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25501
SET1/COMPASS Antibody Sampler Kit

SET1/COMPASS Antibody Sampler Kit #25501

Western Blotting Image 1

Western blot analysis of extracts from 293T and HeLa cells using SET1A (D3V9S) Rabbit mAb.

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Western Blotting Image 2

Western blot analysis of extracts from 293T, F9, and COS-7 cell lines using SET1B (D1U5D) Rabbit mAb.

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Western Blotting Image 3

Western blot analysis of extracts from various cell lines using MLL1 (D2M7U) Rabbit mAb (Amino-terminal Antigen).

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Western Blotting Image 4

Western blot analysis of extracts from MLL1 wild-type (WT) and knockout (KO) mouse embryonic fibroblasts (MEF) using MLL1 (D6G8N) Rabbit mAb (Carboxy-terminal Antigen) (upper) and α-Actinin (D6F6) XP® Rabbit mAb #6487 (lower). MLL1 WT and KO MEF were kindly provided by Dr. Ali Shilatifard of Northwestern University.

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Western Blotting Image 5

Western blot analysis of extracts from various cell lines using MLL2/KMT2B (D6X2E) Rabbit mAb (Carboxy-terminal Antigen) (upper) and α-Actinin (D6F6) XP® Rabbit mAb #6487 (lower). As expected, HCT 116 cells are negative for MLL2 expression.

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Chromatin IP Image 6

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 NCCIT cells and either 10 μl of WDR5 (D9E1I) Rabbit mAb or 2 μl of Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using human nanog promoter primers, SimpleChIP® Human Oct-4 Promoter Primers #4641, 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.

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Western Blotting Image 7

Western blot analysis of extracts from various cell lines using WDR5 (D9E1I) Rabbit mAb.

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Western Blotting Image 8

Western blot analysis of extracts from HCT 116, NIH/3T3, and COS-7 cell lines using WDR82 (D2I3B) Rabbit mAb.

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Western Blotting Image 9

Western blot analysis of extracts from various cell lines using Menin (D45B1) XP® Rabbit mAb.

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Western Blotting Image 10

After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.

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Chromatin IP Image 11

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 HeLa cells and either 5 μL of SET1A (D3V9S) Rabbit mAb or 2 μl of 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.

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IP Image 12

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.

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IP Image 13

Immunoprecipitation of SET1B from HeLa cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is SET1B (D1U5D) Rabbit mAb. Western blot analysis was performed using SET1B (D1U5D) Rabbit mAb.

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Western Blotting Image 14

Western blot analysis of extracts from various cell lines using MLL1 (D6G8N) Rabbit mAb (Carboxy-terminal Antigen).

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IP Image 15

Immunoprecipitation of MLL2/KMT2B from NCCIT cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is MLL2/KMT2B (D6X2E) Rabbit mAb (Carboxy-terminal Antigen). Western blot analysis was performed using MLL2/KMT2B (D6X2E) Rabbit mAb (Carboxy-terminal Antigen).

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IF-IC Image 16

Confocal immunofluorescent analysis of COS-7 cells using Menin (D45B1) XP® Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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IF-IC Image 17

Confocal immunofluorescent analysis of MEF wild-type (left, positive) and MEF MLL (-/-) (right, negative) cells using MLL (D6G8N) Rabbit mAb (Carboxy-terminal Antigen) (green). Actin filaments were labeled with DyLight™ 554 Phalloidin #13054 (red). MLL1 WT and KO MEF were kindly provided by Dr. Ali Shilatifard of Northwestern University.

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IF-IC Image 18

Confocal immunofluorescent analysis of A549 cells (left, positive) and HCT 116 cells (right, negative) using MLL2/KMT2B (D6X2E) Rabbit mAb (Carboxy-terminal Antigen) (green). Actin filaments were labeled with DyLight™ 554 Phalloidin #13054 (red).

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Product Includes Quantity Applications Reactivity MW(kDa) Isotype
SET1A (D3V9S) Rabbit mAb 61702 20 µl
  • WB
  • IP
  • ChIP
H 300 Rabbit IgG
SET1B (D1U5D) Rabbit mAb 44922 20 µl
  • WB
  • IP
H M Mk 320 Rabbit IgG
MLL1 (D2M7U) Rabbit mAb (Amino-terminal Antigen) 14689 20 µl
  • WB
  • IP
H M R Mk 300 Rabbit IgG
MLL1 (D6G8N) Rabbit mAb (Carboxy-terminal Antigen) 14197 20 µl
  • WB
  • IP
  • IF
H M R Mk 180 Rabbit IgG
MLL2/KMT2B (D6X2E) Rabbit mAb (Carboxy-terminal Antigen) 63735 20 µl
  • WB
  • IP
  • IF
H 80 Rabbit IgG
WDR5 (D9E1I) Rabbit mAb 13105 20 µl
  • WB
  • ChIP
H M R Mk 37 Rabbit IgG
WDR82 (D2I3B) Rabbit mAb 99715 20 µl
  • WB
  • IP
H M Mk 30 Rabbit IgG
Menin (D45B1) XP® Rabbit mAb 6891 20 µl
  • WB
  • IF
H M R Mk 68 Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl
  • WB
Goat 

The SET1/COMPASS Antibody Sampler Kit provides an economical means of detecting SET1/COMPASS proteins using control antibodies against SET1A, SET1B, MLL1, MLL2, WDR5, WDR82, and Menin. This kit contains enough primary antibodies to perform at least two western blot experiments.

Each antibody in the SET1/COMPASS Antibody Sampler Kit detects endogenous levels of its target protein. Cross-reactivity was not observed with other family members.

Monoclonal antibodies are produced by immunizing rabbits with synthetic peptides corresponding to residues surrounding Pro383 of human SET1A, Val204 of human SET1B, Val13 of human WDR82, and Val597 of human Menin proteins, or recombinant protein specific to the amino terminus of human MLL1, carboxy terminus of human MLL1, carboxy terminus of human MLL2, and full-length human WDR5 proteins.

The Set1 histone methyltransferase protein was first identified in yeast as part of the Set1/COMPASS histone methyltransferase complex, which methylates histone H3 on lysine 4 and functions as a transcriptional co-activator (1). While yeast contain only one known Set1 protein, mammals contain six Set1-related proteins: SET1A, SET1B, MLL1, MLL2, MLL3 and MLL4, all of which methylate histone H3 on lysine 4 (2,3). These Set1-related proteins are each found in distinct protein complexes, all of which share the common core structural subunits WDR5, RBBP5 and ASH2L (2-6). WDR82 is a core subunit specific to SET1A and SET1B complexes, while Menin is a core subunit specific to the MLL complexes (4,5,7).

Like yeast Set1, all six Set1-related mammalian proteins methylate histone H3 on lysine 4 (2-6). SET1A, SET1B, MLL1 and MLL2 mediate di- and tri-methylation of histone H3 Lys4 at gene promoters to facilitate transcription activation. MLL3 and MLL4 function primarily to mono-methylate histone H3 Lys4 at gene enhancers. MLL1 and MLL2 function as master regulators of both embryogenesis and hematopoiesis, and are required for proper expression of Hox genes (8-10). MLL1 is a large approximately 4000 amino acid protein that is cleaved by the Taspase 1 threonine endopeptidase to form N-terminal (MLL1-N) and C-terminal MLL1 (MLL1-C) fragments, both of which are subunits of the functional MLL1/COMPASS complex (11,12). MLL1 translocations are found in a large number of hematological malignancies, suggesting that Set1 histone methyltransferase complexes play a critical role in leukemogenesis (6). Like MLL1, MLL2 is also a large, approximately 2700 amino acid protein that is cleaved by the Taspase 1 threonine endopeptidase to form N-terminal (MLL2-N) and C-terminal (MLL2-C) fragments, both of which are subunits of the functional MLL2/COMPASS complex. MLL2 has also been implicated as a modulator of hematological malignancies (13). MLL3 and MLL4 proteins are not cleaved by Taspase 1.

  1. Miller, T. et al. (2001) Proc Natl Acad Sci U S A 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.
  7. Yokoyama, A. et al. (2004) Mol Cell Biol 24, 5639-49.
  8. Eissenberg, J.C. and Shilatifard, A. (2010) Dev Biol 339, 240-9.
  9. Smith, E. et al. (2011) Genes Dev 25, 661-72.
  10. Denissov, S. et al. (2014) Development 141, 526-37.
  11. Takeda, S. et al. (2006) Genes Dev 20, 2397-409.
  12. Yokoyama, A. et al. (2002) Blood 100, 3710-8.
  13. Chen, Y. et al. (2017) Cancer Cell 31, 755-770.e6.
Entrez-Gene Id
4221 , 4297 , 9757 , 9739 , 23067 , 11091 , 80335
Swiss-Prot Acc.
O00255 , Q03164 , Q9UMN6 , O15047 , Q9UPS6 , P61964 , Q6UXN9
For Research Use Only. Not For Use In Diagnostic Procedures.

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.

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