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MDC1 / Mediator of DNA damage checkpoint protein 1 · IHC design guide
Design Immunohistochemistry for MDC1
Plan MDC1 chromogenic IHC in paraffin sections around generally nuclear tissue staining (HPA tissue IHC). Use the catalog antibody's 0.5–1 μg/mL IHC range (datasheet A01252-2) and consider damage-associated nuclear foci when interpreting staining (UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
MDC1 Immunohistochemistry Experimental Design Guide
Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before staining
| Expected localisation | Nuclear; chromatin-associated (HPA tissue IHC; UniProt) | |
| Staining pattern | General nuclear staining across tissue cell types (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6 HIER, heat-mediated (datasheet A01252-2) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | None in HPA (detected in all 45 tissues); use no-primary + isotype controls |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A01252-2) | |
| Caveat | Alveolar type I cells show low staining (HPA tissue IHC) | |
| Regulation | DNA damage redistributes MDC1 to nuclear foci (UniProt) | |
| Isoform / epitope | 4 isoforms; antibody epitope coverage is unspecified (UniProt; datasheet A01252-2) |
Section 1
Recommended MDC1 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol with two published MDC1 protocols using formalin-fixed, paraffin-embedded sections (PMC4446443; PMC4515811).
| Sample | Paraffin-embedded human colon cancer tissue; fixative not specified (datasheet A01252-2) |
| Fixation | Image fixative and duration unreported (datasheet A01252-2); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: Citrate pH 6, 20 min (datasheet A01252-2) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A01252-2) |
| Primary antibody | Rabbit anti-MDC1, 0.5-1μg/ml (datasheet A01252-2) |
| Primary incubation | Overnight at 4 °C (datasheet A01252-2) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A01252-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | MDC1-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: General nuclear expression. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated citrate retrieval at pH 6 (datasheet: A01252-2); PMC4515811 reports boiling for retrieval without specifying a buffer.
Section 2
What Is the Expected MDC1 Staining Pattern?
MDC1 should stain nuclei across many cell types: HPA reports general nuclear expression, with high staining in several glandular, hematopoietic, respiratory epithelial and glial cell populations (HPA tissue IHC). Its chromatin association and lack of a transmembrane segment support a nuclear, rather than membrane, pattern (UniProt Q14676 topology and subcellular location). HPA rates the tissue pattern Enhanced, while noting medium consistency with RNA expression (HPA tissue IHC).
| Nuclear staining in adrenal glandular cells, bone marrow hematopoietic cells or bronchial respiratory epithelial cells. | This matches cell populations scored High by HPA; compare signal within the named population rather than treating every cell in the section as equally positive (HPA tissue IHC). |
| Predominantly membrane, extracellular or broad cytoplasmic staining with little nuclear signal. | Treat this as a localisation mismatch and investigate artefact or nonspecific binding: MDC1 is chromatin-associated and has no transmembrane segment (UniProt Q14676 topology and subcellular location). |
| Strong staining chiefly in lung alveolar type I cells or heart cardiomyocytes. | These cells are scored Low by HPA; disproportionate signal raises concern for cross-reactivity or endogenous detection activity, but Low does not mean absent (HPA tissue IHC; general IHC practice). |
| Diffuse colour over tissue, empty spaces and multiple cell compartments. | A slide-wide deposit cannot establish MDC1 localisation; assess nonspecific background and detection chemistry with appropriate controls (UniProt Q14676 subcellular location; general IHC practice). |
| No nuclear signal in an HPA High cell population. | First suspect a failed staining run or unsuitable antibody conditions; absence in one section alone does not establish biological loss of MDC1 (HPA tissue IHC; general IHC practice). |
💡Expected MDC1 appearanceCall a section positive when the appropriate cells show clear nuclear staining, potentially High in HPA High populations; strong membrane or diffuse extranuclear colour is suspect (HPA tissue IHC; UniProt Q14676 subcellular location).
| Reference cell population | HPA scores adrenal glandular, bone marrow hematopoietic and bronchial respiratory epithelial cells High, but lung alveolar type I cells and cardiomyocytes Low; choose comparisons by cell type (HPA tissue IHC). |
| Damage response | MDC1 can redistribute into discrete nuclear foci after DNA damage through phosphorylated H2AX; foci are context dependent, so their absence in routine tissue IHC does not invalidate nuclear staining (UniProt Q14676 subcellular location). |
| Antibody validation | HPA lists HPA006915 and CAB080116 as IHC Enhanced; that supports their reported tissue patterns, but each staining run still needs its own controls (HPA antibodies; general IHC practice). |
| Isoforms and epitope | UniProt lists 4 isoforms and a full-length chain spanning 1–2089; the supplied record gives no antibody epitope, so isoform coverage cannot be predicted (UniProt Q14676 isoforms and processing). |
| Retrieval and detection | Follow the IHC-validated antibody's documented conditions and use suitable detection controls; no MDC1-specific retrieval requirement or fixation sensitivity is established by these sources (general IHC practice; supplied UniProt Q14676 and HPA records). |
| IF/ICC Q&A: Where should MDC1 appear? | In the nucleoplasm and nuclear bodies; HPA supports both locations in ICC-IF, while UniProt also describes damage-associated nuclear foci (HPA subcellular ICC-IF; UniProt Q14676 subcellular location). |
| Situation | Likely cause | Next action |
|---|---|---|
| A reference section lacks nuclear staining. | The run or antibody conditions may have failed; HPA reports High staining in specific reference cell populations (HPA tissue IHC; general IHC practice). | Check a known-positive cell population and run controls, then review the IHC-validated antibody's documented conditions (HPA tissue IHC; general IHC practice). |
| Signal is mainly cytoplasmic or membranous. | The compartment conflicts with MDC1's nuclear, chromatin-associated localisation (UniProt Q14676 topology and subcellular location). | Compare with an HPA High nuclear reference and check staining controls for nonspecific signal (HPA tissue IHC; general IHC practice). |
| Low-scored cells appear stronger than nearby reference cells. | Cross-reactivity or endogenous detection activity is possible; HPA scores alveolar type I cells and cardiomyocytes Low (HPA tissue IHC; general IHC practice). | Identify the stained cell types, inspect reagent controls and compare with an IHC Enhanced antibody where available (HPA tissue IHC and antibodies; general IHC practice). |
| Colour is widespread outside nuclei. | Background or endogenous detection activity may obscure a nuclear pattern (UniProt Q14676 subcellular location; general IHC practice). | Review negative detection controls, blocking and chromogen development, then reassess nuclear localisation (general IHC practice). |
| Nuclear staining is present but discrete foci are absent. | UniProt describes foci following DNA damage; routine tissue staining need not show a visible focus pattern (UniProt Q14676 subcellular location). | Score the nuclear signal against the relevant HPA cell population; reserve a focus-specific interpretation for an appropriate damage context (HPA tissue IHC; UniProt Q14676 subcellular location). |
| Two antibodies give different nuclear intensities. | The supplied records do not identify their epitopes or establish equal isoform recognition; HPA validation does not guarantee identical intensity (UniProt Q14676 isoforms; HPA antibodies). | Check each antibody's documented IHC conditions and compare compartment and cell-type patterns using the same reference tissue (HPA tissue IHC and antibodies; general IHC practice). |
Experiment evidence
IHC & IF images showing MDC1 staining
A01252-2 · IHCIHC analysis of MDC1 using anti-MDC1 antibody (A01252-2). MDC1 was detected in paraffin-embedded section of human colon cancer tissue. Heat
A01252-2 · IHCIHC analysis of MDC1 using anti-MDC1 antibody (A01252-2). MDC1 was detected in paraffin-embedded section of human lung cancer tissue. Heat m
A01252-2 · IFIF analysis of MDC1 using anti-MDC1 antibody (A01252-2). MDC1 was detected in immunocytochemical section of A431 cells. Enzyme antigen retri
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000137337-MDC1/tissue/cerebral+cortex#imid_2296968
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000137337-MDC1/tissue/colon#imid_2296902
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000137337-MDC1/tissue/liver#imid_2296911
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000137337-MDC1/tissue/kidney#imid_2296896
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA006915; green = MDC1, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA006915; green = MDC1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for MDC1 IHC & IF
🧪Run adrenal gland first: its glandular cells should show MDC1 staining (HPA: High in adrenal gland glandular cells). HPA detects MDC1 in all 45 scored tissues, so use no-primary and isotype controls for the negative comparison (HPA: no negative tissue); cells on the positive slide without nuclear signal should remain at background and should not be designated a known MDC1-negative population (UniProt Q14676: nuclear localization).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: None in HPA: MDC1 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead.
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show MDC1 in A-431, U-251MG, U2OS, SiHa, with annotated localisation: Nucleoplasm (supported), Nuclear bodies (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control, a rabbit IgG isotype control matched to the primary concentration, and MDC1-knockout material as a biological negative (caption: rabbit primary and goat anti-rabbit IgG secondary; standard IHC practice). For adrenal gland, check endogenous peroxidase and biotin background before interpreting DAB signal from the biotin-based detection system (caption: SABC with DAB; standard IHC practice).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the selected A01252-2 paraffin-section caption does not state the fixative (caption: fixative unreported). That caption reports citrate retrieval at pH 6 for 20 minutes, but does not establish how strongly MDC1 staining depends on retrieval; HPA ICC-IF images support nuclear localization checks, while the supplied evidence does not establish that frozen sections are easier (caption: retrieval conditions; HPA: nucleoplasm and nuclear bodies). With SABC/DAB detection, endogenous biotin or peroxidase background is a practical concern in the adrenal gland positive tissue (caption: SABC/DAB; standard IHC practice).
HPA tissue IHC evidence for MDC1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: MDC1 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced MDC1 IHC Tips
Use nuclear localisation and matched controls to evaluate MDC1 staining in paraffin sections (UniProt Q14676; HPA tissue IHC).
Which retrieval conditions should I start with for MDC1 in paraffin sections?
Start with heat-mediated retrieval in citrate buffer at pH 6 for 20 minutes (datasheet A01252-2; selected IHC caption). Keep section thickness, heating, cooling and subsequent staining conditions matched across comparison slides so retrieval differences do not become apparent expression differences (standard IHC practice). The selected image used 1 μg/mL primary antibody overnight at 4°C after retrieval, providing a documented starting point for this antibody (selected IHC caption). If nuclear staining remains weak, optimise retrieval duration on control sections while checking tissue integrity and background; document any change before comparing samples (standard IHC practice).
How should I handle fixation when MDC1 nuclear staining is weak?
The selected paraffin-section caption does not state a fixative, and target-specific MDC1 sensitivity to fixation is unknown from the supplied evidence (selected IHC caption). Record the fixative and fixation interval for each specimen, then compare sections processed together before attributing a weak signal to biological variation (standard IHC practice). Start troubleshooting with the documented citrate pH 6, 20-minute retrieval and 1 μg/mL primary antibody conditions, using a matched positive control (selected IHC caption; standard IHC practice). If fixation histories differ, interpret intensity comparisons cautiously and assess nuclear morphology alongside staining; the HPA tissue pattern does not establish a fixation effect (standard IHC practice; HPA tissue IHC).
What MDC1 staining pattern should I expect in chromogenic IHC?
Expect predominantly nuclear staining because MDC1 is associated with chromatin, while HPA describes general nuclear expression in tissue (UniProt Q14676 subcellular; HPA tissue IHC). Distinguish diffuse nuclear signal from discrete nuclear foci where morphology permits; MDC1 relocalises to foci after DNA damage through phosphorylated H2AX binding (UniProt Q14676 subcellular and function). A chromogenic section may not resolve individual foci reliably, so score clearly identifiable nuclei and confirm a focal pattern with an appropriate higher-resolution assay when it matters (standard IHC practice). Predominantly cytoplasmic or extracellular colour should prompt checks of counterstain, detection background and section quality before assignment to MDC1 (UniProt Q14676 subcellular; standard IHC practice).
Can this IHC result distinguish MDC1 isoforms or phosphorylation states?
MDC1 has 4 listed isoforms and numerous modified residues, including phosphorylation sites, but the supplied antibody evidence does not map its epitope (UniProt Q14676 isoforms and modified residues; selected IHC caption). Treat chromogenic nuclear staining as antibody-detected MDC1 signal rather than an isoform-specific or phosphorylation-specific readout unless separate epitope validation establishes that claim (standard IHC practice). The FHA domain spans residues 54–105, and the two BRCT domains lie near the C terminus; neither location identifies what this antibody recognises (UniProt Q14676 domains). For comparisons involving DNA damage, use separately validated state-specific markers and matched controls instead of inferring MDC1 modification from stain intensity alone (standard IHC practice).
How can I follow up an IHC nuclear pattern with multiplex IF?
Use IF as a separate follow-up to chromogenic IHC, with independent validation of antibody performance in that application (standard IF practice). Pair MDC1 with a validated marker for the cell population being assessed and a nuclear counterstain, then confirm that the apparent signal lies within nuclei (UniProt Q14676 subcellular; standard IF practice). Choose spectrally separated fluorophores after inspecting tissue autofluorescence in an unstained section, and include single-stain controls when comparing channels (standard IF practice). Because MDC1 is nuclear and has no transmembrane segment, optimise permeabilisation for access to a nuclear epitope while preserving nuclear structure and potential foci; the supplied IHC caption does not specify IF conditions (UniProt Q14676 topology and subcellular; selected IHC caption; standard IF practice).
How do I distinguish MDC1 signal from chromogenic background?
Assess whether DAB colour remains confined to intact nuclei, the expected compartment for MDC1, and inspect unstained or primary-omission controls for detection-system deposits (UniProt Q14676 subcellular; standard IHC practice). The selected image used 10% goat serum block, biotinylated goat anti-rabbit secondary antibody for 30 minutes at 37°C, and a streptavidin-biotin complex with DAB (selected IHC caption). For this chromogenic workflow, include a peroxidase block and evaluate any endogenous biotin contribution with suitable controls; these are general detection checks, not MDC1-specific findings (standard IHC practice). If colour spreads across stroma or section edges, optimise washing and antibody concentration on matched sections before interpreting nuclear intensity (standard IHC practice).
What should I score when comparing MDC1 across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the analysis region and cell population before scoring, and count only evaluable nuclei with interpretable chromogenic signal (standard IHC practice; UniProt Q14676 subcellular). Report the percentage of positive nuclei and, when intensity is reproducible, an H-score that combines intensity categories with their corresponding cell percentages (standard IHC practice). Normalise counts to the total number of evaluable nuclei in the same annotated compartment; for spatial counts, report positive nuclei per mm² of viable analysed tissue (standard IHC practice). Keep retrieval, detection, counterstain and scoring thresholds consistent across samples, and record excluded necrotic or damaged areas so differences are not driven by tissue quality (standard IHC practice).
When is apparent MDC1 positivity likely to be artefactual?
Give most weight to staining within intact nuclei in the intended cell population, consistent with MDC1 chromatin association and HPA general nuclear expression (UniProt Q14676 subcellular; HPA tissue IHC). Compare that pattern with a matched control section and review cell identity: HPA reports high glandular-cell staining in the appendix and high hematopoietic-cell staining in bone marrow (HPA tissue IHC; standard IHC practice). Diffuse cytoplasmic colour, staining restricted to cut edges or necrotic areas, and signal in a primary-omission control warrant investigation before calling a positive result (UniProt Q14676 subcellular; standard IHC practice). Check endogenous peroxidase and detection background, and avoid interpreting nuclear intensity alone as evidence of DNA damage or repair activity (standard IHC practice; UniProt Q14676 function).
Boster reagents
Best MDC1 / Mediator of DNA damage checkpoint protein 1 IHC Antibodies
A01252-2 has real IHC data from human paraffin sections of colon cancer, lung cancer and testis, plus IF/ICC data from A431 cells (catalog image captions; catalog: Human reactivity).
Real IHC data
Anti-MDC1 Antibody
Cat # A01252-2
A01252-2 is listed for human IHC and IF/ICC (catalog: applications and reactivity). Its images show MDC1 staining in paraffin sections of human colon cancer, lung cancer and testis, and in A431 cells by IF/ICC (catalog image captions).
Which to pick: Choose A01252-2 for paraffin-section IHC: its tissue captions report citrate retrieval at pH 6 and 1 μg/ml primary antibody, but do not report the fixative (A01252-2 IHC image captions). The same SKU is listed for IF/ICC and has an A431 cell IF image at 2 μg/ml (catalog: applications; A01252-2 IF image caption). No cross-species choice is documented; the catalog lists human reactivity only, and clonality is unreported (catalog: reactivity and clone fields).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q14676 (MDC1_HUMAN, Mediator of DNA damage checkpoint protein 1).
- Human Protein Atlas. MDC1 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. MDC1 subcellular location (ICC-IF): Localized to the nucleoplasm and nuclear bodies..
- Human Protein Atlas. MDC1 antibody validation summary (3 antibodies).
- SOX9 activity is induced by oncogenic Kras to affect MDC1 and MCMs expression in pancreatic cancer. Oncogene 2018 — PMC6545484.
- Development of serous ovarian cancer is associated with the expression of homologous recombination pathway proteins. Pathology oncology research : POR 2014 — PMC4177045.
- MDC1 functionally identified as an androgen receptor co-activator participates in suppression of prostate cancer. Nucleic acids research 2015 — PMC4446443.
- MDC1 Enhances Estrogen Receptor-mediated Transactivation and Contributes to Breast Cancer Suppression. International journal of biological sciences 2015 — PMC4515811.
- PubMed PMID:8724849 — UniProt-cited evidence.
- PubMed PMID:17974005 — UniProt-cited evidence.
- PubMed PMID:16702430 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for MDC1 (Mediator of DNA damage checkpoint protein 1). Expected localisation Nuclear; chromatin-associated (HPA tissue IHC; UniProt). Antigen retrieval: Citrate pH 6 HIER, heat-mediated (datasheet A01252-2). Validated IHC antibody: A01252-2 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative None in HPA (detected in all 45 tissues); use no-primary + isotype controls. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human colon cancer tissue; fixative not specified (datasheet A01252-2); 2. Fixation: Image fixative and duration unreported (datasheet A01252-2); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat retrieval: Citrate pH 6 , 20 min (datasheet A01252-2); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A01252-2); 8. Primary antibody: Rabbit anti-MDC1, 0.5-1μg/ml (datasheet A01252-2); 9. Primary incubation: Overnight at 4 °C (datasheet A01252-2). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A01252-2); Caveat: Alveolar type I cells show low staining (HPA tissue IHC); Isoform / epitope: 4 isoforms; antibody epitope coverage is unspecified (UniProt; datasheet A01252-2); Regulation: DNA damage redistributes MDC1 to nuclear foci (UniProt). Evidence: UniProt Q14676, HPA tissue IHC · Enhanced, HPA ICC-IF · nucleoplasm and nuclear bodies., Datasheet A01252-2, PMC4446443, PMC4515811. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/MDC1/immunohistochemistry-ihc-guide.