How to Choose: Enzymatic or Sonication Protocol for ChIP

This video breaks down four questions that will help you find a chromatin fragmentation protocol for your experimental needs.
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Transcript:
Researchers use chromatin immunoprecipitation, or ChIP, to identify and characterize protein DNA interactions in the context of chromatin.ChIP experiments can use varying input samples, and provide ChIP-qPCR or ChIP-seq readouts.
The success of your ChIP experiment depends on the fragmentation of chromatin, a critical step in the ChIP protocol. This can be accomplished with either sonication or enzymatic digestion. But how do you decide which chromatin fragmentation protocol to use in your ChIP experiments? A number of factors can influence your choice, making the decision seem daunting. So let's simplify. Cell Signaling Technology will show you the way.
In this video, we'll walk through four questions that will help you find a protocol and the ideal ChIP kit from Cell Signaling Technology that's suited to your experimental needs. First, what is your input sample type: cells or tissues? Second, what type of protein is being targeted for immunoprecipitation? Histones, transcription factors, and cofactors bind to DNA with varying strength, affecting ChIP efficiency. Other factors to consider are the abundance of the target protein in your sample and your preferred method for chromatin fragmentation. In many instances, you can use either sonication or enzymatic digestion. We'll also introduce some scenarios in which one method for chromatin fragmentation outperforms the other.
Traditional sonication protocols use harsh, denaturing conditions. Cell Signaling Technology scientists have developed specially formulated lysis buffers to protect chromatin integrity and antibody epitopes. This enables you to obtain more reliable results when analyzing transcription factors and cofactors using our ChIP kits. Let's walk through the scenario in which your input samples are cells. If you are looking at histones or high abundance transcription factors or cofactors, both sonication and enzymatic digestion work well. Select a SimpleChIP® kit based on your chromatin fragmentation preference. While sonication can be used with low-abundance transcription factors and cofactors, enzymatic digestion is preferred. The protein-DNA interactions for these proteins are more labile and may be prone to loss in sonication. For optimal results, we recommend using enzymatic fragmentation as a first choice.
Next, we'll consider tissues and how the type of tissue may affect how you choose a ChIP protocol. If you are performing ChIP with any type of tissue, it's critical to homogenize the input material to a single-cell suspension prior to nuclear lysis and chromatin fragmentation. SimpleChIP Plus kits for both sonication and enzymatic protocols are approved for use with most tissue types, including liver, brain, and heart. If you are looking at histones, high abundance transcription factors, or high abundance cofactors in these tissues, you can select either of these kits. For optimal ChIP results with low-abundance transcription factors and cofactors from these tissues, we recommend using the enzymatic protocol.
Note that especially fibrous tissues like skeletal muscle and fatty adipose tissues are more difficult to homogenize. A tissue-specific protocol is required to homogenize these samples into single-cell suspensions prior to chromatin fragmentation. For these fibrous or fatty tissues, Cell Signaling Technology recommends using the sonication protocol with the SimpleChIP Plus Sonication Chromatin IP Kit for all target types. Sonication is more effective than enzymatic digestion due to the significant force required to break apart these tissues, and lyse their cells and nuclei.
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Cell Signaling Technology, CST, and SimpleChIP are registered trademarks of Cell Signaling Technology, Inc.