Flow cytometric analysis of Stat4 (pY693) expression in IFN-α treated human peripheral blood lymphocytes. Human whole blood was either stimulated with 40,000 U/ml of IFN-α for 15 minutes at 37ºC (solid line histogram) or was left unstimulated (dashed line histogram). The erythrocytes were lysed and the leukocytes were fixed with 1× BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) for 10-15 minutes at 37ºC. The leukocytes were washed, permeabilized (BD Phosflow™ Perm Buffer III, Cat. No. 558050) on ice for 30 minutes, washed and then stained with PerCP-Cy™5.5 anti-Stat4 (pY693) (Cat. No.561217). The fluorescence histograms were derived from events with the forward and side light-scatter characteristics of lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
Flow cytometric analysis of Stat4 (pY693) expression in IFN-α treated human peripheral blood lymphocytes. Human whole blood was either stimulated with 40,000 U/ml of IFN-α for 15 minutes at 37ºC (solid line histogram) or was left unstimulated (dashed line histogram). The erythrocytes were lysed and the leukocytes were fixed with 1× BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) for 10-15 minutes at 37ºC. The leukocytes were washed, permeabilized (BD Phosflow™ Perm Buffer III, Cat. No. 558050) on ice for 30 minutes, washed and then stained with PerCP-Cy™5.5 anti-Stat4 (pY693) (Cat. No.561217). The fluorescence histograms were derived from events with the forward and side light-scatter characteristics of lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
製品詳細
BD Phosflow™
Signal transducer and activator of transcription 4; SLEB11
Human (QC Testing), Mouse (Reactivity Confirmed in Development)
Mouse IgG2b, κ
Phosphorylated Human Stat4 (pY693)
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl
AB_10643004
Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
RUO
Preparation and Storage
Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated with PerCP-Cy5.5 under optimum conditions, and unconjugated antibody and free PerCP-Cy5.5 were removed. Storage of PerCP-Cy5.5 conjugates in unoptimized diluent is not recommended and may result in loss of signal intensity.
Product Notices
- 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).
- Source of all serum proteins is from USDA inspected abattoirs located in the United States.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- Cy is a trademark of Amersham Biosciences Limited. This conjugated product is sold under license to the following patents: US Patent Nos. 5,486,616; 5,569,587; 5,569,766; 5,627,027.
- Please observe the following precautions: Absorption of visible light can significantly alter the energy transfer occurring in any tandem fluorochrome conjugate; therefore, we recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to prevent exposure of conjugated reagents, including cells stained with those reagents, to room illumination.
- 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.
- PerCP-Cy5.5–labelled antibodies can be used with FITC- and R-PE–labelled reagents in single-laser flow cytometers with no significant spectral overlap of PerCP-Cy5.5, FITC, and R-PE fluorescence.
- PerCP-Cy5.5 is optimized for use with a single argon ion laser emitting 488-nm light. Because of the broad absorption spectrum of the tandem fluorochrome, extra care must be taken when using dual-laser cytometers, which may directly excite both PerCP and Cy5.5™. We recommend the use of cross-beam compensation during data acquisition or software compensation during data analysis.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- This product is subject to proprietary rights of Amersham Biosciences Corp. and Carnegie Mellon University and made and sold under license from Amersham Biosciences Corp. This product is licensed for sale only for research. It is not licensed for any other use. If you require a commercial license to use this product and do not have one return this material, unopened to BD Biosciences, 10975 Torreyana Rd, San Diego, CA 92121 and any money paid for the material will be refunded.
関連製品
Stain Buffer (FBS) RUO
サイズ
500 mL
カタログ番号
554656
Lyse/Fix Buffer 5X RUO
サイズ
250 mL
カタログ番号
558049
Perm Buffer III RUO
サイズ
125 mL
カタログ番号
558050
561217 Rev. 2
抗体の詳細
38/p-Stat4
The Stat proteins function both as cytoplasmic signal transducers and as activators of transcription. Seven mammalian Stat proteins have been identified: Stat1-4, Stat5a, 5b, and Stat6. Stat4 has been shown to play an important role in development of T helper cells, specifically the Th1 subset. Stat4 is activated by IL-12 and by type 1 interferons. Knockout mice supported the role that Stat4 plays in IL-12 signaling because lymphocytes from Stat 4-/- mice could neither differentiate into Th1 cells nor produce IFN-γ in response to treatment with IL-12. IFN-γ plays an important role in host defense. A key component in the activation of Stat4 is the phosphorylation on tyrosine and serine residues in response to IL-12 stimulation. IL-12 stimulation leads to the phosphorylation of Stat4 on tyrosine 693 and serine 721. Transcriptional activity of Stat4 has been shown to be significantly reduced when residues Y693 and S721 are mutated.
561217 Rev. 2
フォーマットの詳細
PerCP-Cy5.5
PerCP-Cy5.5 dye is part of the BD blue family of dyes. This tandem fluorochrome is comprised of a fluorescent protein complex (PerCP) with an excitation maximum (Ex Max) of 482 nm and an acceptor dye with an emission maximum (Em Max) at 676 nm. PerCP-Cy5 is designed to be excited by the blue laser (488-nm) and detected using an optical filter centered near 680 nm (e.g., a 695/40 nm bandpass filter). The donor dye can be partially excited by the Violet (405-nm) laser resulting in cross-laser excitation and fluorescence spillover. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
PerCP-Cy5.5
Blue 488 nm
482 nm
676 nm
561217 Rev.2
引用＆参考文献
Development References (2)
-
Kisseleva T, Bhattacharya S, Braunstein J, Schindler CW. Signaling through the JAK/STAT pathway, recent advances and future challenges. Gene. 2002; 285:1-24. (Biology).
-
Visconti R, Gadina M, Chiariello M, et al. Importance of the MKK6/p38 pathway for interleukin-12−induced STAT4 serine phosphorylation and transcriptional activity. Blood. 2000; 96:1844-1852. (Biology).
561217 Rev. 2
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.
