Chromatin-Associated Interactome Profiling
Identify native protein complexes bound to transcription factors, cofactors, or chromatin regulators from nuclear extracts.
RIME-MS Analysis Service
Map Endogenous Chromatin-Bound Protein Interactions
RIME-MS (Rapid Immunoprecipitation–Mass Spectrometry) identifies endogenous, chromatin-anchored protein complexes that drive transcriptional regulation. At Creative Proteomics, we help researchers reveal transcription factor–cofactor assemblies, detect condition-specific rewiring, and validate chromatin-bound mechanisms that cannot be captured by conventional proteomics.
Problems We Solve
- Difficulty mapping nuclear or chromatin-bound interactors at endogenous levels.
- Limited sensitivity for weak or transient transcriptional complexes.
- Gaps between ChIP-seq binding profiles and complex composition data.
Our Advantages
- Optimized Orbitrap-based RIME-MS workflow for high-confidence nuclear interactomes.
- Endogenous, tag-free capture preserving native PTMs and stoichiometry.
- Integrative bioinformatics linking interactome data with ChIP-seq, ATAC-seq, and RNA-seq.
From transcription factor biology to chromatin pharmacology, RIME-MS empowers deeper mechanistic insight into gene regulation.
What Is RIME-MS?
RIME-MS (Rapid immunoprecipitation–mass spectrometry of endogenous proteins) maps chromatin-anchored protein interactions in their native regulatory context. The workflow enriches an endogenous bait by immunoprecipitation, stabilizes nuclear assemblies via gentle crosslinking, and identifies co-purifying partners using high-resolution LC–MS/MS. It complements ChIP-seq by linking genomic occupancy to complex composition without genetic tagging or overexpression.
Typical Scientific Questions RIME-MS Can Answer
- Which cofactors co-assemble with my transcription factor on chromatin?
- How do interaction partners change across conditions or genotypes?
- Which interactors are high-confidence versus background associations?
- Do partners indicate known complexes or novel assemblies worth validation?
- Which chromatin remodelers or epigenetic regulators co-purify with the bait?
- How do PTMs or domain mutations rewire the native interactome?
- Which candidates best align with pathway signals from RNA-seq/ATAC-seq?
- What short list should I prioritize for orthogonal follow-ups?
Advantages of Our RIME-MS Service
Endogenous, Tag-Free Capture — Preserve Native Biology
No overexpression or epitope tags; complexes are profiled in situ with intact PTMs and stoichiometry.
High-Confidence IDs — 1% FDR at PSM/Peptide/Protein Levels
Target–IgG controls, decoy databases, and common-contaminant filters keep false discoveries at or below 1%.
Reproducible Quantitation — CV ≤10–15% Across Replicates
LFQ/TMT pipelines with median normalization and batch correction deliver stable interactor intensities suitable for condition comparisons.
Low Background Pull-downs — Matched Controls to Reduce Nonspecifics
Stringent wash schemes and IgG/bead-only controls suppress carryover; enrichment scores highlight true interactors over background.
Nuclear Focus, Chromatin Ready — Optimized for Transcriptional Complexes
Crosslinking, nuclear isolation, and on-bead digestion are tuned for chromatin-anchored assemblies and transient interactions.
Sensitive Detection — Broad Dynamic Range for Low-Abundance Partners
High-resolution Orbitrap acquisition and optimized gradients improve coverage of weakly represented nuclear cofactors.
Technical Services
Service Scope
Workflow and Instrumentation
Input Requirements
Deliverables
Use Cases
How to Choose
FAQ
Get a Custom Proposal
RIME-MS Services: What We Offer at Creative Proteomics
Condition-Specific Interaction Mapping
Compare interaction networks across treatments, genotypes, or time points to reveal dynamic rewiring of transcriptional assemblies.
Epigenetic Complex Characterization
Resolve the composition of histone-modifying and chromatin-remodeling complexes involved in gene activation or repression.
Novel Partner Discovery for Regulatory Proteins
Uncover previously unknown interactors of transcription factors or nuclear signaling proteins to support mechanistic insights.
Drug Target and Mode-of-Action Validation
Analyze how candidate compounds alter chromatin-bound protein complexes, aiding in early-stage pharmacology studies.
Tissue- or Species-Specific Complex Mapping
Profile nuclear interactomes in primary tissues or across species to explore context-specific regulatory mechanisms.
Multi-Omics Integration Support
Provide protein interaction context for RNA-seq, ATAC-seq, or phosphoproteomic data to strengthen regulatory interpretations
Step-by-Step Workflow for RIME-MS
1
Define Target & Controls
Select bait, validated antibody, IgG/bead controls; set contrasts and replicates.
2
Crosslink & Enrich Nuclei
Stabilize chromatin assemblies; isolate nuclei under gentle conditions.
3
Endogenous IP
Pull down native complexes; add reference peptides if quantitation is needed.
4
Wash & On-Bead Digest
Remove nonspecifics; reduce/alkylate and digest to MS-ready peptides.
5
NanoLC–Orbitrap MS/MS
High-accuracy acquisition with broad dynamic range.
6
ID, Quant & QC
Database search with decoys; control FDR; report calibration and replicate metrics.
7
Background Subtraction & Scoring
Subtract controls, rank by enrichment/reproducibility, flag contaminants.
8
Pathway/Network Annotation
Map confident interactors to complexes and regulatory pathways.
9
Decision-Ready Report
Editable tables, plots, network views, and a prioritized validation shortlist.
Instrumentation & Platform Capabilities for RIME-MS
- Separation & MS: NanoLC with low-dispersion flow and cooled autosampler + high-resolution Orbitrap (≤3 ppm, up to 120k); optional FAIMS for low-abundance gains.
- Acquisition Modes: DDA for discovery, DIA for robust quant, PRM for targeted verification; 60–120 min gradients balance depth and reproducibility.
- Quantitation: Label-free LFQ (default), optional TMT multiplexing; practical linear dynamic range ~4–5 orders of magnitude.
- Quality Control: iRT retention tracking; mass calibration per batch; FDR ≤1% at PSM/protein; IgG/blank controls and spike-in recovery (typ. 80–120%).
- Background Handling: Control-based background subtraction, reproducibility scoring, and contaminant filtering (CRAPome-style).
Orbitrap Exploris 480
Q Exactive HF-X
Sample Requirements for HDX-MS Projects
| Item | Accepted Matrices | Input Guideline | re-Analytical State | Storage & Logistics | Provide with Submission |
| Cultured cells | Adherent or suspension mammalian cells (cell lines or primary) | ≥ 1–2 × 107 cells per IP condition | Mild formaldehyde crosslinking; harvest as clean nuclear pellets; avoid harsh detergents/SDS | Snap-freeze pellets; ship on dry ice | Cell ID, passage, medium/treatments; crosslink and lysis buffer recipes |
| Tissue samples | Fresh-frozen mammalian tissues | ≥ 200–300 mg per condition | Finely minced; optionally crosslinked before freezing; minimize blood content | Store at –80 °C; ship on dry ice | Species, tissue/region, collection method, pre-processing notes |
| Nuclear extracts (optional) | Nuclei isolated from cells or tissues | From ≥ 1 × 107 starting cells | Crosslinked nuclei; gentle lysis to preserve complexes | Freeze aliquots with protease inhibitors; dry ice shipping | Nuclei isolation protocol; estimated DNA/protein content |
| Client-prepared IP lysates (optional) | Crosslinked nuclear lysates | ≥ 1 mg total protein per IP | Not boiled; detergent level compatible with IP (no SDS) | Freeze immediately after preparation; dry ice | Full lysis/IP buffer composition and steps taken |
| Antibody | Target-specific, IP-validated antibody | ≥ 10–20 μg per IP (or provide catalog/clone) | Recognizes native epitope; avoid antibodies requiring denaturation | Ship chilled/frozen per datasheet | Datasheet/clone, lot, species, epitope info; suggested bead type |
Notes
- If antibody performance in IP is unknown, request a pilot pull-down prior to full RIME-MS.
- For integrative projects (e.g., with ChIP-seq/ATAC-seq), align conditions and controls across assays.
- RUO only; no clinical or diagnostic use.
Deliverables: What You Get from Our HDX-MS Service
- Interactor List — High-confidence proteins with UniProt IDs, gene names, and enrichment scores.
- Quant Comparison — Fold change tables and volcano plots (if multiple conditions are analyzed).
- Functional Annotation — GO terms, complex membership (CORUM), and pathway enrichment (Reactome).
- Network Visualization — Editable interaction maps in Cytoscape-ready formats (.SIF/.XLSX).
- Raw Data Files — LC–MS/MS files (.RAW and .mzML) for reprocessing or archival.
- Processed Outputs — Peptide and protein ID tables (.XLSX/.TXT), background-filtered candidate lists.
- QC Report — Run summaries including FDR, replicate correlation, and calibration metrics.
- Method Overview — Summary of IP conditions, antibodies used, controls, and sample handling.
Differential Interactome Volcano Plot
Condition-Resolved Heatmap with Clustering
Interaction Network Map with Complex Annotations
Orthogonal Validation Panel (Multi-modal)
RIME-MS Use Cases
Decode TF–cofactor logic on chromatin
Clarify activator/repressor assemblies underlying expression programs.
Reveal condition-specific rewiring
See which partners appear, disappear, or shift with treatment or genotype.
Prioritize functional candidates
Shortlist interactors that align with pathways or phenotypes for follow-up tests.
Assess variant or domain effects
Determine how mutations/domains reshape nuclear complex composition.
Contextualize multi-omics signals
Anchor RNA-seq/ATAC-seq findings with direct interaction evidence.
Differentiate core vs context-dependent partners
Separate stable complex members from state-specific adapters.
Support mechanism-of-action studies
Link compound exposure to changes in chromatin-bound assemblies.
Map tissue/lineage specificity
Compare interactomes across primary tissues or developmental states.
RIME-MS Compared with AP-MS, BioID, Co-IP, XL-MS, and ChIP–MS
Capability Matrix (✔︎ = strong, ○ = partial/possible, — = limited)
| Decision factor | RIME-MS | AP-MS (tagged) | BioID/TurboID | Co-IP MS (endogenous) | XL-MS | ChIP–MS / ChIP-SICAP |
| Endogenous (no genetic tag) | ✔︎ | — | — | ✔︎ | ✔︎/○ | ✔︎ |
| Chromatin-anchored complexes | ✔︎ | ○ | — | ○ | ○ | ✔︎ (locus-linked) |
| Captures transient/weak partners | ✔︎ (crosslink-stabilized) | — | ✔︎ (time-windowed proximity) | — | ○ | ○ |
| Locus-specific view (factor-bound DNA) | ○ | — | — | — | — | ✔︎ |
| Live-cell spatial neighborhood | — | — | ✔︎ | — | — | — |
| Direct contact restraints / topology | — | — | — | — | ✔︎ | — |
| High-yield stable complex mapping | ○ | ✔︎ | ○ | ✔︎ (strong binders) | — | — |
| Artifact risk (tag/overexpression) | Low | Medium–High | Medium | Low | Low–Medium (crosslinking) | Medium (antibody dependency) |
| Primary inputs typically accepted | Crosslinked cells/tissues | Edited cell lines | Edited cell lines | Cells/tissues | Enriched complexes | Crosslinked chromatin |
How to choose
- "I need endogenous nuclear partners with chromatin context." → RIME-MS
- "I have a clean tagged bait and want broad, stable partners fast." → AP-MS (tagged)
- "I suspect weak, transient, or membrane-adjacent neighbors in live cells." → BioID/TurboID
- "I just need to confirm a few strong hits from discovery." → Co-IP MS (endogenous)
- "I need distance constraints or topology within a complex." → XL-MS
- "Who co-exists with my factor on DNA at its bound loci?" → ChIP–MS / ChIP-SICAP
Practical pairings that work
- RIME-MS + ChIP-seq: connect who binds with where it binds.
- RIME-MS → Co-IP MS: orthogonally confirm the strongest candidates without crosslinking.
- BioID/TurboID → RIME-MS: refine broad proximity hits to chromatin-anchored interactors.
- RIME-MS + XL-MS: move from membership to contact-level evidence.
You May Want to Know
What is RIME-MS used for?
Profiling endogenous protein complexes on chromatin—especially transcription factor/cofactor assemblies—by combining antibody pull-down of fixed nuclei with high-resolution MS; it naturally complements ChIP-seq to link occupancy and complex composition.
Does RIME-MS require genetic tagging or overexpression?
No—targets are enriched at endogenous levels, reducing tag- or overexpression-driven artifacts seen in some AP-MS designs.
Can RIME-MS detect weak or transient interactions?
Yes; formaldehyde crosslinking stabilizes short-lived chromatin assemblies prior to IP, improving capture of low-affinity partners.
How is RIME-MS different from BioID/TurboID proximity labeling?
RIME-MS enriches chromatin-anchored complexes around an antibody-defined bait without genetic editing, whereas BioID/TurboID biotinylates proteins in a spatial neighborhood of a tagged enzyme in live cells; use BioID/TurboID when live-cell spatial reach is the priority.
How does RIME-MS compare with ChIP–MS/ChIP-SICAP?
RIME-MS maps bait-centred nuclear complexes globally; ChIP-MS/ChIP-SICAP reports proteins co-occupying factor-bound DNA and tends to return cleaner, locus-linked proteomes at the cost of breadth.
Can RIME-MS be quantitative across conditions?
Yes; multiplexed adaptations (e.g., TMT-based qPLEX-RIME) enable condition-resolved comparisons of chromatin-associated complexes.
What controls strengthen RIME-MS specificity?
Matched IgG or bead-only controls and condition-matched inputs support background modeling and enrichment scoring in downstream analysis.
Which samples are suitable?
Crosslinked cultured cells or tissues prepared for nuclear enrichment are typical inputs; maintain IP-compatible buffers to preserve complexes.
What outputs should I expect?
Background-filtered interactor tables with identifiers, confidence metrics, and functional/network annotations; optional quantitative contrasts when designed upfront.
When is RIME-MS not the best fit?
If the question centers on live-cell spatial proximity or membrane-adjacent neighborhoods, prefer proximity labeling; for direct distance restraints within a complex, consider XL-MS.