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BD Pharmingen™ PerCP-Cy™5.5 Mouse anti-Ki-67

Name: PerCP-Cy™5.5 Mouse anti-Ki-67
Brand: PerCP-Cy™5.5 Mouse anti-Ki-67
SKU: 561284

Clone B56 (RUO)

Two-color flow cytometric analysis of Ki-67 expression by proliferating MOLT-4 cells. Proliferating cells from the human MOLT-4 (T lymphoblastic leukemia, ATCC CRL-1582) cell line was fixed and permeabilized with 70% ice cold ethanol. The cells were washed twice with BD Pharmingen™ Stain Buffer (FBS) (Cat. No. 554656) and stained with either PerCP-Cy5.5 Mouse IgG1, κ Isotype Control (Cat. No. 561824; Left Figure) or PerCP-Cy™5.5 Mouse anti-Ki-67 antibody (Cat. No. 561284; Right Figure) according to the BD Biosciences support protocol, Flow Cytometry Staining Protocol for Detection of Ki-67. The cells were then counterstained with 4',6-Diamidino-2-phenylindole dihydrochloride (DAPI) (Sigma, Cat. No. D-9542) to stain double-stranded DNA. Two-color flow cytometric dot plots showing the correlated expression patterns of DAPI (DNA) staining versus Ki-67 were derived from gated events with the forward and side light-scatter characteristics of intact cells.

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Product Details

Brand: BD Pharmingen™

Alternative Name: MKI67; Antigen identified by monoclonal antibody Ki-67; KIA

Reactivity: Human (QC Testing), Mouse (Tested in Development), Rat, Rhesus (Reported)

Isotype: Mouse IgG1, κ

Immunogen: Human Ki-67

Application: Intracellular staining (flow cytometry) (Routinely Tested)

Vol. Per Test: 5 µl

RRID: AB_10611574

Storage Buffer: Aqueous buffered solution containing BSA and ≤0.09% sodium azide.

Regulatory Status: 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.

Recommended Assay Procedures

BD® CompBeads can be used as surrogates to assess fluorescence spillover (compensation). When fluorochrome conjugated antibodies are bound to BD® CompBeads, they have spectral properties very similar to cells. However, for some fluorochromes there can be small differences in spectral emissions compared to cells, resulting in spillover values that differ when compared to biological controls. It is strongly recommended that when using a reagent for the first time, users compare the spillover on cell and BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.

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).
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.
Source of all serum proteins is from USDA inspected abattoirs located in the United States.
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.
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.
Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
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.
Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
Cy is a trademark of Global Life Sciences Solutions Germany GmbH or an affiliate doing business as Cytiva.

Companion Products

Stain Buffer (FBS) RUO

Size 500 ML Cat No. 554656

561284 Rev. 4

Antibody Details

B56

The B56 monoclonal antibody specifically binds to the Ki-67 antigen that is expressed in the nucleus of cycling cells (G1, S, G2, M cell cycle phases). During the G0 phase, the antigen cannot be detected. During interphase of the cell cycle, it is associated with nucleolar components, and it is on the surface of the chromosomes during M phase. Ki-67 is a large protein having 2 alternatively spliced isoforms, an N-terminal forkhead-associated domain, a C-terminal domain that binds to heterochromatin proteins, and multiple phosphorylation sites, the functions of which are still unclear. Because of the strict association of Ki-67 expression with cell proliferation, anti-Ki-67 antibodies are useful for the identification, quantification, and monitoring of growing cell populations.

561284 Rev. 4

Format Details

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.

Format: PerCP-Cy5.5

Excitation Max: 482 nm

Emission Max: 676 nm

561284 Rev.4

Citations & References

View product citations for antibody "561284" on CiteAb

Development References (11)

Bruno S, Crissman HA, Bauer KD, Darzynkiewicz Z. Changes in cell nuclei during S phase: progressive chromatin condensation and altered expression of the proliferation-associated nuclear proteins Ki-67, cyclin (PCNA), p105, and p34. Exp Cell Res. 1991; 196(1):99-106. (Biology: Flow cytometry). View Reference
Bruno S, Darzynkiewicz Z. Cell cycle dependent expression and stability of the nuclear protein detected by Ki-67 antibody in HL-60 cells. Cell Prolif. 1992; 25(1):31-40. (Biology: Flow cytometry). View Reference
Byeon IJ, Li H, Song H, Gronenborn AM, Tsai MD. Sequential phosphorylation and multisite interactions characterize specific target recognition by the FHA domain of Ki67.. Nat Struct Mol Biol. 2005; 12(11):987-93. (Biology). View Reference
Ho DWY, Fan ST, To J, et al. Selective plasma filtration for treatment of fulminant hepatic failure induced by D-galactosamine in a pig model. Gut. 2002; 50:869-876. (Clone-specific).
Kill IR. Localisation of the Ki-67 antigen within the nucleolus: evidence for a fibrillarin-deficient region of the dense fibrillar component. J Cell Sci. 1996; 109(6):1253-1263. (Biology). View Reference
Kouro T, Medina KL, Oritani K, Kincade PW. Characteristics of early murine B-lymphocyte precursors and their direct sensitivity to negative regulators. Blood. 2001; 97(9):2708-2715. (Clone-specific: Flow cytometry). View Reference
Pitcher CJ, Hagen SI, Walker JM, et al. Development and homeostasis of T cell memory in rhesus macaque. J Immunol. 2002; 168(1):29-43. (Clone-specific: Flow cytometry). View Reference
Scholzen T, Endl E, Wohlenberg C, et al. The Ki-67 protein interacts with members of the heterochromatin protein 1 (HP1) family: a potential role in the regulation of higher-order chromatin structure.. J Pathol. 2002; 196(2):135-44. (Biology). View Reference
Scholzen T, Gerdes J. The Ki-67 protein: from the known and the unknown.. J Cell Physiol. 2000; 182(3):311-22. (Biology). View Reference
Spargo LDJ, Cleland LG, Cockshell MP, Mayrhofer Graham. Recruitment and proliferation of CD4+ T cells in synovium following adoptive transfer of adjuvant-induced arthritis. Int Immunol. 2006; 18(6):897-910. (Clone-specific: Flow cytometry, Immunofluorescence).
Starborg M, Gell K, Brundell E, Höög C. The murine Ki-67 cell proliferation antigen accumulates in the nucleolar and heterochromatic regions of interphase cells and at the periphery of the mitotic chromosomes in a process essential for cell cycle progression. J Cell Sci. 1996; 109(1):143-153. (Biology). View Reference

View All (11)

561284 Rev. 4

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.