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Flow cytometric analysis of perforin expression in human peripheral blood mononuclear cells. Human peripheral blood mononuclear cells were fixed and permeabilized with BD Cytofix/Cytoperm™ Fixation and Permeabilization Solution (Cat. No. 554722). The cells were then washed with and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with either BD Horizon™ PE-CF594 Mouse IgG2b, κ Isotype Control (Cat. No. 562305; dashed line histogram) or BD Horizon™ PE-CF594 Mouse Anti-Human Perforin antibody (Cat. No. 563763; solid line histogram). The fluorescence histograms were derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.

hPerforin_PE-CF594_Overlay_new.png Orange-Cap-Blue-Label.jpg

Flow cytometric analysis of perforin expression in human peripheral blood mononuclear cells. Human peripheral blood mononuclear cells were fixed and permeabilized with BD Cytofix/Cytoperm™ Fixation and Permeabilization Solution (Cat. No. 554722). The cells were then washed with and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with either BD Horizon™ PE-CF594 Mouse IgG2b, κ Isotype Control (Cat. No. 562305; dashed line histogram) or BD Horizon™ PE-CF594 Mouse Anti-Human Perforin antibody (Cat. No. 563763; solid line histogram). The fluorescence histograms were derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.

Product Details
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BD Horizon™
PRF1; P1; PERF; PFN1; PFP; Perforin-1; Cytolysin; FLH2; HPLH2
Human (QC Testing), Cow (Reported)
Mouse BALB/c IgG2b, κ
Purified Granules from the Human Lymphoma Cell Line YT
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl
AB_2738410
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 BD Horizon™ PE-CF594 under optimum conditions, and unconjugated antibody and free PE-CF594 were removed.
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Product Notices

  1. 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).
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. 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.
  4. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
  5. 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.
  6. Because of the broad absorption spectrum of the tandem fluorochrome, extra care must be taken when using multi-laser cytometers, which may directly excite both PE and CF™594.
  7. When excited by the yellow-green (561-nm) laser, the fluorescence may be brighter than when excited by the blue (488-nm) laser.
  8. Texas Red is a registered trademark of Molecular Probes, Inc., Eugene, OR.
  9. CF™ is a trademark of Biotium, Inc.
  10. This product is provided under an Agreement between BIOTIUM and BD Biosciences. The manufacture, use, sale, offer for sale, or import of this product is subject to one or more patents or pending applications owned or licensed by Biotium, Inc. This product, and only in the amount purchased by buyer, may be used solely for buyer’s own internal research, in a manner consistent with the accompanying product literature. No other right to use, sell or otherwise transfer (a) this product, or (b) its components is hereby granted expressly, by implication or by estoppel. This product is for research use only. Diagnostic uses require a separate license from Biotium, Inc. For information on purchasing a license to this product including for purposes other than research, contact Biotium, Inc., 3159 Corporate Place, Hayward, CA 94545, Tel: (510) 265-1027. Fax: (510) 265-1352. Email: btinfo@biotium.com.
  11. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  12. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
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563763 Rev. 1
Antibody Details
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δG9

Perforin has a key role in cell-mediated cytotoxicity. It is a 70 kDa cytolytic protein that is expressed in the cytoplasmic granules of cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells. CTLs are involved in eliminating virally infected cells, in anti-tumor immune responses, in allograft rejections, and in some autoimmune diseases. NK cells are important for tumor surveillance and destruction and are involved in allograft rejections. Cytotoxic cells release the contents of their cytotoxic granules, including perforin upon recognition of their target cell. In the presence of calcium, perforin forms transmembrane channels or pores in the membrane of the target cell leading to a cell death that resembles apoptosis. The ability to detect perforin-positive cells with specific antibody should be useful in identifying and understanding perforin-mediated reactions.

Clone δG9 reacts with human and bovine perforin. It does not cross-react with mouse perforin. Purified granules from the human lymphoma cell line YT were used as immunogen. Clone δG9 was initially characterized by immunoprecipitation and immunohistochemistry of frozen tissue sections. The antibody stains scattered lymphocytes in red pulp of spleen, and scattered infiltrated lymphocytes in lymphoma.

This antibody is conjugated to BD Horizon™ PE-CF594, which has been developed exclusively by BD Biosciences as a better alternative to PE-Texas Red®. PE-CF594 excites and emits at similar wavelengths to PE-Texas Red® yet exhibits improved brightness and spectral characteristics. Due to PE having maximal absorption peaks at 496 nm and 564 nm, PE-CF594 can be excited by the blue (488-nm), green (532-nm) and yellow-green (561-nm) lasers and can be detected with the same filter set as PE-Texas Red® (eg 610/20-nm filter).

563763 Rev. 1
Format Details
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PE-CF594
BD Horizon™ PE-CF594 dye is a part of the BD PE family of dyes. This tandem fluorochrome is comprised of a R-Phycoerythrin (PE) donor that has excitation maxima (Ex Max) of 496-nm and 566-nm and an acceptor dye with an emission maximum (Em Max) at 615-nm. PE-CF594, driven by BD innovation, is designed to be excited by the blue (488-nm), Green (532-nm) and Yellow-Green (561-nm) lasers and detected using an optical filter centered near 615 nm (e.g., a 610/20-nm bandpass filter). The donor dye can be excited by the Blue (488-nm), Green (532-nm) and Yellow-Green (561-nm) lasers and the acceptor dye can be excited by the Green (532-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.
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PE-CF594
496 nm, 566 nm
615 nm
563763 Rev.1
Citations & References
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View product citations for antibody "563763" on CiteAb

Development References (7)

  1. Endsley JJ, Furrer JL, Endsley MA, et al. Characterization of bovine homologues of granulysin and NK-lysin. J Immunol. 2004; 173(4):2607-2614. (Clone-specific: Flow cytometry, Western blot). View Reference
  2. Fox WM 3rd, Hameed A, Hutchins GM, et al. Perforin expression localizing cytotoxic lymphocytes in the intimas of coronary arteries with transplant-related accelerated arteriosclerosis. Hum Pathol. 1993; 24(5):477-482. (Clone-specific: Immunohistochemistry). View Reference
  3. Hameed A, Fox WM, Kurman RJ, Hruban RH, Podack ER. Perforin expression in endometrium during the menstrual cycle. Int J Gynecol Pathol. 1995; 14(2):143-150. (Clone-specific: Flow cytometry). View Reference
  4. Hameed A, Fox WM, Kurman RJ, Hruban RH, Podack ER. Perforin expression in human cell-mediated luteolysis. Int J Gynecol Pathol. 1995; 14(2):151-157. (Clone-specific: Immunohistochemistry). View Reference
  5. Hameed A, Olsen KJ, Cheng L, Fox WM 3rd, Hruban RH, Podack ER. Immunohistochemical identification of cytotoxic lymphocytes using human perforin monoclonal antibody. Am J Pathol. 1992; 140(5):1025-1030. (Immunogen: Immunohistochemistry, Immunoprecipitation). View Reference
  6. Hameed A, Podack ER, Fox WM, Schafer RW, Sherman ME. Detection of perforin in human peritoneal fluid T-lymphocytes. Acta Cytol. 1996; 40(3):401-407. (Clone-specific: Immunohistochemistry). View Reference
  7. Rukavina D, Balen-Marunic S, Rubesa G, Orlic P, Vujaklija K, Podack ER. Perforin expression in peripheral blood lymphocytes in rejecting and tolerant kidney transplant recipients. Transplantation. 1996; 61(2):285-291. (Clone-specific: Flow cytometry). View Reference
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563763 Rev. 1

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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.