H3K27bu - Chromatin Antibodies

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SNAP-ChIP Luminex

For final validation, we test each antibody using SNAP-ChIP^® (Sample Normalization & Antibody Profiling for ChIP) Spike-in Control Panels added to a ChIP experiment. Passing requires that an antibody display <20% cross-reactivity (relative to on-target binding) and >5% enrichment of on-target PTM vs. Input.

Heatmaps are sorted by PTM, and are structured such that the most specific antibodies that passed testing against the full SNAP-ChIP spike-in panel are ranked at the top. These antibodies are recommended for ChIP-seq. The remaining antibodies in the heatmap are separated into three groups: those that failed testing against the full SNAP-ChIP panel, antibodies that were only tested against a subset of spike-ins and passed, and those tested against a subset of spike-ins that failed. Within each subgroup, antibodies are ranked by specificity.

* Denotes antibodies that were tested in Shah et al. 2018.

Efficiency (% Recovered)

< 5% 5-10% > 10%

Efficiency, or enrichment, is the percent recovery of on-target PTM compared to input. Efficiency scores are binned into three categories, shown on the scale. >5% efficiency is required for SNAP-ChIP Certification.

Specificity (% On-Target)

0 20 40 60 80 100 120

Antibody binding is expressed relative to the on-target PTM. Binding is presented as a gradient from blue (weak activity) to red (strong activity). Increasing red across the entire panel denotes antibodies with high cross-reactivity.

There are no tests for this method.

We use Luminex^® multiplexing technology to screen antibody candidates against modified nucleosome substrates. To pass this triage experiment, antibodies must display <10% cross-reactivity relative to the on-target PTM.

Antibodies are sorted in the heatmap by PTM and then by overall specificity, with antibodies that passed Luminex and have highest specificity ranked at the top. Note that passing Luminex does not guarantee an antibody will work well in ChIP experiments; so be sure to confirm that your antibody also passes testing against SNAP-ChIP spike-ins.

Specificity (% On-Target)

0 20 40 60 80 100 120

K-AcylStat

H3K27bu

Unmodified

H3K4ac

H3K9ac

H3K9bu

H3K9cr

H3K14ac

H3K18ac

H3K18bu

H3K18cr

tetraAc-H3

H3K23ac

H3K27ac

H3K27bu

H3K27cr

H3K27acS28ph

H3K36ac

H4K5ac

H4K8ac

H4K12ac

H4K16ac

tetraAc-H4

H4K20ac

tetraAc-H2A

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