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Safety Data Sheet (SDS) Brilliant Violet 605 Anti-mouse Ki-67 Antibody     Product Data Sheet (PDF)    
Brilliant Violet 605™ Anti-mouse Ki-67 Antibody
3862065 50 µg $260.00       
Clone: 16A8
Isotype: Rat IgG2a, κ
Reactivity: Mouse, Human
Immunogen: E. coli expressed partial mouse Ki-67 recombinant protein, 1816-2163 aa.
Formulation: Phosphate-buffered solution, pH 7.2, containing 0.09% sodium azide and BSA (origin USA).
Preparation: The antibody was purified by affinity chromatography and conjugated with Brilliant Violet 605™ under optimal conditions. The solution is free of unconjugated Brilliant Violet 605™ and unconjugated antibody.
Concentration: 0.2 mg/ml
Storage & Handling: The antibody solution should be stored undiluted between 2°C and 8°C, and protected from prolonged exposure to light. Do not freeze.
Application:

ICFC - Quality tested
IF - Reported in the literature

Application Notes:

Ki-67 Staining Protocol:

1. Prepare 70% ethanol and chill at -20°C.
2. Prepare target cells of interest and wash 2X with PBS by centrifuge at 350xg for 5 minutes.
3. Discard supernatant and loosen the cell pellet by vortexing.
4. Add 3 ml cold 70% ethanol drop by drop to the cell pellet while vortexing.
5. Continue vortexing for 30 seconds and then incubate at -20°C for 1 hour.
6. Wash 3X with Cell Staining Buffer and then resuspend the cells at the concentration of 0.5-10 x 106/ml.
7. Mix 100 µl cell suspension with proper fluorochrome-conjugated Ki-67 antibody and incubate at room temperature in the dark for 30 minutes.
8. Wash 2X with Cell Staining Buffer and then resuspend in 0.5 ml cell staining buffer for flow cytometric analysis.

Recommended Usage:

Each lot of this antibody is quality control tested by our Ki-67 staining protocol below. For flow cytometric staining, the suggested use of this reagent is ≤0.25 µg per million cells in 100 µl volume. It is recommended that the reagent be titrated for optimal performance for each application.

Brilliant Violet 605™ excites at 405 nm and emits at 603 nm. The bandpass filter 610/20 nm is recommended for detection, although filter optimization may be required depending on other fluorophores used. Be sure to verify that your cytometer configuration and software setup are appropriate for detecting this channel. Refer to your instrument manual or manufacturer for support. Brilliant Violet 605™ is a trademark of Sirigen Group Ltd.

This product is subject to proprietary rights of Sirigen Inc. and is made and sold under license from Sirigen Inc. The purchase of this product conveys to the buyer a non-transferable right to use the purchased product for research purposes only. This product may not be resold or incorporated in any manner into another product for resale. Any use for therapeutics or diagnostics is strictly prohibited. This product is covered by U.S. Patent(s), pending patent applications and foreign equivalents.

Application References:

1. Medina-Reyes EI, et al. 2015. Environ Res. 136:424. PubMed
2. Guillaumond F, et al. 2015. PNAS. 112:2473. PubMed
3. Sharma SK, et al. 2015. J Immunol. 194:5529. PubMed
4. Rodero MP, et al. 2014. J. Invest. Dermatol. 7:1991-7. PubMed

Con A+IL-2-stimulated (3 days) C57BL/6

Con A+IL-2-stimulated (3 days) C57BL/6 mouse splenocytes were fixed and permeabilized with 70% ethanol, and then stained with Ki-67 (clone 16A8) Brilliant Violet 605™ (filled histogram) or rat IgG2a, κ Brilliant Violet 605™ isotype control (open histogram).



Description:

The nuclear protein Ki-67 was first identified by the monoclonal antibody Ki-67, which was generated by immunizing mice with nuclei of the L428 Hodgkin lymphoma cell line. Ki-67 protein plays an essential role in ribosomal RNA transcription and cell proliferation. Expression of Ki-67 occurs during G1, S, G2, and M phase, while in G0 phase the Ki-67 protein is not detectable. Ki-67 is strongly expressed in proliferating cells and has been reported as a prognostic marker in various tumors.

Other Names: KIA, proliferation-related Ki-67 antigen
Structure: 325 kD protein containing a forkhead-associated domain (FHA) and 13 tandem repeats
Distribution: Nucleus and chromosome
Function: Required for cell cycle progression and proliferation
Interaction: Interacts with KIF15; binds to MKI67IP through FHA domain
Antigen References:

1. Starborg M, et al. 1996. J. Cell. Sci. 109:143.
2. Byeon IJ, et al. 2005. Nat. Struct. Mol. Biol. 12:987.
3. Yerushalmi R, et al. 2010. Lancet. Oncol. 11:174.
4. Beltrami AP, et al. 2001. N. Engl. J. Med. 344:1750.
5. Sachsenberg N, et al. 1998. J. Exp. Med. 187:1295.
6. Nagy Z, et al. 1997. Acta. Neuropathol. 93:294.