TIP-ChIP(TM) Assay Kit

TIP-ChIP is a high-throughput ChIP alternative with these key features:

• Map histone modifications in multiple samples simultaneously
• Chromatin is tagmented in an unbiased fashion
• Built-in normalization using barcoded inputs
• Works with low cell (150,000 to 300,000) input
• Significant reduction in hands-on-time compared to ChIP-seq

TIP-ChIP (Tagmented, Indexed, and Pooled ChIP-Seq is a streamlined cost-effective method for performing ChIP-Seq on up to 96 samples in a single reaction, with lower input and reduced variability than traditional Chromatin Immunoprecipitation. TIP-ChIP is a high-resolution, two-step chromatin immunoprecipitation (ChIP) epigenetic technique designed to precisely map protein–DNA interactions genome-wide from individual samples that are indexed and pooled to enable high-throughput ChIP from 300,000 to 1 million cells per sample.

This method is especially useful to perform ChIP in multiple samples in the same experiment. Because samples in TIP-ChIP are pooled together per target, there are fewer steps – only one IP per target as opposed to IPs per sample in traditional ChIP.

ChIP-Seq is a well-established technique for mapping protein-DNA interactions and histone modifications genome-wide; however, there are some limitations to ChIP-Seq including the large sample input requirement of 1 to 10 Million cells, time-consuming and cumbersome protocol, difficult sample chromatin preparation, one sample processed per reaction, and batch effects.

TIP-ChIP overcomes many of these limitations by combining samples into one reaction per immunoprecipitation (IP). By pooling samples, 300,000 to 1 million cells are needed per sample, and only 8 samples are needed to pool for one IP. Additionally, TIP-ChIP does not rely on traditional chromatin preparation, and instead utilizes Tn5 transposase to incorporate unique barcodes per sample. This streamlines the assay workflow, improves data quality, and eliminates batch effects.

How does TIP-ChIP work?

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Figure 1. TIP-ChIP Assay Workflow. TIP-ChIP enables treamlined high-throughput ChIP-Seq with an unbiased tagmentation method followed by IP pull-down.

What antibodies work in TIP-ChIP?

Active Motif specializes in manufacturing high-quality antibodies to histones, histone modifications, chromatin proteins, and other factors, including a growing list of antibodies that we have experimentally validated in-house to work well in TIP-ChIP assays. TIP-ChIP works well with many antibodies that work in ChIP-Seq, but not all antibodies that have been validated for ChIP will work well in TIP-ChIP, and it is important to choose validated antibodies for the application. Guidelines for optimizing antibodies in TIP-ChIP are included in the manual.

TIP-ChIP Assay Advantages

By utilizing Tn5 transposase, indexing and tagmentation are done simultaneously. Tn5 in traditional ATAC-Seq methods is biased toward open chromatin regions, and in TIP-ChIP a modification to these steps results in unbiased tagmentation.

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Figure 2. TIP-ChIP workflow starts with unbiased tagmentation. Tn5 tagmentation in TIP-ChIP is not biased to preferentially tagment only open chromatin regions.

TIP-ChIP Assay Kit Experimental Design

The TIP-ChIP Assay Kit includes H3K27ac, H3K4me3, and RNA pol II phospho Ser2 antibodies to profile histone marks and transcription activation. These three antibodies together give a powerful view of active gene regulation and transcriptional dynamics. Depending on experimental questions and sample amounts, the breakaway 96-well plate can be set up to use 24 wells of each antibody, or in other combinations to optimize antibodies for other targets of interest as illustrated below.

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TIP-ChIP™ Assay Kit Contents

• TIP-ChIP Assembled Transposomes Plate, store at -20°C
• Hypotonic Quench, store at 4°C
• 5X TC Tag Buffer, store at -20°C
• 10X PBS, store at 4°C
• Digitonin (1%), store at -20°C
• 10% Tween 20, store at RT
• Chromatin Opening Buffer, store at RT
• SPRI Beads, store at 4°C
• Releasing Buffer, store at RT
• Protease Inhibitor Cocktail (PIC) store at -20°C
• 5M NaCl, store at RT
• Proteinase K, store at -20°C
• RNase A, store at -20°C
• BSA 9(10 mg/ml), store at -20°C
• blocker, store at -20°C
• Blocking Reagent AM1, store at -20°C
• ChIP Buffer without SDS, Store at RT
• ChIP Buffer, store at RT
• LiCl Buffer, store at RT
• Wash Buffer AM1, store at RT
• TE, pH 8.0, store at RT
• Elution Buffer AM4, store at RT
• ChIP Filtration Columns, store at RT
• TIP-ChIP Columns, store at RT
• AM qPCR Dye, store at -20°C

• TIP-ChIP DNA Binding Buffer, store at RT
• TIP-ChIP DNA Wash Buffer, store at RT
• DNA Purification Elution Buffer, store at RT
• Protein A Agarose Beads, store at 4°C
• Q5 Polymerase NGS MM, store at -20°C
• AM i5-001 Primer, store at -20°C
• AM i5-002 Primer, store at -20°C
• AM i5-003 Primer, store at -20°C
• AM i5-004 Primer, store at -20°C
• AM i5-005 Primer, store at -20°C
• AM i7-001 Primer, store at -20°C
• AM i7-002 Primer, store at -20°C
• AM i7-003 Primer, store at -20°C
• AM i7-004 Primer, store at -20°C
• AM i7-005 Primer, store at -20°C
• Stop Solution, store at RT
• Diversi-Phi Indexed PhiX, store at -20°C
• AM qPCR Standard 1, store at -20°C
• AM qPCR Standard 2, store at -20°C
• AM qPCR Standard 3, store at -20°C
• AM qPCR Standard 4, store at -20°C
• AbFlex® Histone H3K27ac antibody (rAb), store at -20°C
• Histone H3K4 me3 antibody (pAb), store at -20°C
• AbFlex® RNA pol II CTD phospho Ser2 antibody (rAb), store at -20°C
• Tagmentation Stop Solution, store at 4°C

TIP-ChIP™ Assay Kit Data

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Figure 3. TIP-ChIP Data Demonstrates Strong Concordance with ENCODE ChIP-Seq Data. H3K27ac was profiled in 12 barcoded K562 samples pooled into a single TIP-ChIP reaction and demultiplexed by barcode. Genomic browser tracks show pseudobulk signal from the pooled data (red), per-sample demultiplexed signal (purple) and ENCODE ChIP-Seq tracks (gold). Concordant peak positions and amplitudes indicate that TIP-ChIP multiplexing and deconfolution preserve locus-level resolution and data quality relative to confentional ChIP-Seq.