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MAGnify™ Chromatin Immunoprecipitation System (Applied Biosystems™)

The MAGnify™ Chromatin Immunoprecipitation System provides a streamlined, optimized assay for the enrichment of chromatin/protein complexes and DNA recovery using magnetic bead capture technology. The isolated DNA is ready for downstream analysis by methods such as PCR- or qPCR-based assays, or massive parallel DNA sequencing.

Chromatin immunoprecipitation (ChIP) is a powerful technique for studying the association of certain proteins with specific regions of the genome. These sequence-specific DNA-binding proteins are believed to play a role in such cellular processes as DNA replication, recombination, repair, and segregation; chromosomal stability; cell-cycle progression; and epigenetic silencing. In a standard ChIP assay, a cell is fixed via formaldehyde treatment and the chromatin is sheared and immunoprecipitated via a highly specific antibody. The researcher then analyzes the DNA to identify the genomic regions where the chromatin-associated proteins bind to the chromatin in vivo. This kit enables researchers to start with lower sample amounts than traditional ChIP workflows, thereby preserving precious samples, and the protocol can be completed in a single day, compared with 2–3 days for a traditional ChIP assay. The kit can be used with our suite of ChIP-validated antibodies, and is also complementary with MethylCode™ and NCode™ products for downstream epigenetics research.

Using the MAGnify™ system
When using the MAGnify™ system, you treat cells or tissue with formaldehyde to generate protein-protein and protein-DNA crosslinks between molecules in close proximity within the chromatin complex. The cells are then lysed, and the chromatin is released from the nuclei and sheared by sonication to reduce the average DNA fragment size to 200–500 bp for analysis by quantitative real-time PCR (qPCR) or 100–300 bp for analysis by massive parallel DNA sequencing. You then immunoprecipitate and isolate the crosslinked protein of interest using a specific ChIP-qualified antibody conjugated to Dynabeads® Protein A/G. The formaldehyde crosslinking is reversed by heat treatment, and the DNA associated with that protein is purified. The DNA is now ready for downstream analyses such as end-point PCR or quantitative PCR (qPCR), genome-wide analyses using promoter-tiling arrays, or next-generation sequencing. In PCR/qPCR analysis, primers are designed to span the desired DNA sequence of interest, and the data demonstrates whether the specific protein of interest is associated in vivo with that DNA region.

Pierce™ Agarose ChIP Kit (Thermo Scientific™)

The Thermo Scientific Pierce Agarose ChIP Kit provides a complete set of reagents and a simple, fast and reproducible protocol to perform chromatin immunoprecipitation (ChIP) assays.

Features of the Agarose ChIP Kit:

• Simple and fast ChIP assay protocol (< 8 hours)
• Highly efficient isolation and lysis of nuclei
• Easy and reproducible enzymatic digestion
• Low-background and high-binding capacity Protein A/G agarose resin
• Highly specific positive controls included: RNA polymerase II antibody and GAPDH PCR primers
• Fast and reproducible spin-column format
• High-recovery DNA purification
• ChIP-validated antibodies available

The Pierce Agarose ChIP Kit contains sufficient reagents to perform 30 ChIP assays with appropriate controls using an optimized protocol and a convenient spin-column format. ChIP-validated and quality-guaranteed antibodies are also available for use the the Pierce Agarose ChIP Kit. Because antibodies that work for western blotting may not function in ChIP assays, we have assembled our most popular ChIP-validated monoclonal and polyclonal antibodies here to help you move from experimental setup to results more quickly.

The strength of the ChIP assay is its ability to capture a snapshot of specific protein-DNA interactions as they occur in living cells, and then quantitate the interactions using standard or quantitive PCR. A successful ChIP assays requires a number of critical steps to be carried out (crosslinking, chromatin preparation, immunoprecipitation) prior to detecting the target genomic DNA sequence. Individually, each step in the ChIP assay protocol can be time consuming to optimize. The Pierce Agarose ChIP Assay Kit simplifies the process, making accurate and reproducible results easier to obtain.

To perform ChIP assays with the Pierce Agarose ChIP Kit, protein-DNA complexes are stabilized and then extracted. In vivo crosslinking is traditionally achieved with formaldehyde. Crosslinking performed directly in cells locks in the protein-DNA complexes, trapping these unstable and sometimes transient interactions. To lyse, extract and solubilize the crosslinked complexes, the ChIP assay kit includes the Thermo Scientific Chromatin Prep Module (also sold separately, Part No. 26158). Together, this complete ChIP assay kit provides a simple, reliable and convenient means for isolating chromatin-bound DNA and enriching samples for the proteins of interest with less than 15% contamination from other cellular compartments - without a Dounce homogenizer.

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Pierce™ Magnetic RNA-Protein Pull-Down Kit (Thermo Scientific™)

The Thermo Scientific Pierce Magnetic RNA-Protein Pull-Down Kit provides researchers with a streamlined, robust method to enrich protein-RNA interactions using end-labeled RNA as the bait.

Features of the Magnetic RNA-Protein Pull-Down Kit:

Direct – capture ribonucleoprotein complexes directly with end-labeled RNA; does not use or require antibodies for pull-down
Easy to use – no spin cups or centrifugation necessary for the enrichment of RBP; procedure streamlined for minimal hands-on time (less than 3 hours) after RNA labeling reaction
Flexible – use in vitro transcribed RNA or synthetic RNA for labeling of various lengths and complexity; proteins successfully enriched using endogenous, over-expressed, and in vitro translated lysates
Specific – low bead background; unrelated RNA or mutated RNA does not significantly enrich specified RBPs
Economical – less expensive than purchasing commercially synthesized end-labeled RNA, magnetic beads, and reagents separately
Complete – contains both labeling and enrichment modules with buffers necessary for assay; positive control RNA, negative control RNA, and RBP antibody included

The Magnetic RNA-Protein Pull-Down Kit provides reagents to efficiently enrich RNA Binding Proteins (RBPs) using RNA end-labeled with desthiobiotin and streptavidin magnetic beads. The complete kit contains sufficient reagents for 20 RNA labeling reactions and 20 protein-RNA pull-down assays. This direct enrichment of the protein-RNA interaction provides an alternative to antibody capture of protein-RNA complexes or moieties incorporated into the nucleic acid. An added advantage to the kit is that it includes validated controls for both the labeling and pull-down assay. The kit is amenable to several downstream applications, including Western blotting and Mass Spectrometry (MS).

Complete kit contains the Pierce RNA 3'-End Desthiobiotinylation Kit, positive and negative RNA controls, nucleic-acid compatible streptavidin magnetic beads, and buffers for RBP enrichment and elution

User-supplied RNA and lysate (experimental sample)

• Mutational analysis
• Structure function analysis
• Identification of RNA:Protein interactions
• MS analysis of unknown RNA:protein binding pairs or complexes

Utilizing labeled RNA as bait for RNA-binding protein enrichment is advantageous over antibody enrichment in that it captures the interaction directly. Included in this kit is the Pierce RNA 3'-End Desthiobiotinylation Kit, which uses T4 RNA ligase to attach a single cytidine bisphosphate nucleotide to the 3'-ends of single-stranded RNA. Desthiobiotin may be used for detection or as an elutable affinity handle. The length of the spacer between the nucleotide and desthiobiotin has been optimized for efficient attachment to the beads without compromising the accessibility of the RNA to protein.

The procedure for enrichment of RNA binding proteins has been optimized for ease-of-use. The labeled RNA is first captured to the beads to orient the RNA for protein binding. RNA-bound beads are then equilibrated in Protein RNA Binding buffer before protein lysate is added. After washing, sample can be eluted using either nondenaturing Biotin Elution Buffer or SDS-PAGE loading buffer. Eluted samples are ready for a variety of downstream applications, including Western blotting or Mass spectrometry.

The control system for the pull-down assay utilizes 3'-untranslated region androgen receptor (AR) RNA, poly(A)25 RNA, and mammalian cell lysate. The proximal 3'-untranslated region (UTR) of androgen receptor RNA contains UC-rich regions for HuR and Poly(C) Binding Proteins (CP1 and 2). These RNA binding proteins regulate mRNA stability (HuR) and mRNA turnover and translation (CP1 and 2). The negative control RNA, poly(A)25 RNA, does not contain HuR or poly(C) BP binding sites.

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Pierce™ Magnetic ChIP Kit (Thermo Scientific™)

The Thermo Scientific Pierce Magnetic ChIP Kit provides a convenient method for efficient isolation of chromatin-bound DNA by immunoprecipitation (chromatin IP) for subsequent quantitation by PCR.

Features of the Magnetic ChIP Kit:

Rapid—obtain purified DNA that is ready for quantitative PCR in about 8 hours
Efficient and reproducible—micrococcal nuclease digestion and nuclear lysis are highly optimized
Sensitive—obtain results with as few as 10,000 cells (1 x 10^4)
Low background—Pierce Protein A/G Magnetic Beads are blocked in a non-DNA-containing reagent to minimize background
Complete—optimized positive control reagents are included: RNA polymerase II antibody and GAPDH promoter PCR primers

The Pierce Magnetic ChIP Kit contains sufficient reagents to perform 30 chromatin immunoprecipitation (ChIP) assays with appropriate controls using an optimized protocol. The kit includes reagents for cell lysis, capture of protein-DNA complexes, reversal of crosslinking, and DNA isolation. The blocked Pierce Protein A/G Magnetic Beads provide high binding capacity, low non-specific background, and work with many antibody species. These beads can be used manually with a magnetic stand or with automated platforms, such as the Thermo Scientific KingFisher Instruments. ChIP-validated and quality-guaranteed antibodies are also available for use with the Pierce Magnetic ChIP Kit.

Kit contains reagents for chromatin preparation, IP, DNA purification, and positive controls (antibody and primers)

In vivo crosslinker (such as formaldehyde), micro-tip sonicator (such as Misonix™ Sonicator 3000), ChIP qualified antibody of choice, PCR primers for DNA sequence of interest, PCR master mix (containing a dye such as SYBR™ Green, if qPCR is desired), and a qPCR instrument

• Determine sites of specific protein-DNA interactions on genomic DNA by PCR or ChIP-Seq
• Monitor the effects of histone modifications or chemical agents on DNA binding

A successful ChIP assay requires a number of critical steps (crosslinking, chromatin preparation, immunoprecipitation) prior to detecting the target genomic DNA. Individually, each step in the ChIP protocol can be time-consuming to optimize. The Thermo Scientific Pierce Magnetic ChIP Kit simplifies the ChIP process, enabling accurate and reproducible results.

Protein-DNA complexes are first crosslinked in vivo with formaldehyde. The kit contains reagents to lyse cells and extract and solubilize the crosslinked complexes. The complexes are then incubated with a specific antibody and isolated using Pierce Protein A/G Magnetic Beads. After reversing crosslinks and digesting protein, the resulting DNA fragments are purified and are then ready for standard or quantitative PCR.

More Product Data
Analysis of androgen-dependent and -independent regulation of transcriptional activity

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SOLiD™ ChIP-Seq Kit, with ChIP magnet (Applied Biosystems™)

The SOLiD™ ChIP Seq Kit is a complete and optimized system for genome wide Chromatin Immunoprecipitation (ChIP) analysis on SOLiD™ and includes ChIP preparation reagents and library generation reagents for 20 samples along with a protocol optimized for library preparation with reduced amount of DNA from 1-10ng samples. Chromatin immunoprecipitation (ChIP) experiments examine histone modifications and genomic DNA sequences bound to specific regulatory proteins and when combined with next generation sequencing is a powerful method to examine genome wide protein DNA interactions. This configuration also includes the DynaMAG®-PCR magnet for processing ChIP samples.

Pierce™ Chromatin Prep Module (Thermo Scientific™)

The Thermo Scientific™ Pierce™ Chromatin Prep Module provides a simple, fast, and reproducible method to isolate chromatin for use in assays that monitor transcription regulation through histone modification1 (epigenetics) or transcription factor-DNA binding interactions, such as chromatin immunoprecipitation (ChIP). The module contains all reagents necessary to isolate crosslinked chromatin from mammalian cells.

This Chromatin Prep Module is also included as part of our complete Agarose ChIP Kit.

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