Flow cytometric analysis of HLA-ABC expression on human peripheral blood lymphocytes. Whole blood was stained with either PE-Cy™5 Mouse Anti-Human HLA-ABC (Cat. No. 555554; solid line histogram) or PE-Cy™5 Mouse IgG1 κ Isotype Control (Cat. No. 555750; dashed line histogram). Erythrocytes were lysed with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899). Fluorescence histograms depicting HLA-ABC (or Ig isotype control) expression were derived from gated events with the forward and side light-scatter characteristics of viable lymphocytes. Flow cytometry was performed on a BD FACScan™ system.
Flow cytometric analysis of HLA-ABC expression on human peripheral blood lymphocytes. Whole blood was stained with either PE-Cy™5 Mouse Anti-Human HLA-ABC (Cat. No. 555554; solid line histogram) or PE-Cy™5 Mouse IgG1 κ Isotype Control (Cat. No. 555750; dashed line histogram). Erythrocytes were lysed with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899). Fluorescence histograms depicting HLA-ABC (or Ig isotype control) expression were derived from gated events with the forward and side light-scatter characteristics of viable lymphocytes. Flow cytometry was performed on a BD FACScan™ system.
Product Details
BD Pharmingen™
Major histocompatibility complex, class I, A,B,C; HLA class I A,B,C
Human (QC Testing), Rhesus, Cynomolgus, Baboon (Tested in Development)
Mouse IgG1, κ
Flow cytometry (Routinely Tested)
20 µl
AB_395937
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 PE-Cy5 (formerly known as BD Cy-Chrome™) under optimum conditions, and unconjugated antibody and free PE-Cy5 were removed.
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.
- 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.
- PE-Cy5 is a tandem fluorochrome composed of R-phycoerythrin (PE), which is excited by the 488 nm light of an Argon ion laser and serves as an energy donor, coupled to the cyanine dye Cy5, which acts as an energy acceptor and fluoresces at 670 nm. BD Biosciences Pharmingen has maximized the fluorochrome energy transfer in PE-Cy5, thus maximizing its fluorescence emission intensity, minimizing residual emission from PE, and minimizing lot-to-lot variation.
- PE-Cy5 is optimized for use with a single argon ion laser emitting 488-nm light. Because of the broad absorption spectrum of the PE-Cy5 tandem fluorochrome, extra care must be taken when using dual-laser cytometers which may directly excite both PE and Cy5™.
- 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.
- PE-Cy5 tandem fluorochromes have been reported to bind some classes of human macrophages and granulocytes via Fc receptors, and PE has been reported to bind to mouse B lymphocytes via Fc receptors. Preincubation of mouse leukocytes with Mouse BD Fc Block™ purified anti-mouse CD16/CD32 mAb 2.4G2 can reduce the non-specific binding of PE-Cy5-conjugated reagents to mouse B cells. However, PE-Cy5 conjugated reagents should not be used to stain splenocytes of SJL, NOD, and MRL mice as B lymphocytes and/or other leukocytes have been reported to non-specifically stain regardless of the use of Mouse BD Fc Block™ (the CD72c complex has been implicated for PE-Cy5 binding in these strains). Reagents conjugated to PE, PerCP, PerCP-Cy5.5, APC, and APC-Cy7 tandem fluorochrome can be used on leukocytes from these mouse strains.
- Cy is a trademark of GE Healthcare.
- Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
Companion Products
Stain Buffer (FBS) RUO
Size
500 mL
Cat No.
554656
Stain Buffer (BSA) RUO
Size
500 mL
Cat No.
554657
Lysing Solution 10X Concentrate CE_IVD
Size
100 mL
Cat No.
349202
Lysing Buffer RUO
Size
100 mL
Cat No.
555899
PE-Cy™5 Mouse IgG1 κ Isotype Control RUO
Size
100 Tests
Cat No.
555750
555554 Rev. 10
Antibody Details
G46-2.6
The Human Leukocyte Antigen (HLA) complex is the human version of the MHC, helping the immune system distinguish the body's own proteins versus those from foreign invaders, such as viruses. Humans have three main MHC class I genes, known as HLA-A, HLA-B and HLA-C. Major histocompatibility complex (MHC) class I molecules, which are widely found on the surface of nucleated cells, function by binding peptides and displaying them on the cell surface to cytotoxic T-cells. Intracellular degradation of cytosolic proteins by the proteasome generates many of the peptides that load MHC class I molecules. MHC class I may also serve as an inhibitory ligand for natural killer (NK) cell receptors (KIR, Killer Immunoglobulin-like Receptors), which viruses may modulate expression levels for to evade immune detection. The G46-2.6 monoclonal antibody binds to a monomorphic epitope on the alpha chain of HLA-A, HLA-B and HLA-C.
555554 Rev. 10
Format Details
PE-Cy5
PE-Cy5 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, Cy™5, with an emission maximum (Em Max) at 670-nm. PE 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 670-nm (e.g., a 670/20-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 Red (627-640-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.
PE-Cy5
Yellow-Green 488 nm, 532 nm, 561 nm
496 nm, 566 nm
670 nm
555554 Rev.10
Citations & References
Development References (5)
-
Barclay NA, Brown MH, Birkeland ML, et al, ed. The Leukocyte Antigen FactsBook. San Diego, CA: Academic Press; 1997.
-
Crisa L, Cirulli V, Ellisman MH, Ishii JK, Elices MJ, Salomon DR. Cell adhesion and migration are regulated at distinct stages of thymic T cell development: the roles of fibronectin, VLA4, and VLA5. J Exp Med. 1996; 184(1):215-228. (Clone-specific: Flow cytometry). View Reference
-
Kap YS, van Meurs M, van Driel N, et al. A monoclonal antibody selection for immunohistochemical examination of lymphoid tissues from non-human primates. J Histochem Cytochem. 2009; 57(12):1159-1167. (Clone-specific: Immunohistochemistry). View Reference
-
Koppelman B, Neefjes JJ, de Vries JE, de Waal Malefyt R. Interleukin-10 down-regulates MHC class II alphabeta peptide complexes at the plasma membrane of monocytes by affecting arrival and recycling. Immunity. 1997; (6):861-871. (Clone-specific: Flow cytometry). View Reference
-
Xia H, Liu H, Zhang G, Zheng Y. Phenotype and function of monocyte-derived dendritic cells from chinese rhesus macaques. Cell Mol Immunol. 2009; 6(3):159-165. (Clone-specific: Flow cytometry). View Reference
555554 Rev. 10
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.
