Invitrogen™

OncoStat Panel, SNAP-ChIP™ Spike-in

Catalog Number	Quantity
A47343	10 reactions

Catalog number A47343

Price (EUR)

447,00

Quantity:

10 reactions

Price (EUR)

447,00

SNAP-ChIP (Sample Normalization and Antibody Profiling Chromatin ImmunoPrecipitation), a proprietary technology from EpiCypher, uses DNA-barcoded semi-synthetic nucleosomes bearing distinct histone mutations as spike-in ChIP controls.

• Homogenous, fully defined standards faithfully represent target mononucleosomes in the experimental sample
• Nucleosomes are subjected to rigorous quality control for lot-to-lot consistency
• Unique DNA barcodes can be distinguished from experimental sample genomes
• Spike-ins provide a direct readout of antibody performance in ChIP
• Ability to monitor technical variability between samples
• Defined standards enable universal normalization across experiments

SNAP-ChIP panels are directly compatible with your current ChIP workflow, with semi-synthetic nucleosomes bearing the mutation of interest immunoprecipitated and processed alongside sample chromatin.

Users can directly monitor antibody capability (specificity and enrichment) in a ChIP experiment by measuring the recovery of on- and off-target nucleosomes. The SNAP spike-ins can be further used as a defined standard to normalize your ChIP data, thereby controlling for technical variability across experiments.

The OncoStat Panel consists of a pool of nucleosomes harboring seven well-studied histone H3.3 mutations that have been implicated in cancer (histone H3.3K4M, H3.3K9M, H3.3K27M, H3.3G34R, H3.3G34V, H3.3G34W and H3.3K36M) plus a wild-type H3.3 control. Further, each of these eight nucleosomes is wrapped by two independent DNA barcodes (16 total barcodes in the panel), allowing for an internal technical replicate in each ChIP reaction. A single spike-in of the panel allows users to check antibody specificity by examining the post-IP recovery of on- versus off-target SNAP-ChIP nucleosomes, supporting the generation of high quality ChIP data.

For Research Use Only. Not for use in diagnostic procedures.

Specifications

DescriptionRecombinant Nucleosome Panel

No. of Tests10 Reactions

Quantity10 reactions

Product LineOncoStat, SNAP-ChIP

Product TypeSNAP-ChIP Panel

Unit SizeEach

Contents & Storage

Store at -5 to -30°C.

Frequently asked questions (FAQs)

Can you please recommend specific histone PTM antibodies?

Thermo Fisher Scientific (https://www.thermofisher.com/us/en/home.html), in collaboration with EpiCypher, has performed extensive antibody testing for various histone PTMs, using SNAP-ChIP spike-in control panels to validate antibody specificity. For more information, please go to:
https://www.thermofisher.com/us/en/home/life-science/antibodies/invitrogen-antibody-validation/SNAP-ChIP-antibody-validation.html.

Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

Will the SNAP-ChIP spike-ins affect the required sequencing depth?

The SNAP-ChIP spike-ins represent <<1% of total nucleosomes in the sample, so sequencing depth is unaffected.

Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

With SNAP-ChIP panels, do you have specific guidelines for running the sequencing?

Paired-end sequencing is recommended for several reasons:
- Because the reverse barcodes are unique to each dNuc within a panel, they will not be reachable from the top strand in many NGS configurations (e.g., single end 50, 75, or 100 bp sequencing). Thus, half of the data associated with the SNAP-ChIP spike-ins (those from the top strand) cannot be confidently aligned to a specific nucleosome in the panel and will be discarded. The read depth associated with the SNAP-ChIP spike-ins will be concomitantly reduced.
- Paired-end sequencing allows read filtering to eliminate data associated with dinucleosomes (immunoprecipitated more efficiently than mononucleosomes* and thus overrepresented in the sequencing data). This bias can be mitigated by excluding fragments sized >220 bp from analysis.
* Grzybowski, A. T., Chen, Z. & Ruthenburg, A. J. Calibrating ChIP-Seq with Nucleosomal Internal Standards to Measure Histone Modification Density Genome Wide. Molecular Cell 58: 886-899, doi:10.1016/j.molcel.2015.04.022 (2015).

Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

Can you please recommend a native ChIP protocol to use with SNAP-ChIP spike-in?

Please see the following reference for a detailed native ChIP method:
Brand, M., Rampalli, S., Chaturvedi, C. P. & Dilworth, F. J. Analysis of epigenetic modifications of chromatin at specific gene loci by native chromatin immunoprecipitation of nucleosomes isolated using hydroxyapatite chromatography. Nature Protocols 3: 398-409, doi:10.1038/nprot.2008.8 (2008).

Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

I am planning to use SNAP-ChIP panels. Why should I use a native ChIP protocol with micrococcal nuclease (MNase) digestion vs. crosslinking/sonication?

SNAP-ChIP spike-in is directly compatible with both native and crosslinked approaches though the former is recommended. Crosslinking can impact antibody specificity and enrichment because crosslinked chromatin becomes more sticky and susceptible to epitope masking. In our experience, signal-to-noise ratios are often decreased in crosslinked samples compared to native ChIP.
In contrast, a native nuclei preparation that is micrococcal nuclease-digested to yield >95% pure mononucleosomes will yield samples that more closely resemble the SNAP-ChIP spike-ins (i.e., unfixed mononucleosomes). As a result, data obtained from the SNAP-ChIP controls will be most representative of the experimental samples.

Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

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