High-Sensitivity ChIP Kit

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.

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 resources and products, products, and other .