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MAPK1 / Mitogen-activated protein kinase 1 · IHC design guide
Design Immunohistochemistry for MAPK1
Plan chromogenic MAPK1 IHC in paraffin sections around the cytoplasmic and nuclear staining seen in most tissues (HPA tissue IHC). Use consistent fixation and score the two compartments separately, since phosphorylation can increase nuclear localization (UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
MAPK1 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 in most tissues (HPA tissue IHC) | |
| Staining pattern | Most tissues show cytoplasmic and nuclear staining (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen) | |
| 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 paraffin sections. (standard IHC practice; not target-specific) | |
| Caveat | Phosphorylation can increase nuclear localization (UniProt) | |
| Regulation | Expression regulation is not annotated (UniProt) | |
| Isoform / epitope | 2 isoforms; check antibody epitope coverage (UniProt) |
Section 1
Recommended MAPK1 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is followed by published MAPK1 IHC protocols for parathyroid adenoma, breast cancer, and mouse liver (PMC11279064; PMC5591392; PMC12587933).
| Sample | Paraffin-embedded Human melanoma tissue; fixative not specified (datasheet M00030) |
| Fixation | Image fixative and duration unreported (datasheet M00030); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in Tris-EDTA buffer, pH 9.0, 20 min at 95–98 °C (standard rule: nuclear antigen) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Rabbit monoclonal (clone II-13) anti-MAPK1, 1:50 recommended; image 1:100 (datasheet M00030) |
| Primary incubation | Overnight at 4 °C (standard) |
| Detection | HRP-polymer secondary, DAB chromogen 5–10 min (standard) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | MAPK1-positive staining in adipocytes of adipose tissue (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic and nuclear expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with the page’s Tris-EDTA pH 9.0 retrieval at 95–98 °C for 20 min; two published protocols used pH 6 (page retrieval; PMC11279064; PMC5591392).
Section 2
What Is the Expected MAPK1 Staining Pattern?
MAPK1/ERK2 is expected in the cytoplasm and nuclei of many cell types; HPA reports cytoplasmic and nuclear staining in most tissues, with high staining in selected adipocytes, epithelial cells, glia, and neurons (HPA: tissue IHC). Its lack of a transmembrane segment supports intracellular interpretation (UniProt P28482 topology). HPA rates the tissue profile Enhanced but reports only medium consistency between antibody staining and RNA expression (HPA: tissue IHC).
| Cytoplasmic and nuclear chromogen in adipocytes, bronchial respiratory epithelium, or cortical neurons (HPA: High in these cells). | This fits the broad tissue pattern (HPA: cytoplasmic and nuclear expression in most tissues). Judge intensity within identifiable cells and alongside controls; a nuclear-to-cytoplasmic difference alone does not establish a change in total MAPK1 abundance (UniProt P28482 subcellular location). |
| Predominantly nuclear or cytoplasmic staining within otherwise credible positive cells. | Either distribution can be plausible: MAPK1 occupies both compartments, and phosphorylation or binding partners can affect its localisation (UniProt P28482 subcellular location). Interpret a shift in the context of the antibody and specimen; total-MAPK1 staining alone does not identify the phosphorylated species (UniProt P28482 modified residues). |
| An isolated extracellular deposit or broad acellular chromogen is scored as MAPK1. | That is a compartment mismatch: MAPK1 has no transmembrane segment and is documented in intracellular locations, with additional caveola and focal-adhesion association (UniProt P28482 topology and subcellular location). Reassess deposits as possible IHC artefact before assigning a cellular score (general IHC practice). |
| Strong staining appears chiefly in cells reported as low, such as esophageal squamous epithelium or smooth muscle (HPA: Low). | Low means some staining is possible; it is not a negative-control designation (HPA: tissue IHC). An unexpected dominant signal warrants checks for antibody cross-reactivity, tissue pigment, or endogenous detection activity (general IHC practice). |
| No cellular signal appears in a well-preserved section containing bronchial respiratory epithelial cells (HPA: High). | This conflicts with the HPA reference pattern but does not by itself prove MAPK1 absence (HPA: High; tissue reliability Enhanced, medium RNA–staining consistency). Check assay controls and section handling before interpreting the specimen as negative (general IHC practice). |
💡Expected MAPK1 appearanceCall a positive result when identifiable HPA-high cells show clear intracellular cytoplasmic and/or nuclear staining; treat acellular deposits or widespread chromogen outside cells as suspect, not a positive MAPK1 pattern (HPA: tissue IHC; UniProt P28482 subcellular location; general IHC practice).
| Reference cells and tissue range | HPA reports High staining in adipocytes, bronchial respiratory epithelium, cortical neurons, glia, and other listed cells; it reports Low staining in several other cell groups and lists no negatives (HPA: tissue IHC). Choose a documented high-staining cell population when judging whether the run produced the expected pattern. |
| Localisation and signalling state | MAPK1 is reported in cytoplasm and nucleus, with spindle, centrosome, caveola, and focal-adhesion associations; retention and phosphorylation can influence localisation (UniProt P28482 subcellular location). A nuclear fraction is therefore plausible, but distribution alone cannot establish pathway activation or the abundance of a particular phospho-form (UniProt P28482 modified residues). |
| Antibody and molecular scope | The HPA tissue profile is Enhanced, while individual listed IHC antibodies are Approved except CAB079933, which is Enhanced (HPA: antibodies; tissue IHC). MAPK1 has two isoforms and multiple modified residues; those facts do not establish which forms any given antibody detects (UniProt P28482 isoforms and modified residues). Check the selected antibody's stated specificity before comparing stains. |
| IF/ICC Q&A: Should its localisation set the IHC scoring rule? | No. HPA IF/ICC reports mainly nucleoplasmic MAPK1, plus cytosol and other locations, but cautions that its subcellular summary uses antibodies targeting proteins from multiple genes (HPA: subcellular ICC-IF). Use the cytoplasmic-and-nuclear tissue IHC profile for paraffin-section interpretation (HPA: tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| The documented high-staining cells are blank (HPA: tissue IHC). | A failed run or weak detection is possible; a blank section alone does not identify the cause (general IHC practice). | Confirm that the chosen cells are present, then inspect the run controls, primary-antibody application, chromogen development, and counterstain (general IHC practice). HPA gives reference staining, not a target-specific retrieval requirement (HPA: tissue IHC). |
| The whole section has a brown haze that obscures cell borders. | Non-specific reagent retention or detection background can obscure a cellular pattern (general IHC practice). | Compare with an appropriate no-primary control; review blocking, washing, detection reagents, and development time (general IHC practice). Score MAPK1 only where cellular staining remains distinguishable (HPA: cytoplasmic and nuclear tissue pattern). |
| Pigmented or blood-rich areas seem positive despite weak cellular staining. | Intrinsic pigment or endogenous enzyme activity can mimic chromogenic signal (general IHC practice). | Inspect the no-primary control and tissue morphology; apply the detection system's endogenous-activity control where relevant (general IHC practice). Do not count acellular colour as cytoplasmic or nuclear MAPK1 (HPA: tissue IHC). |
| Only nuclei stain, or nuclei stain more strongly than cytoplasm. | A nuclear component is biologically plausible; localisation can vary with MAPK1 regulation (UniProt P28482 subcellular location). | Check that the signal is in intact cells and that controls are acceptable (general IHC practice). Record the compartmental pattern without calling activation unless the antibody is validated for the relevant phosphorylated form (UniProt P28482 modified residues). |
| A low-staining reference cell group gives the section's strongest signal (HPA: Low). | The pattern may reflect specimen variation, cross-reactivity, or detection background; HPA Low is not equivalent to absent (HPA: tissue IHC; general IHC practice). | Compare the same run with a documented HPA-high population and an appropriate control; review the selected antibody's validation before attributing the difference to MAPK1 (HPA: tissue IHC and antibodies; general IHC practice). |
| Two antibodies yield different nuclear-to-cytoplasmic patterns. | HPA assigns antibody-specific IHC validation levels, and MAPK1 has multiple isoforms and modified residues; neither fact alone explains the disagreement (HPA: antibodies; UniProt P28482 isoforms and modified residues). | Compare each antibody's stated target and validation, then inspect matched controls and cellular morphology (general IHC practice). Report the disagreement rather than merging the patterns into one MAPK1 score. |
Experiment evidence
IHC & IF images showing MAPK1 staining
M00030 · IHCImmunohistochemical analysis of paraffin-embedded Human melanoma, using the Antibody at 1:100 dilution.
M00030 · IHCImmunohistochemical analysis of paraffin-embedded Human renal cancer, using the Antibody at 1:100 dilution.
M00030 · IFImmunofluorescent analysis using the Antibody at 1:50 dilution.
A00030Y222 · IHCImmunohistochemical analysis of paraffin-embedded Human-lung tissue. 1, ERK 1/2 (phospho Tyr222/205) Polyclonal Antibody was diluted at 1:20
A00030Y222 · IHCImmunohistochemical analysis of paraffin-embedded Human-Appendix tissue. 1, ERK 1/2 (phospho Tyr222/205) Polyclonal Antibody was diluted at
A00030Y222 · IFImmunofluorescence analysis of Hela cell. 1, ERK 1/2 (phospho Tyr222/205) Polyclonal Antibody (red) was diluted at 1:200 (4° overnight). β-t
A00030 · IHCFormalin-fixed and paraffin-embedded human breast carcinoma tissue reacted with MAPK1 Antibody (C-term) (Cat. #A00030), which was peroxidase
PA1049 · IFIF analysis of MAPK1/3 using anti-MAPK1/3 antibody (PA1049) and anti-Tubulin Alpha antibody (M03989-3). MAPK1/3 was detected in immunocytoch
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000100030-MAPK1/tissue/cerebral+cortex#imid_2389232
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000100030-MAPK1/tissue/colon#imid_2328230
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000100030-MAPK1/tissue/liver#imid_2328215
HPA · PancreasHuman Protein Atlas IHC, Pancreas. Source: https://www.proteinatlas.org/ENSG00000100030-MAPK1/tissue/pancreas#imid_2328174
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA030069; green = MAPK1, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA030069; green = MAPK1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for MAPK1 IHC & IF
🧪Run adipose tissue first and confirm staining in adipocytes (HPA: High in adipocytes). HPA detects MAPK1 in all 45 scored tissues, so there is no supported negative tissue or validated negative cell population within the positive slide; use no-primary and isotype controls to establish background, and do not assume that an unstained cell is MAPK1-negative (HPA: no negative rows).
Positive control tissue: Adipose tissue (Adipocytes, HPA High)
Negative control tissue: None in HPA: MAPK1 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 MAPK1 in A-431, U-251MG, U2OS, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), KOLF2.1J, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only section and a control antibody matched to the primary antibody’s host species and immunoglobulin class, with clonality matched where applicable; use a MAPK1 knockout sample as a biological negative. Block endogenous peroxidase for chromogenic detection, and assess adipose tissue autofluorescence if using IF (standard IHC/IF practice).
⚠️Feasibility: No target-specific fixation window, fixation effect, or retrieval dependence is reported in the supplied evidence; optimize antigen retrieval empirically. The selected M00030 caption documents paraffin-section IHC at 1:100 in human melanoma but does not report a fixative (selected-SKU tissue-IHC caption); the relative ease of frozen sections or IF is unreported. In paraffin-processed adipose tissue, lipid extraction can leave empty vacuoles, so score intact adipocytes rather than empty spaces (standard histology practice).
HPA tissue IHC evidence for MAPK1
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 |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | High | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Breast | Adipocytes | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Caudate | Glial cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: MAPK1 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced MAPK1 IHC Tips
Troubleshoot MAPK1 staining in paraffin sections by checking retrieval, compartment pattern, controls and scoring before interpreting differences between samples.
How should I retrieve MAPK1 in paraffin sections with weak nuclear staining?
Start with Tris-EDTA at pH 9.0, heated to 95–98 °C for 20 minutes for this page’s nuclear-antigen IHC workflow (page retrieval setting). Cool sections consistently before staining, and compare a retrieval control from the same run because heating and cooling affect epitope access (standard IHC practice). Nuclear and cytoplasmic signal are both plausible for MAPK1, so assess both compartments when judging whether retrieval worked (HPA: cytoplasmic and nuclear expression in most tissues). If staining remains weak, compare a citrate-buffer retrieval condition on adjacent sections while holding antibody dilution and detection constant (standard IHC optimization). Excessive retrieval can damage tissue morphology or raise background, so retain the condition that preserves interpretable cells (standard IHC practice).
Could fixation explain weak MAPK1 staining in my paraffin sections?
Target-specific fixation sensitivity for MAPK1 is unknown from the supplied IHC evidence; the selected 1:100 melanoma image states paraffin embedding but does not report a fixative (M00030 caption). Record the actual fixative, fixation duration and processing history for each specimen before comparing staining intensity (standard IHC practice). If staining varies across a batch, compare sections with matched processing and a shared control under the same retrieval conditions (standard IHC practice). Evaluate tissue preservation and staining together: poor morphology or uneven labeling can indicate a processing problem, but cannot establish a MAPK1-specific fixation effect (standard IHC practice). Do not use expression patterns alone to assign a fixation cause (HPA: cytoplasmic and nuclear expression in most tissues).
Is nuclear MAPK1 staining expected alongside cytoplasmic staining?
Yes: MAPK1 is reported in the nucleus and cytoplasm, with additional associations including spindle, centrosome, caveola and focal adhesion (UniProt P28482: subcellular locations). In tissue IHC, HPA describes cytoplasmic and nuclear expression in most tissues, so score those compartments separately rather than merging their signals (HPA: tissue IHC profile). Phosphorylation at Ser-246, Ser-248 and Thr-190 can promote nuclear localization, whereas PEA15 binding and phosphorylated DAPK1 promote cytoplasmic retention (UniProt P28482: subcellular annotation). A total-MAPK1 stain cannot by itself establish which modification caused a compartment shift (UniProt P28482: modified residues; standard IHC interpretation). Check that nuclear signal follows intact nuclei and cytoplasmic signal follows preserved cell boundaries (standard IHC practice).
How can I assess isoform coverage and phospho-epitope sensitivity?
MAPK1 has 2 annotated isoforms and a protein kinase domain spanning residues 25–313 (UniProt P28482: isoforms and domains). Check the catalog antibody’s stated immunogen and mapped epitope before claiming that it recognizes both isoforms; those details are not supplied here (standard antibody validation practice). MAPK1 carries regulated phosphorylation sites, including Thr-185 and Tyr-187, so total-protein and phospho-specific staining answer different questions (UniProt P28482: modified residues). If modification dependence matters, compare matched sections with appropriately validated total and phospho-specific reagents under their respective conditions (standard IHC practice). Treat a staining difference as an epitope-access or specificity question until controls support a biological interpretation (standard IHC interpretation).
How should I investigate MAPK1 localization with multiplex IF?
For the separate IF/ICC workflow, pair MAPK1 with a validated marker identifying the expected cell population and include a nuclear counterstain to resolve compartment boundaries (standard IF practice). HPA reports mainly nucleoplasmic localization, with additional cytosolic and microtubule-associated signal, but cautions that its subcellular assessment uses antibodies targeting proteins from multiple genes (HPA: subcellular summary). Choose fluorophores after checking specimen autofluorescence and place the weaker signal in a spectrally clear channel (standard IF practice). Because MAPK1 has no transmembrane segment, use controlled permeabilisation to expose intracellular epitopes and compare with a no-primary control (UniProt P28482: topology; standard IF practice). Validate antibody compatibility and channel separation before interpreting apparent overlap with the cell marker (standard IF practice).
What should I check when MAPK1 chromogenic staining looks diffuse?
First compare a no-primary control and inspect whether color appears over tissue, extracellular material or slide edges under the same detection conditions (standard chromogenic IHC practice). Quench endogenous peroxidase for a peroxidase-based assay and optimize protein blocking, washes and chromogen development time as general workflow steps (standard chromogenic IHC practice). The selected paraffin melanoma image used the catalog antibody at 1:100, which is a documented image condition rather than a guaranteed working dilution for every section (M00030 caption). Excess chromogen or antibody can obscure compartment boundaries, so adjust one variable at a time against a matched control (standard IHC optimization). Preserve plausible nuclear and cytoplasmic staining while rejecting pigment or precipitate that does not follow cells (HPA: tissue IHC profile; standard IHC interpretation).
How should I score MAPK1 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define cell populations and compartments before scoring because MAPK1 has documented nuclear and cytoplasmic tissue staining (HPA: tissue IHC profile). For each compartment, report the percentage of positive cells and an H-score based on the proportions at each intensity, using the same thresholds across sections (standard IHC scoring practice). If spatial abundance matters, count positive cells per mm² of viable tissue and report the area assessed (standard digital pathology practice). Normalize comparisons to the relevant cell population or viable tissue area, with matched staining runs and exposure to chromogen (standard IHC practice). Record exclusion rules for folds, necrosis, edges and pigment before scoring to reduce operator-dependent differences (standard IHC quantification practice).
How can I distinguish true MAPK1 signal from staining artefacts?
A credible MAPK1 pattern follows intact cells and can include both nucleus and cytoplasm in tissue sections (HPA: cytoplasmic and nuclear expression in most tissues). Compare the labeled cell population with the specimen’s morphology; HPA reports broad expression and low tissue specificity, so cell identity alone cannot prove antibody specificity (HPA: tissue IHC profile and RNA specificity). Strong color restricted to cut edges, necrotic areas or tissue folds warrants review as a processing or staining artefact (standard IHC interpretation). Check no-primary and peroxidase-block controls when unexpected color might reflect detection reagents or endogenous enzyme activity (standard chromogenic IHC practice). Interpret a compartment shift cautiously: total MAPK1 localization does not independently demonstrate kinase activation (UniProt P28482: phosphorylation-linked localization; standard IHC interpretation).
Boster reagents
Best MAPK1 / Mitogen-activated protein kinase 1 IHC Antibodies
Catalog images show MAPK1/ERK2 IHC in human, mouse and rat paraffin sections (M00030 IHC captions) and MAPK1/3 IF in U2OS cells (PA1049 IF caption).
Real IHC data
Anti-ERK2 Monoclonal Antibody
Cat # M00030
Real IHC data
Anti-Phospho-ERK 1/2 (Y222/205) MAPK1 Antibody
Cat # A00030Y222
Real IHC data
Anti-MAPK1 Antibody (C-term)
Cat # A00030
Real IF data
Anti-MAPK1/3 Antibody ®
Cat # PA1049
M00030 has IHC images from paraffin-embedded human melanoma and renal cancer, rat lung and mouse ovary (M00030 IHC captions); A00030Y222 has IHC images from paraffin-embedded human lung and appendix, rat lung and mouse liver (A00030Y222 IHC captions). A00030 has IHC-P data from formalin-fixed, paraffin-embedded human breast carcinoma (A00030 IHC caption); PA1049 has IF/ICC data from U2OS cells (PA1049 IF caption).
Which to pick: For tissue IHC across human, mouse and rat, consider M00030: it is monoclonal clone II-13 with IHC listed and paraffin-section images in all three species; its captions do not report a fixative (M00030 catalog and IHC captions). For human paraffin-section, paraffin-embedded tissue, A00030 has direct breast carcinoma DAB evidence; choose A00030Y222 when phospho ERK1/2 is the target, noting that its paraffin-section captions do not report a fixative (A00030 IHC caption; A00030Y222 title and IHC captions). For IF/ICC, PA1049 has a U2OS cell image and both applications listed, while M00030 also lists IF and ICC (PA1049 IF caption and applications; M00030 applications). The selected M00030 tissue-IHC caption documents paraffin sections, but does not specify the fixative (selected-SKU IHC image M00030).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P28482 (MK01_HUMAN, Mitogen-activated protein kinase 1).
- Human Protein Atlas. MAPK1 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. MAPK1 subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the cytosol, microtubules, primary cilium, primary cilium tip and basal body. Caution: Based on antibodies targeting proteins from multiple genes..
- Human Protein Atlas. MAPK1 antibody validation summary (6 antibodies).
- Sporadic Parathyroid Adenoma: A Pilot Study of Novel Biomarkers in Females. Medicina (Kaunas, Lithuania) 2024 — PMC11279064.
- Increased MAPK1/3 Phosphorylation in Luminal Breast Cancer Related with PIK3CA Hotspot Mutations and Prognosis. Translational oncology 2017 — PMC5591392.
- Protective effect of capsaicin on AKT1 and MAPK1 expression in the liver of mice (Mus musculus) induced by aflatoxin B1. Open veterinary journal 2025 — PMC12587933.
- The vasculogenic mimicry related signature predicts the prognosis and immunotherapy response in renal clear cell carcinoma. BMC cancer 2024 — PMC10996246.
- PubMed PMID:1540184 — UniProt-cited evidence.
- PubMed PMID:1319925 — UniProt-cited evidence.
- PubMed PMID:10591208 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for MAPK1 (Mitogen-activated protein kinase 1). Expected localisation Cytoplasmic and nuclear staining in most tissues (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen). Validated IHC antibody: M00030 at 1:50 recommended; image 1:100 (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 workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded Human melanoma tissue; fixative not specified (datasheet M00030); 2. Fixation: Image fixative and duration unreported (datasheet M00030); 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-induced epitope retrieval in Tris-EDTA buffer, pH 9.0 , 20 min at 95–98 °C…; 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Rabbit monoclonal (clone II-13) anti-MAPK1, 1:50 recommended; image 1:100 (datasheet M00030); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation conditions consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Phosphorylation can increase nuclear localization (UniProt); Isoform / epitope: 2 isoforms; check antibody epitope coverage (UniProt); Regulation: Expression regulation is not annotated (UniProt). Evidence: UniProt P28482, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet M00030, PMC11279064, PMC5591392. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/MAPK1/immunohistochemistry-ihc-guide.