-4):265-277. (Immunogen: ELISA). View Reference
-
Prussin C, Metcalfe DD. Detection of intracytoplasmic cytokine using flow cytometry and directly conjugated anti-cytokine antibodies. J Immunol Methods. 1995; 188(1):117-128. (Methodology: IC/FCM Block). View Reference
561480 Rev. 1
Please refer to Support Documents for Quality Certificates
Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described
Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims. Comparisons are not made against non-BD technologies, unless otherwise noted.
For Research Use Only. Not for use in diagnostic or therapeutic procedures.
