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MED15 / Mediator of RNA polymerase II transcription subunit 15 · IHC design guide
Design Immunohistochemistry for MED15
Plan MED15 staining in paraffin sections using IHC-validated antibody A03568-1 (datasheet A03568-1). Assess cytoplasmic and nuclear staining while allowing for tissue-dependent intensity (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
MED15 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 | Cytoplasmic and nuclear staining (HPA tissue IHC) | |
| Staining pattern | Ubiquitous cytoplasmic and nuclear staining (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6 HIER, heat-mediated (datasheet A03568-1) | |
| Positive control | Adipose tissue+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 | Keep fixation conditions consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Salivary glandular cells show low staining (HPA tissue IHC) | |
| Regulation | Expression varies by tissue (UniProt) | |
| Isoform / epitope | 3 isoforms; epitope coverage is unreported (UniProt) |
Section 1
Recommended MED15 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol (datasheet A03568-1) with published MED15 IHC methods for liver, prostate and bladder cancer specimens (PMC5844107; PMC5352374; PMC6096071).
| Sample | Paraffin-embedded human colon cancer tissue; fixative not specified (datasheet A03568-1) |
| Fixation | Image fixative and duration unreported (datasheet A03568-1); 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 A03568-1) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A03568-1) |
| Primary antibody | Rabbit anti-MED15, 0.5-1μg/ml (datasheet A03568-1) |
| Primary incubation | Overnight at 4 °C (standard) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A03568-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | MED15-positive staining in adipocytes of adipose tissue (HPA tissue IHC: High). HPA tissue profile: Ubiquitous cytoplasmic and nuclear expression. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated citrate pH 6 retrieval (datasheet A03568-1). The HCC study specifies citrate retrieval but gives no pH or heating conditions (PMC5844107).
Section 2
What Is the Expected MED15 Staining Pattern?
MED15 should show cytoplasmic and nuclear staining across many cell types in paraffin sections (HPA tissue IHC: ubiquitous cytoplasmic and nuclear expression). Nucleoplasmic localisation is supported by ICC-IF (HPA subcellular). This distribution fits a protein found in the cytoplasm and nucleus with no transmembrane segment (UniProt Q96RN5). Interpret intensity by cell type: HPA rates its tissue IHC evidence Approved, with medium consistency between staining and RNA expression (HPA tissue IHC).
| Nuclear and cytoplasmic staining appears in several cell types; adipocytes or bone marrow hematopoietic cells stain strongly. | This matches the broad tissue pattern, including High staining in those cells (HPA tissue IHC). Assess nuclear and cytoplasmic signal separately; a single field need not reproduce every tissue's intensity (HPA tissue IHC: levels vary by cell type). |
| Signal is confined to the plasma membrane or extracellular material, without convincing intracellular staining. | That distribution conflicts with the reported cytoplasmic and nuclear IHC pattern (HPA tissue IHC), supported nucleoplasmic IF location (HPA subcellular), and lack of a transmembrane segment (UniProt Q96RN5). Treat it as suspect staining and review the detection controls. |
| Only one unexpected cell population stains while neighboring cells show no signal. | A sharply restricted pattern warrants scrutiny because HPA describes ubiquitous expression and supplies no negative tissue in this record (HPA tissue IHC). Cross-reactivity or endogenous detection activity are possibilities, not diagnoses; compare a known-positive section and the appropriate detection control (general IHC practice). |
| Brown precipitate is diffuse across cells, stroma, or the whole section, obscuring compartment boundaries. | Unstructured staining cannot establish MED15 localisation against the nuclear and cytoplasmic reference pattern (HPA tissue IHC). Background may arise from nonspecific binding, detection chemistry, or inadequate washing; assess a matched control before scoring cells (general IHC practice). |
| A known-positive tissue has no convincing cellular signal. | Absence of signal in a cell type rated High, such as bone marrow hematopoietic cells, conflicts with the HPA reference image summary (HPA tissue IHC). First check that the assay and controls worked; a negative slide alone does not establish absent MED15 protein (general IHC practice). |
💡Expected MED15 appearanceCall a result positive when identifiable cells show nuclear and cytoplasmic chromogenic signal, with strong examples in HPA High cell types; isolated membrane or extracellular staining is suspect (HPA tissue IHC; UniProt Q96RN5 topology).
| Tissue and cell selection | HPA lists High staining in adipocytes, bone marrow hematopoietic cells, and several epithelial or neural cell groups, but Low staining in salivary gland glandular cells (HPA tissue IHC). Choose a documented high cell type when checking assay performance. |
| IHC antibody evidence | HPA003179 is IHC Approved; the tissue profile has medium consistency with RNA data (HPA antibodies; HPA tissue IHC). This supports comparison with the reported pattern, while leaving ambiguous or discrepant staining for control-based review. |
| Subcellular interpretation | UniProt lists both cytoplasm and nucleus (UniProt Q96RN5); HPA tissue IHC likewise reports both compartments (HPA tissue IHC). A nuclear signal is therefore expected, while HPA ICC-IF more specifically supports nucleoplasmic localisation (HPA subcellular). |
| IF/ICC: where should MED15 appear? | Expect nucleoplasmic fluorescence as the supported main location (HPA subcellular). HPA003179 has ICC Supported status (HPA antibodies). Interpret that evidence on the separate IF/ICC page; it does not define a paraffin IHC retrieval or staining condition. |
| Isoforms and epitope coverage | UniProt lists 3 isoforms (UniProt Q96RN5). The supplied records do not locate the antibody epitope or establish isoform-specific staining, so a difference in signal cannot be assigned to an isoform from these data. |
| Fixation and retrieval | No MED15-specific fixation sensitivity or retrieval requirement is supplied. If optimisation is needed, vary retrieval against a documented positive tissue and compare controls (general IHC practice); do not infer an effect from HPA staining levels or UniProt topology. |
| Situation | Likely cause | Next action |
|---|---|---|
| No signal in a documented high cell type | An assay or detection failure is possible when HPA reports High staining for that cell type (HPA tissue IHC; general IHC practice). | Verify tissue identity, antibody application, detection reagents, and the positive control; then review the assay's existing retrieval conditions (general IHC practice). Do not conclude that MED15 is absent from one failed slide. |
| Weak signal in salivary gland glandular cells | HPA rates this cell type Low, so weak staining alone can fit the reference pattern (HPA tissue IHC). | Compare a documented High cell type before changing assay conditions (HPA tissue IHC; general IHC practice). Score the salivary cells against local background rather than treating them as a negative control. |
| Diffuse stain prevents nuclear scoring | Nonspecific binding, incomplete washing, or detection background may obscure the expected compartments (general IHC practice; HPA tissue IHC: nuclear and cytoplasmic pattern). | Inspect a matched detection control, check blocking and wash steps, and adjust antibody concentration within the assay's validated range (general IHC practice). Score MED15 only when cell boundaries and nuclei remain interpretable. |
| Membrane-only or extracellular signal dominates | This is discordant with cytoplasmic and nuclear localisation and the absence of a transmembrane segment (HPA tissue IHC; UniProt Q96RN5). | Compare with a documented positive section and a detection control; review staining distribution at the cellular level before calling it MED15 (general IHC practice). A chromogenic deposit alone does not identify the antigen. |
| Unexpected isolated cells stain strongly | Cross-reactivity or endogenous detection activity could mimic a restricted positive pattern (general IHC practice); HPA describes broad expression and provides no negative tissue here (HPA tissue IHC). | Check the same cell type in another field and a documented positive section. If using enzyme-based detection, inspect the appropriate endogenous-activity control (general IHC practice). |
| Nuclear and cytoplasmic signal differ from an IF image | Tissue IHC reports both compartments, whereas HPA ICC-IF identifies the nucleoplasm as its supported main location (HPA tissue IHC; HPA subcellular). | Judge the paraffin section against the tissue IHC pattern and its controls (HPA tissue IHC; general IHC practice). Use the separate IF/ICC guide for fluorescence interpretation; the IF location does not supply an IHC protocol. |
Experiment evidence
IHC & IF images showing MED15 staining
A03568-1 · IHCIHC analysis of MED15 using anti-MED15 antibody (A03568-1). MED15 was detected in paraffin-embedded section of human colon cancer tissue. He
A03568-1 · IHCIHC analysis of MED15 using anti-MED15 antibody (A03568-1). MED15 was detected in paraffin-embedded section of human lung cancer tissue. Hea
A03568-1 · IFIF analysis of MED15 using anti-MED15 antibody (A03568-1). MED15 was detected in immunocytochemical section of U20S cells. Enzyme antigen re
M03568 · IFIF analysis of MED15 using anti-MED15 antibody (M03568). MED15 was detected in immunocytochemical section of A431 cells. Enzyme antigen retr
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000099917-MED15/tissue/cerebral+cortex#imid_1036028
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000099917-MED15/tissue/colon#imid_1035946
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000099917-MED15/tissue/liver#imid_1035955
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000099917-MED15/tissue/kidney#imid_1035956
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA003179; green = MED15, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA003179; green = MED15, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for MED15 IHC & IF
🧪Run adipose tissue first and look for staining in adipocytes (HPA: High in adipocytes). HPA reports MED15 in all 45 scored tissues, so there is no supported negative tissue; no-primary and isotype controls carry the negative comparison, and any cells without nuclear signal on the positive slide are only a local background reference, not validated MED15-negative cells (HPA: no negative rows; HPA subcellular: nucleoplasm).
Positive control tissue: Adipose tissue (Adipocytes, HPA High)
Negative control tissue: None in HPA: MED15 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 MED15 in A-431, U-251MG, U2OS, KOLF2.1J, NIH 3T3, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and a rabbit IgG isotype control matched to the catalog primary’s format; use MED15 knockout material as a biological specificity control, or a peptide block if the immunizing peptide is available (caption: rabbit anti-MED15; standard IHC practice). Quench endogenous peroxidase and check for endogenous biotin when using the caption’s biotinylated secondary and SABC–DAB detection (caption: biotinylated goat anti-rabbit IgG, SABC and DAB; standard IHC practice).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the selected A03568-1 tissue-IHC caption does not state the fixative (caption: fixative unreported). The paraffin-section example uses heat retrieval in citrate buffer at pH 6 for 20 minutes; the supplied evidence does not establish that frozen sections or tissue IF are easier (caption: retrieval conditions; HPA: ICC-IF images in cell lines). Adipose sections can show lipid-extraction spaces that complicate interpretation of chromogenic signal (standard histology practice).
HPA tissue IHC evidence for MED15
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | High | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Breast | Adipocytes | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: MED15 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced MED15 IHC Tips
Troubleshoot MED15 staining in paraffin section IHC using the catalog image, HPA expression patterns, and MED15 localisation as reference points.
What retrieval should I try first when MED15 staining is weak?
Start with heat mediated citrate buffer at pH 6 for 20 minutes (datasheet A03568-1). The catalog image used this retrieval on a paraffin section of human colon cancer, although its fixative was not reported (datasheet A03568-1). If staining remains weak, compare a second retrieval condition on adjacent sections while keeping antibody concentration and detection constant (standard IHC practice). Include a known staining reference section in each run so a retrieval change can be distinguished from a failed detection step (standard IHC practice). Record nuclear and cytoplasmic signal separately because both are reported for MED15 (HPA: tissue IHC profile).
Could fixation explain weak or patchy MED15 staining?
Target specific MED15 sensitivity to fixation is unknown from the supplied evidence; the catalog paraffin image does not state a fixative (datasheet A03568-1). Check the specimen's recorded fixative and fixation duration before comparing staining across blocks (standard IHC practice). Process reference and test sections with the same citrate pH 6, 20 minute retrieval and detection conditions (datasheet A03568-1; standard IHC practice). If signal varies within a section, review tissue preservation and staining at cut edges before attributing the pattern to MED15 biology (standard IHC practice). Do not infer fixation sensitivity from MED15 localisation or modified residues (UniProt Q96RN5: localisation and modified residues).
How should I assess nuclear versus cytoplasmic MED15 staining?
Score nuclear and cytoplasmic DAB staining separately in intact cells (standard IHC practice). MED15 is reported in both nucleus and cytoplasm, while HPA tissue IHC describes ubiquitous cytoplasmic and nuclear expression (UniProt Q96RN5: subcellular location; HPA: tissue IHC profile). HPA cell imaging supports nucleoplasmic localisation, so a nuclear signal has an independent localisation reference (HPA: subcellular summary). Compare compartments in the same cell population and under the same counterstain and exposure conditions (standard IHC practice). Diffuse colour over tissue folds, empty spaces, or damaged cells should be reviewed as possible staining artefact before assigning a compartment (standard IHC practice).
Could MED15 isoforms or epitope accessibility change the IHC pattern?
MED15 has 3 annotated isoforms, but the supplied catalog caption does not identify the antibody's epitope or establish isoform selectivity (UniProt Q96RN5: isoforms; datasheet A03568-1). Treat staining as MED15 immunoreactivity unless an epitope map and appropriate validation support a narrower claim (standard IHC interpretation). MED15 has no annotated transmembrane segment, and its listed modifications include dimethylarginine at 349 and phosphothreonine at 603 (UniProt Q96RN5: topology and modified residues). Those annotations alone do not show whether this antibody detects either modified state (UniProt Q96RN5: modified residues; datasheet A03568-1). When samples disagree, compare epitope information and retrieval records before attributing the difference to an isoform (standard IHC practice).
How can IF help assess a disputed MED15 IHC pattern?
Use IF/ICC as an independent localisation check, with its own validated conditions rather than assuming that the paraffin section IHC conditions transfer (standard IF practice; datasheet A03568-1: tissue IHC caption). Multiplex MED15 with a marker for the cell population being assessed, and compare nuclear signal with the DNA counterstain (standard IF practice; HPA: nucleoplasmic localisation). Choose fluorophores and controls that separate specific signal from tissue autofluorescence, particularly when examining tissue sections (standard IF practice). For intracellular MED15, assess permeabilisation under the IF antibody's validated conditions so the epitope is accessible (UniProt Q96RN5: nuclear and cytoplasmic localisation; standard IF practice).
What should I check when MED15 DAB staining looks diffuse?
Inspect a no primary antibody control and the counterstained section for diffuse deposit, tissue folds, and staining outside intact cells (standard IHC practice). The catalog example used 10% goat serum blocking, 1 µg/mL primary antibody, a biotinylated secondary, and DAB development (datasheet A03568-1). Compare background after adjusting blocking, primary concentration, washing, and chromogen development one variable at a time (standard IHC practice). With biotin based detection, assess background from the detection system using an appropriate control (datasheet A03568-1: detection method; standard IHC practice). Retain compartment specific signal that tracks intact cells as a candidate for MED15 staining (HPA: tissue IHC profile).
How should I quantify MED15 across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and score nuclear and cytoplasmic staining separately before comparing specimens (standard IHC practice; HPA: tissue IHC profile). An H-score combines the percentage of cells at each intensity level into a 0–300 score; alternatively report the percentage of positive cells (standard IHC scoring practice). For spatial questions, count positive cells per mm² of viable tissue and report the sampled area (standard IHC quantification practice). Normalise comparisons to the same cell population, viable tissue area, staining batch, and reference section (standard IHC practice). Set intensity thresholds against controls before scoring, and document how necrotic regions and section edges were excluded (standard IHC practice).
When is a MED15 IHC signal convincing rather than artefactual?
Look for reproducible staining in intact cells with nuclear or cytoplasmic localisation, both reported for MED15 (UniProt Q96RN5: subcellular location; HPA: tissue IHC profile). A solely extracellular deposit or colour confined to damaged edges needs review against the no primary antibody control (standard IHC practice). Exclude necrotic areas and check endogenous enzyme activity with detection controls before calling diffuse DAB signal positive (standard IHC practice). MED15 expression is widespread, and HPA reports high staining in several cell populations, including adipocytes and hematopoietic cells (HPA: tissue IHC profile). Interpret differences among cell populations with morphology and matched controls; HPA rates agreement between antibody staining and RNA expression as medium (HPA: Approved reliability description).
Boster reagents
Best MED15 / Mediator of RNA polymerase II transcription subunit 15 IHC Antibodies
Validated MED15 antibodies have IHC data from human colon cancer, lung cancer and placenta sections, and mouse brain sections (A03568-1 image captions); IF/ICC data come from U20S and A431 cells (catalog IF captions).
Real IHC data
Anti-MED15 Antibody ®
Cat # A03568-1
Real IF data
Anti-MED15 Antibody ® (monoclonal, 6F4)
Cat # M03568
A03568-1 is listed for IHC and IF, with paraffin section images from human colon cancer, lung cancer and placenta, and mouse brain, plus an IF/ICC image from U20S cells (catalog applications and image captions). M03568 is listed for IF/ICC, with an image from A431 cells; IHC is not listed for it (catalog applications and IF caption).
Which to pick: Choose A03568-1 for tissue IHC: its rabbit antibody has paraffin section images, with citrate retrieval at pH 6 for 20 minutes and primary antibody at 1 μg/ml (catalog A03568-1 IHC captions). The captions do not report the fixative (catalog A03568-1 IHC captions). For IF/ICC, M03568 is a mouse monoclonal, clone 6F4, with an A431 cell image; choose A03568-1 when Human, Mouse or Rat reactivity is needed, while M03568 lists Human reactivity only (catalog entries and IF captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q96RN5 (MED15_HUMAN, Mediator of RNA polymerase II transcription subunit 15).
- Human Protein Atlas. MED15 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. MED15 subcellular location (ICC-IF): Localized to the nucleoplasm..
- Human Protein Atlas. MED15 antibody validation summary (1 antibodies).
- MED15-TFE3 rearranged renal cell carcinoma: a subtype of TFE3-rearranged renal cell carcinoma with unique clinicopathologic features and better prognosis. World journal of surgical oncology 2025 — PMC12538908.
- Increased mediator complex subunit 15 expression is associated with poor prognosis in hepatocellular carcinoma. Oncology letters 2018 — PMC5844107.
- MED15 overexpression in prostate cancer arises during androgen deprivation therapy via PI3K/mTOR signaling. Oncotarget 2017 — PMC5352374.
- The knockdown of the Mediator complex subunit MED15 restrains urothelial bladder cancer cells' malignancy. Oncology letters 2018 — PMC6096071.
- PubMed PMID:11024300 — UniProt-cited evidence.
- PubMed PMID:11414760 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for MED15 (Mediator of RNA polymerase II transcription subunit 15). Expected localisation Cytoplasmic and nuclear staining (HPA tissue IHC). Antigen retrieval: Citrate pH 6 HIER, heat-mediated (datasheet A03568-1). Validated IHC antibody: A03568-1 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adipose tissue (Adipocytes); 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 A03568-1); 2. Fixation: Image fixative and duration unreported (datasheet A03568-1); 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 A03568-1); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A03568-1); 8. Primary antibody: Rabbit anti-MED15, 0.5-1μg/ml (datasheet A03568-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation conditions consistent across sections. (standard IHC practice; not target-specific); Caveat: Salivary glandular cells show low staining (HPA tissue IHC); Isoform / epitope: 3 isoforms; epitope coverage is unreported (UniProt); Regulation: Expression varies by tissue (UniProt). Evidence: UniProt Q96RN5, HPA tissue IHC · Approved, HPA ICC-IF · nucleoplasm., Datasheet A03568-1, PMC5844107, PMC5352374. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/MED15/immunohistochemistry-ihc-guide.