Revision 4

#61702Store at -20C

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IP, ChIP, ChIP-seq, C&R

REACTIVITY:

H

SENSITIVITY:

Endogenous

MW (kDa):

300

Source/Isotype:

Rabbit IgG

UniProt ID:

#O15047

Entrez-Gene Id:

9739

Product Information

Product Usage Information

For optimal ChIP and ChIP-seq results, use 5 μL of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP. This antibody has been validated using SimpleChIP® Enzymatic Chromatin IP Kits.

The CUT&RUN dilution was determined using CUT&RUN Assay Kit #86652.
Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:200
Chromatin IP 1:100
Chromatin IP-seq 1:100
CUT&RUN 1:100

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Specificity / Sensitivity

SET1A (D3V9S) Rabbit mAb recognizes endogenous levels of total SET1A protein. This antibody does not cross-react with other SET1/COMPASS proteins, including SET1B, MLL1, MLL2, MLL3, and MLL4.

Species Reactivity:

Human

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Pro383 of human SET1A protein.

Background

The Set1 histone methyltransferase protein was first identified in yeast as part of the Set1/COMPASS histone methyltransferase complex, which methylates histone H3 at Lys4 and functions as a transcriptional co-activator (1). While yeast contain only one known Set1 protein, six Set1-related proteins exist in mammals: SET1A, SET1B, MLL1, MLL2, MLL3, and MLL4, all of which assemble into COMPASS-like complexes and methylate histone H3 at Lys4 (2,3). These Set1-related proteins are each found in distinct protein complexes, all of which share the common subunits WDR5, RBBP5, ASH2L, CXXC1 and DPY30. These subunits are required for proper complex assembly and modulation of histone methyltransferase activity (2-6). MLL1 and MLL2 complexes contain the additional protein subunit, menin (6). Like yeast Set1, all six Set1-related mammalian proteins methylate histone H3 at Lys4 (2-6). MLL translocations are found in a large number of hematological malignancies, suggesting that Set1/COMPASS histone methyltransferase complexes play a critical role in leukemogenesis (6).

  1. Miller, T. et al. (2001) Proc Natl Acad Sci USA 98, 12902-7.
  2. Shilatifard, A. (2008) Curr Opin Cell Biol 20, 341-8.
  3. Tenney, K. and Shilatifard, A. (2005) J Cell Biochem 95, 429-36.
  4. Lee, J.H. and Skalnik, D.G. (2005) J Biol Chem 280, 41725-31.
  5. Lee, J.H. et al. (2007) J Biol Chem 282, 13419-28.
  6. Hughes, C.M. et al. (2004) Mol Cell 13, 587-97.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IP: Immunoprecipitation ChIP: Chromatin IP ChIP-seq: Chromatin IP-seq C&R: CUT&RUN

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
SimpleChIP is a registered trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Revision 4
#61702

SET1A (D3V9S) Rabbit mAb

Western Blotting Image 1: SET1A (D3V9S) Rabbit mAb Expand Image
Western blot analysis of extracts from 293T and HeLa cells using SET1A (D3V9S) Rabbit mAb.
Immunoprecipitation Image 1: SET1A (D3V9S) Rabbit mAb Expand Image
Immunoprecipitation of SET1A from 293T cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is SET1A (D3V9S) Rabbit mAb. Western blot analysis was performed using SET1A (D3V9S) Rabbit mAb.
Chromatin Immunoprecipitation Image 1: SET1A (D3V9S) Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from HeLa cells and SET1A (D3V9S) Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using SimpleChIP® ChIP-seq DNA Library Prep Kit for Illumina® #56795. The figure shows binding across GAPDH, a known target gene of SET1A (see additional figure containing ChIP-qPCR data). For additional ChIP-seq tracks, please download the product datasheet.
Chromatin Immunoprecipitation Image 2: SET1A (D3V9S) Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from HeLa cells and SET1A (D3V9S) Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using SimpleChIP® ChIP-seq DNA Library Prep Kit for Illumina® #56795. The figure shows binding across chromosome 12 (upper), including GAPDH (lower), a known target gene of SET1A (see additional figure containing ChIP-qPCR data).
Chromatin Immunoprecipitation Image 3: SET1A (D3V9S) Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from HeLa cells and either SET1A (D3V9S) Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by Real-Time PCR using SimpleChIP® Human β-Actin Promoter Primers #13653, SimpleChIP® Human GAPDH Promoter Primers #4471, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
CUT and RUN Image 1: SET1A (D3V9S) Rabbit mAb Expand Image
CUT&RUN was performed with HeLa cells and SET1A (D3V9S) Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA library was prepared using DNA Library Prep Kit for Illumina Systems (ChIP-seq, CUT&RUN) #56795. The figure shows binding across PRKAA1 gene, a known target gene of SET1A (see additional figure containing CUT&RUN-qPCR data).
CUT and RUN Image 2: SET1A (D3V9S) Rabbit mAb Expand Image
CUT&RUN was performed with HeLa cells and SET1A (D3V9S) Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA library was prepared using DNA Library Prep Kit for Illumina Systems (ChIP-seq, CUT&RUN) #56795. The figures show binding across chromosome 5 (upper), including PRKAA1 (lower), a known target gene of SET1A (see additional figure containing CUT&RUN-qPCR data).
CUT and RUN Image 3: SET1A (D3V9S) Rabbit mAb Expand Image
CUT&RUN was performed with HeLa cells and either SET1A (D3V9S) Rabbit mAb or Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT&RUN) #66362, using CUT&RUN Assay Kit #86652. The enriched DNA was quantified by real-time PCR using human PRKAA1 promoter primers, human KANSL1L promoter primers, and human PAD14 promoter primers. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.