High-Sensitivity ChIP Kit (ab185913)

Submit a review Q&A (1)References (14)
High abundance protein enrichment:
  • Low abundance protein enrichment:
  • Histone H3K18ac ChIP assay example data

Key features and details

  • Sample type: Adherent cells, Suspension cells, Tissue

Overview

  • Product name

    High-Sensitivity ChIP Kit
    See all ChIP Kit kits

  • Sample type

    Tissue, Adherent cells, Suspension cells

  • Species reactivity

    Reacts with: Mammals, Other species

  • Product overview

    High-Sensitivity ChIP Kit (ab185913) is a complete set of optimized reagents to carry out a successful chromatin immunoprecipitation procedure in a high throughput format starting from mammalian cells or tissues. The highly specific and sensitive kit is suitable for selective enrichment of a chromatin fraction containing specific DNA sequences using various mammalian cell/tissues. The optimized protocol and kit components reduce non-specific background ChIP levels to allow capture of low abundance protein/transcription factors and increased specific enrichment of target protein/DNA complexes. The target protein bound DNA prepared with the High-Sensitivity ChIP Kit can be used for various downstream applications including PCR (ChIP-PCR), microarrays (ChIP-on-chip), and sequencing (ChIP-seq).


     


    Starting Materials
    Starting materials can include various tissue or cell samples such as cells from flask or plate cultured cells, fresh and frozen tissues, etc. In general, the amount of cells and tissues for each reaction can be 2 x 103 to 1 x 106 and 0.5 mg to 50 mg, respectively. For optimal preparation, the input amount should be 1-2 x 105 cells or 10-20 mg tissues since the enrichment of target proteins to genome loci may vary. For the target proteins that are low abundance transcription factors, the input amount should be 5-6 x 105 cells or 50 to 60 mg tissues.


     


    Primers


    The GAPDH primers provided with the kit are for the human sequence. If using the kit with a difference species, GAPDH primers for that species will need to be acquired.


     

  • Notes

    Protein-DNA interaction plays a critical role for cellular functions such as signal transduction, gene transcription, chromosome segregation, DNA replication and recombination, and epigenetic silencing. Identifying the genetic targets of DNA binding proteins and knowing the mechanisms of protein-DNA interaction is important for understanding cellular processes. Chromatin immunoprecipitation (ChIP) offers an advantageous tool for studying such protein-DNA interactions. It allows for the detection of a specific protein bound to a specific gene sequence in living cells using PCR (ChIP-PCR), microarrays (ChIP-chip), or sequencing (ChIP-seq). For example, measurement of the amount of methylated histone H3 at lysine 9 (meH3-K9) associated with a specific gene promoter region under various conditions can be achieved through a ChIP-PCR assay, while the recruitment of methylated H3-K9 to the promoters on a genome-wide scale can be detected by ChIP-on-chip or ChIP-sequencing. ChIP analysis requires that ChIPed DNA contains minimal background in order to reliably identify true TF-enriched regions. High background in ChIP is mainly caused ineffective wash buffers, insufficient cross-link reversal, inappropriate DNA fragment length, and residual RNA interference. To effectively capture TF/DNA complexes, which are often in low abundance, an ideal ChIP method requires having maximum sensitivity with minimized background levels. This method should also be able to enrich highly abundant protein/DNA complexes using a small amount of cells or tissues in a high throughput format. The High-Sensitivity ChIP Kit is designed to achieve these goals by maximizing sensitivity and minimizing non-specific background signals.

    ChIP assay products and guides

    Find more ChIP assay / chromatin immunoprecipitation resources and products, ChIP antibody products, and other ChIP assay kits and related reagents.

  • Tested applications

    Suitable for: ChIPmore details

Properties

Applications

Our Abpromise guarantee covers the use of ab185913 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
ChIP Use at an assay dependent concentration.

The GAPDH primers provided with the kit are for the human sequence. If using the kit with a difference species, GAPDH primers for that species will need to be acquired.

Images

  • High abundance protein enrichment:
    High abundance protein enrichment:

    Sheared chromatin isolated from different numbers of MBD-231 cells was used for ChIP-qPCR analysis of RNA polymerase II enrichment in GAPDH promoters using ab185913 and a quantitative PCR Fast Kit.

  • Low abundance protein enrichment:
    Low abundance protein enrichment:

    Sheared chromatin isolated from different numbers of MCF7 cells was used for ChIP-qPCR analysis of ER-a enrichment in TFF1 promoters using ab185913 and a quantitative PCR Fast Kit.

  • Histone H3K18ac ChIP assay example data
    Histone H3K18ac ChIP assay example data

    Histone H3K18ac ChIP assay was carried out using ab185913.

    ChIP-Seq reads align with the same peak sites as ENCODE data.

References (14)

Publishing research using ab185913? Please let us know so that we can cite the reference in this datasheet.

ab185913 has been referenced in 14 publications.

  • Zhou P  et al. Transforming growth factor beta (TGF-ß) is activated by the CtBP2-p300-AP1 transcriptional complex in chronic renal failure. Int J Biol Sci 16:204-215 (2020). PubMed: 31929749
  • Mu G & Chen F Oncogenic Roles Of A Histone Methyltransferase SETDB2 In AML1-ETO Positive AML. Cancer Manag Res 12:783-792 (2020). PubMed: 32099474
  • Lu W  et al. The CARM1-p300-c-Myc-Max (CPCM) transcriptional complex regulates the expression of CUL4A/4B and affects the stability of CRL4 E3 ligases in colorectal cancer. Int J Biol Sci 16:1071-1085 (2020). PubMed: 32140074
  • Chen FX  et al. Inflammation-dependent downregulation of miR-532-3p mediates apoptotic signaling in human sarcopenia through targeting BAK1. Int J Biol Sci 16:1481-1494 (2020). PubMed: 32226296
  • Pignatti E  et al. Beta-Catenin Causes Adrenal Hyperplasia by Blocking Zonal Transdifferentiation. Cell Rep 31:107524 (2020). PubMed: 32320669
View all Publications for this product

Customer reviews and Q&As

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Question

For the pulldown, is the enhancer mix protein A, G, or both? I have a goat primary so want to make sure it's going to work. Any information you can provide on how the protein A or G pulldown works would be helpful as well (beads in the enhancer or protein a/g already bound to the wells?)

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Answer

Protein G is pre-coated on the wells, so it should work for goat primary antibodies.

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