Product Pathways - PathScan ELISA
PathScan® Di-Methyl-Histone H3 (Lys4) Sandwich ELISA Kit #7124
|7124C||1 Kit (96 assays)||---||In Stock||---|
|7124||carrier free and custom formulation / quantity||email request|
When ordering five or more kits, please contact us for processing time and pricing at firstname.lastname@example.org.
|Kit Includes||Volume||Solution Color|
|Histone H3 Rabbit Antibody Coated Microwells||96 tests|
|Di-Methyl-Histone H3 (Lys4) Rabbit Detection mAb (Biotinylated)||1 ea||Green (Lyophilized)|
|HRP-Linked Streptavidin (ELISA Formulated)||1 ea||Red (Lyophilized)|
|Detection Antibody Diluent||11 ml|
|HRP Diluent||11 ml|
|TMB Substrate #7004||11 ml||Colorless|
|STOP Solution #7002||11 ml||Colorless|
|ELISA Wash Buffer (20X)||25 ml||Colorless|
|ELISA Sample Diluent||25 ml||Blue|
|Cell Lysis Buffer (10X) #9803||15 ml||Yellowish|
Note: 12 8-well modules – Each module is designed to break apart for 8 tests.
Storage: Kit should be stored at 4°C with the exception of Lysis Buffer, which is stored at –20°C (packaged separately).
The PathScan® Di-Methyl-Histone H3 (Lys4) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of histone H3 when di-methylated at Lys4. A total Histone H3 Rabbit monoclonal Antibody has been coated onto the microwells. After incubation with cell lysates, total Histone H3 is captured by the coated antibody. Following extensive washing, biotinylated di-methyl Histone H3(lys4) is added to detect the di-methyl Histone H3 (Lys4) protein. HRP-linked streptavidin is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of histone H3 di-methylated at Lys4.
Antibodies in kit are custom formulations specific to kit.
Specificity / Sensitivity
CST's PathScan® Di-Methyl-Histone H3 (Lys4) Sandwich ELISA Kit #7124 detects endogenous levels of histone H3 when di-methylated at Lys4. As shown in Figure 1 using the Di-Methyl-Histone H3 (Lys4) Sandwich ELISA Kit #7124, a high level of di-methylation at Lys4 on histone H3 is detected in COS cells when treated with TSA. The level of total histone H3 (modified and unmodified) remains unchanged as shown by Western analysis (Figure 1). Similar results are obtained when NIH/3T3 and Jurkat cells are treated with TSA (data not shown). This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.
ELISA - Western correlation
Figure 1. Treatment of COS cells with trichostatin A (TSA) increases the di-methylation of Histone H3 at Lys4 detected by PathScan® Di-Methyl-Histone H3 (Lys4) Sandwich ELISA Kit #7124. TSA treatment does not affect the level of histone H3 that is detected by Western analysis. COS cells (70-80% confluent) were treated for 16-18 hours with 0.4 μM TSA at 37ºC. Absorbance readings at 450 nm are shown in the top figure while the corresponding Western blots using Histone H3 Antibody #9715 (left panel) or Di-Methyl-Histone H3 (Lys4) (C64G9) Rabbit mAb #9725 (right panel) are shown in the bottom figure.
Modulation of chromatin structure plays an important role in the regulation of transcription in eukaryotes. The nucleosome, made up of DNA wound around eight core histone proteins (two each of H2A, H2B, H3, and H4), is the primary building block of chromatin (1). The amino-terminal tails of core histones undergo various post-translational modifications, including acetylation, phosphorylation, methylation, and ubiquitination (2-5). These modifications occur in response to various stimuli and have a direct effect on the accessibility of chromatin to transcription factors and, therefore, gene expression (6). In most species, histone H2B is primarily acetylated at Lys5, 12, 15, and 20 (4,7). Histone H3 is primarily acetylated at Lys9, 14, 18, 23, 27, and 56. Acetylation of H3 at Lys9 appears to have a dominant role in histone deposition and chromatin assembly in some organisms (2,3). Phosphorylation at Ser10, Ser28, and Thr11 of histone H3 is tightly correlated with chromosome condensation during both mitosis and meiosis (8-10). Phosphorylation at Thr3 of histone H3 is highly conserved among many species and is catalyzed by the kinase haspin. Immunostaining with phospho-specific antibodies in mammalian cells reveals mitotic phosphorylation at Thr3 of H3 in prophase and its dephosphorylation during anaphase (11).
- Workman, J.L. and Kingston, R.E. (1998) Annu Rev Biochem 67, 545-79.
- Hansen, J.C. et al. (1998) Biochemistry 37, 17637-41.
- Strahl, B.D. and Allis, C.D. (2000) Nature 403, 41-5.
- Cheung, P. et al. (2000) Cell 103, 263-71.
- Bernstein, B.E. and Schreiber, S.L. (2002) Chem Biol 9, 1167-73.
- Jaskelioff, M. and Peterson, C.L. (2003) Nat Cell Biol 5, 395-9.
- Thorne, A.W. et al. (1990) Eur J Biochem 193, 701-13.
- Hendzel, M.J. et al. (1997) Chromosoma 106, 348-60.
- Goto, H. et al. (1999) J Biol Chem 274, 25543-9.
- Preuss, U. et al. (2003) Nucleic Acids Res 31, 878-85.
- Dai, J. et al. (2005) Genes Dev 19, 472-88.
- Cantone, L. et al. (2011) Environ Health Perspect 119, 964-9. Applications: ELISA.
Have you published research involving the use of our products? If so we'd love to hear about it. Please let us know!
- 9803 Cell Lysis Buffer (10X)
- 9725 Di-Methyl-Histone H3 (Lys4) (C64G9) Rabbit mAb
- 9717 Histone H3 (3H1) Rabbit mAb
- 9715 Histone H3 Antibody
- 7121 PathScan® Acetyl-Histone H3 (Lys9) Sandwich ELISA Kit
- 7123 PathScan® Mono-Methyl-Histone H3 (Lys4) Sandwich ELISA Kit
- 7125 PathScan® Tri-Methyl-Histone H3 (Lys4) Sandwich ELISA Kit
- 7232 PathScan® Acetylated Histone H3 Sandwich ELISA Kit
- 9808 Phosphate Buffered Saline (PBS-20X)
- 9809 Phosphate Buffered Saline with Tween® 20 (PBST-20X)
- 7002 STOP Solution
- 7004 TMB Substrate
This product is intended for research purposes only. The product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.
PathScan is a trademark of Cell Signaling Technology, Inc.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.