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PathScan® Acetylated Histone H3 Sandwich ELISA Kit #7232

cell based assay   cell-based assay   Histone H3 elisa   sandwich elisa   screening  


No. Size Price
7232C 1 Kit ( 96 assays ) ¥6,345.00 现货查询 购买询价
Product Includes Volume Solution Color
ELISA Sample Diluent 25 ml
Anti-rabbit IgG, HRP-linked Antibody (ELISA Formulated) 1 ea Red (Lyophilized)
Detection Antibody Diluent 11 ml Green
HRP Diluent 11 ml Red
Acetylated-Lysine Rabbit Detection mAb 1 ea Green (Lyophilized)
STOP Solution #7002 11 ml Colorless
TMB Substrate #7004 11 ml Colorless
ELISA Wash Buffer (20X) 25 ml Colorless
Cell Lysis Buffer (10X) #9803 15 ml Yellowish
Sealing Tape 2 sheets
Histone H3 Ab Coated Microwells 96 tests

Specificity / Sensitivity

CST's PathScan® Acetylated Histone H3 Sandwich ELISA Kit detects endogenous levels of Acetylated Histone H3. Using this Sandwich ELISA Kit #7232, acetylated lysines on Histone H3 are detected when treated with TSA in Jurkat cells. However, the levels of Histone H3 remains unchanged, as shown by Western analysis using the Histone H3 Antibody #9715 (figure 1). COS and HIH 3T3 cells treated with TSA show similar results (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.

CST's PathScan® Acetylated Histone H3 Sandwich ELISA Kit能够检测内源性乙酰化的Histone H3蛋白水平。用TSA处理Jurkat细胞后,使用Sandwich ELISA Kit #7232检测Histone H3蛋白的乙酰化的赖氨酸。然而,使用Histone H3 Antibody #9715进行免疫印迹(western blot)分析,Histone H3蛋白水平没有改变(图1)。使用TSA处理的COS和HIH 3T3细胞显示类似的结果(数据未显示)。内部验证显示,该试剂盒检测已列出的物种,但是可能也会检测其它物种的同源性蛋白。


The PathScan® Acetylated Histone H3 Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of acetylated lysines on Histone H3. A Histone H3 Antibody has been coated onto the microwells. After incubation with cell lysates, Histone H3 is captured by the coated antibody. Following extensive washing, an Acetylated-Lysine Mouse mAb is added to detect the acetylated lysines on the Histone H3 protein. Anti-mouse IgG, HRP linked Antibody #7076 is then used to recognize the bound detection antibody. HRP substrate, TMB is added to develop color. HRP substrate, TMB is added to develop color. The magnitude of optical density for this developed color is proportional to the quantity of acetylated Histone H3. 
 Antibodies in kit are custom formulations specific to kit.

该PathScan® Acetylated Histone H3 Sandwich ELISA Kit是一个固相夹心酶联免疫吸附法,其能检测内源性赖氨酸乙酰化的Histone H3蛋白。一个Histone H3 Antibody已经被包被在微孔板上。在与细胞裂解液一起孵化后,Histone H3蛋白被包被的抗体吸附。清洗彻底后,加入Acetylated-Lysine Mouse mAb鼠单抗检测赖氨酸乙酰化组蛋白H3的水平。再加入Anti-mouse IgG, HRP 偶联抗体 #7076作为二抗来识别之前绑定的检测抗体。最后加入HRP底物(TMB)进行显色。该显色的吸光度值的大小与在乙酰化的Histone H3蛋白量成正比例。试剂盒里的抗体是针对该试剂盒特定制备的。

ELISA - Western correlation

ELISA - Western correlation

Figure 1: Treatment of Jurkat cells with TSA causes accumulation of acetylation on Histone H3, detected by Sandwich ELISA kit #7232, but does not affect the level of total Histone H3 protein, detected by western analysis. OD 450nm readings are shown in the top figure, while the corresponding western blot using the Acetylated Lysine Mouse mAb (Ac-K-103) #9681 (left panel) or Histone H3 Antibody #9715 (right panel), is shown in the bottom figure.

图1:使用TSA刺激Jurkat细胞可以引起乙酰化的histone H3的蛋白堆积,随后用Sandwich ELISA kit #7232检测,通过免疫印迹检测发现不影响histone H3的总蛋白水平。在450 nm吸光度值显示在最上图中,然而通过使用Acetylated Lysine Mouse mAb (Ac-K-103) #9681 (左图)和Histone H3 Antibody #9715 (右图),相应的免疫印迹(western blot)显示在下图中。



Figure 2: The relationship between protein concentration of lysates from untreated and TSA treated HeLa cells and kit assay optical density readings. HeLa cells were treated with TSA (4.0 µM overnight) and lysates were prepared using Cell Lysis Buffer (10X) #9803.

图2:HeLa细胞裂解的蛋白浓度(未处理或TSA处理)与试剂盒分析的光密度读数的关系。HeLa细胞使用TSA(4.0 µM 过夜)处理,然后使用Cell Lysis Buffer (10X) #9803裂解。


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).

染色质结构的修饰在调节真核细胞的转录中扮演着重要的角色。由DNA围绕和八聚体组蛋白(H2A,H2B,H3和H4各两个)共同组成的核小体是染色质的主要组成(1)。核小体的组蛋白氨基酸尾端进行不同的转录后修饰,包括乙酰化,磷酸化,甲基化和泛素化(2-5)。这些修饰通过不同的刺激产生并且对转录因子能否接近染色质有着直接的影响,所以也影响着基因的表达(6)。在大多数的物种中组蛋白H2B主要在Lys5,,12,,15和20位点上发生乙酰化(4,7)。组蛋白H3主要是在Lys9,14,18,23,27和56位点上发生乙酰化。在某些物种里H3上的Lys9位点发生乙酰化并应该在组蛋白沉积和染色质组装中扮演着重要的角色(2,3)。组蛋白H3上Ser10,Ser28和Thr11位点的磷酸化在有丝分裂和无丝分裂中都与染色质的缩合紧密相连(8-10)。H3的Thr3位点的磷酸化在许多物种中都是高度保守的,是由kinase haspin所催化的。在哺乳动物中用磷酸化特异性的抗体做免疫组化显示H3的Thr3位点在有丝分裂的前期发生磷酸化,后期发生去磷酸化(11)。

  1. Workman, J.L. and Kingston, R.E. (1998) Annu Rev Biochem 67, 545-79.
  2. Hansen, J.C. et al. (1998) Biochemistry 37, 17637-41.
  3. Strahl, B.D. and Allis, C.D. (2000) Nature 403, 41-5.
  4. Cheung, P. et al. (2000) Cell 103, 263-71.
  5. Bernstein, B.E. and Schreiber, S.L. (2002) Chem Biol 9, 1167-73.
  6. Jaskelioff, M. and Peterson, C.L. (2003) Nat Cell Biol 5, 395-9.
  7. Thorne, A.W. et al. (1990) Eur J Biochem 193, 701-13.
  8. Hendzel, M.J. et al. (1997) Chromosoma 106, 348-60.
  9. Goto, H. et al. (1999) J Biol Chem 274, 25543-9.
  10. Preuss, U. et al. (2003) Nucleic Acids Res 31, 878-85.
  11. Dai, J. et al. (2005) Genes Dev 19, 472-88.

Application References

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

PathScan is a trademark of Cell Signaling Technology, Inc.

Cell Signaling Technology® is a trademark of Cell Signaling Technology, Inc.

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