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EpiQuik Chromatin Immunoprecipitation (ChIP) Kit

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For immunoprecipitating chromatin specifically from cell input samples via microplate format

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Suggested Workflow
Chromatin Isolation
 
 
Chromatin Shearing
 
 
ChIP
 
 
PCR Analysis
 
Schematic procedure of the EpiQuik™ Chromatin Immunoprecipitation (ChIP) Kit.
ChIP was performed according to the protocol of the EpiQuik™ Chromatin Immunoprecipitation (ChIP) Kit using U2OS cells (1 x 106/well).
Comparative overview of the EpiQuik™ Chromatin Immunoprecipitation (ChIP) Kit.
Input Type: Chromatin
Research Area: Chromatin & Transcription
Target Application: Immunoprecipitation
Vessel Format: 96-Well Plate
100% Guarantee: 6 months
Catalog No.SizePriceQty
P-2002-124 reactions $234.00 
P-2002-248 reactions $380.00 
Availability: Usually Ships in 1 Day or Same day delivery Same Day NY Delivery 
Product Overview

The EpiQuik™ Chromatin Immunoprecipitation (ChIP) Kit is a complete set of optimized reagents to perform chromatin immunoprecipitation (ChIP) via a convenient, microplate-based format. The kit is ready-to-use and provides all the essential components needed to carry out a successful ChIP experiment. The EpiQuik™ ChIP kits are suitable for combining the specificity of immunoprecipitation with qualitative and quantitative PCR, ChIP-Seq, and ChIP-on-chip. This kit has the following advantages:

  • Fast and easy microplate-based procedure, to completed within 5 hours.
  • Strip microwell format makes the assay flexible: manual or high throughput.
  • Columns for DNA purification are included: save tremendous amounts of time and reduce unnecessary physical labor.
  • Compatible with all DNA amplification-based approaches.
  • Achieves very reliable and consistent assay conditions.

See also a quick chart to compare ChIP kits.

Background Information
Protein-DNA interaction play 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 process. Chromatin Immunoprecipitation (ChIP) offers an advantageous tool for studying protein-DNA interactions. Unlike other methods such as EMASA, DNA microarrays, and report gene assayswhich analyze direct interactions between protein and DNA in vitro- ChIP can detect that a specific protein binds to the specific sequences of a gene in living cells. 

Principle & Procedure
This ChIP kit includes a positive control antibody (RNA polymerase II), a negative control normal mouse IgG, and GAPDH primers that can be used as a positive control to demonstrate the efficacy of the kit reagents and protocol. RNA polymerase II is considered to be enriched in the GAPDH gene promoter that is expected to be undergoing transcription in most growing mammalian cells and can be immunoprecipitated by RNA polymerase II but not by normal mouse IgG. In this ChIP, cells are cross-linked with formaldehyde and chromatin is extracted. The chromatin is then sheared and added into the microwell immobilized with affinity antibodies. Cross-linked DNA is released from antibody-captured protein-DNA complex, reversed and purified through the specifically designed F-Spin Column. Eluted DNA can be used for various down-stream applications.

Starting Materials
Starting materials can include various cell samples. In general, the input amount should be from 500,000 to 2,000,000 cells for each reaction.

Product Components

CP1 (Wash Buffer)
CP2 (Antibody Buffer)
CP3A (Lysis Buffer)
CP3B (Lysis Buffer)
CP4 (ChIP Dilution Buffer)
CP5 (DNA Release Buffer)
CP6 (Reverse Buffer)
CP7 (Binding Buffer)
CP8 (Elution Buffer)
Protease Inhibitor Cocktail (100X)*
Normal Mouse IgG (1 mg/ml)*
Anti-RNA Polymerase II (1mg/ml)*
Proteinase K (10 mg/ml)*
Control Primer (GAPDH)
     Forward (20 µM)*
     Reverse (20 µM)*
8-Well Assay Strips (with Frame)
8-Well Strip Caps
F-Spin Column
F-Collection Tube
User Guide

* Spin the solution down to the bottom before use.

Frequently Asked Q's

1. Can IP be overnight?
Yes.

2. Is there any background DNA by mouse IgG IP?
Yes, a little < 2 ng.

3. Can any species Ab can be used for the IP?
Yes, if it is an IgG antibody.

4. Can kit P-2002 be used for frozen tissues?
Yes, but it is not as good as fresh samples.

5. How much volume of elution should be added into PCR reaction?
2 µl.

6. Are cell extracts containing 1%SDS compatible with CP4?
No, the SDS concentration in IP solution should be less than 0.1%.

7. Why is there no difference of CT values between the negative control and samples?
A. Increase washing time by one.
B. Check if antibody is chip grade.
C. There is no enrichment on the promoter.

8. Why is the difference between negative control and positive control not significant?
A. For conventional PCR, reduce PCR cycles to 35-36 or optimize the PCR cycles.
B. Increase washing time by one additional wash.

User Guide & MSDS

[User Guide]*
*Always use the actual User Guide that shipped with your product. Is the above file locked? You can also request user guides by emailing info@epigentek.com along with your contact information and institution name.

[Material Safety Data Sheet]
Product Citations

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Teng Y et. al. (November 2007). AS1411 alters the localization of a complex containing protein arginine methyltransferase 5 and nucleolin. Cancer Res. 67(21):10491-500.

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Customer Reviews

Rating By m*****@ucdenver.edu Verified Purchase Reviewed on: Tuesday 20 December, 2016
Application Description
The product I used is EpiQuick Chromatin Immunoprecipitation (ChiP) Kit (P-2002), and CtBP1 antibody (A-2705). I used this for a Chip assay to examine genes that bind with CtBP1 protein.

Pros: The antibody works good with human cells on CHiP assay. The ChiP kit is quick. Does not require huge amount of cells to start with.

Cons: The tiny tubes provided are kind of hard to work with. Not easy to label or to put in and get out of the heater we used.

Other Thoughts
The user guide should write more details

#1: Since the kit is shipped in two parts. One is stored at room temperature and the other is stored at 4C. I once accidentally stored all at room temperature for a couple of days. I called the technique support and asked if I can still use the part should be stored at 4C. I hope this can be written in the storage section.

The following suggestions are for people who has no previous experience with ChiP assay:

#2: The negative (normal mouse IgG) and positive control (Anti-RNA Polymerase II) are for the whole assay, not for each individual sample.

#3: When collecting cells, what temperature should people centrifuge the cells with? room temperature or 4C? I assume it is room temperature because other steps are done at room temperature.

#4: step 1 during cell lysis: since use ice cold PBS, so centrifuge also at 4C? after centrifuge, remove PBS

#5: step 2 during cell lysis: after votext 10second, centrifuge at 4C?

#6: last step of DNA shearing: what is the temperature of centrifuge? cross-linked DNA reversal/DNA purification procedure:

#7: step 3: add 150ul of CP7 to the samples and transfer to the columns. How about "input DNA"?

#8: step 4-7: should people do the same thing for the "input DNA" vials?

I hope I described everything clearly. In general, please treat user as beginners and write as much details as possible. If you can, also write down the purpose of each step.
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