FFPE ChIP-seq Service

Formalin-fixed, paraffin-embedded (FFPE) samples represent the gold standard for clinical tissue preservation. However, traditional chromatin-based assays are often incompatible with FFPE due to heavy crosslinking and fragmented nucleic acids. CD BioSciences's FFPE ChIP-seq service overcomes these challenges using optimized protocols that restore chromatin accessibility and deliver reproducible, high-quality ChIP-seq data from archived samples.

Introduction to FFPE ChIP-seq

FFPE ChIP-seq (Chromatin Immunoprecipitation followed by sequencing) enables genome-wide analysis of histone modifications or chromatin-associated proteins from formalin-fixed tissues. Unlike fresh or frozen samples, FFPE tissues suffer from extensive crosslinking and nucleic acid damage, making chromatin extraction inefficient.

Recent advances—including the Chrom-EX PE method—have introduced a critical tissue-level crosslink reversal step by heat incubation (e.g., 65°C), which significantly improves chromatin solubility, DNA fragment size control, and compatibility with downstream sequencing. By optimizing crosslink reversal temperature and sonication conditions, FFPE ChIP-seq now allows reliable recovery of histone marks (e.g., H3K4me3, H3K27ac, H3K27me3) and even non-histone proteins such as RNA Polymerase II.

Fig.1 A tissue-level cross-link reversal allows efficient extraction of high-quality chromatin from FFPE tissues. (Zhong, J., et al., 2019)

Applications of FFPE ChIP-seq

Our Services

To address the specific challenges of FFPE samples, CD BioSciences integrates controlled crosslink reversal and chromatin stabilization strategies into every step.

Sample Evaluation & Project Design

1. Review of sample format (block/section), fixation details, and target protein
2. Antibody selection and input estimation (as little as two 20-μm sections per IP)
3. Inclusion of proper controls (IgG, intergenic background)

Deparaffinization & Crosslink Reversal

1. Paraffin removal and progressive rehydration
2. Tissue-level crosslink reversal via overnight incubation in chromatin stabilization buffer at optimized temperatures (typically 65°C)

Chromatin Fragmentation

1. MNase digestion and/or mild sonication to obtain nucleosomal or sub-nucleosomal DNA (100–500 bp)
2. Fragmentation pattern adjusted according to desired ChIP target (broad vs. narrow peaks)

Chromatin Immunoprecipitation (ChIP)

1. Enrichment with validated antibodies for histone modifications or chromatin-bound proteins (e.g., H3K27ac, H3K4me3, RNA Pol II)
2. Extensive washing to reduce background

Library Preparation & Sequencing

1. Low-input–compatible library prep kits
2. Paired-end sequencing (Illumina platforms)
3. Typical output: 20–50M reads per sample

Bioinformatics & Comparative Analysis

1. Alignment, peak calling, signal normalization
2. Cross-sample correlation and replicate reproducibility checks (Pearson r > 0.9 in published validations)
3. Optional comparison with matched frozen sample data or public datasets

Supported Sample Types

1. FFPE Tissue Sections
2. Archived Clinical Tumor Specimens
3. Tissue Microarrays (TMA)

1. Laser Capture Microdissected FFPE Regions
2. Other Paraffin-Embedded Archival Samples
3. Other Types (Please Inquire)

Our Advantages

1. Crosslink Reversal Expertise: Thermal and enzymatic de-crosslinking tuned for FFPE preservation artifacts.
2. Histone and Pol II Compatibility: Validated for H3K27ac, H3K4me3, Pol II, and selected TFs.
3. Versatile Chromatin Recovery: Supports both MNase digestion and sonication methods.
4. Clinically Relevant Input Handling: Effective with archival blocks aged 3–10 years.
5. Comparative Analysis Ready: Enables integration with matched fresh/frozen ChIP data.
6. Full Workflow Optimization: Tailored steps from deparaffinization to sequencing for FFPE performance.

Unlock the epigenetic insights hidden in your archived tissue samples. CD BioSciences's FFPE ChIP-seq service delivers the sensitivity, reproducibility, and analytical depth needed to bring your clinical epigenomics forward. Contact us today to evaluate your FFPE material or request a pilot run.

Reference

1. Zhong, J., et al. (2019). Enhanced and controlled chromatin extraction from FFPE tissues and the application to ChIP-seq. BMC genomics, 20(1), 249.