ECM1 / Extracellular matrix protein 1 · IHC design guide

Design Immunohistochemistry for ECM1

Plan paraffin section ECM1 IHC around cytoplasmic staining in squamous epithelia and epididymal glands (HPA tissue IHC). The catalog antibody was demonstrated at 1:50 (datasheet M02861-1); interpret staining with ECM1 secretion in mind (UniProt).

Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use

Immunohistochemistry protocol sheet for ECM1 (IHC for ECM1): expected localisation Observed cytoplasmic; molecularly secreted to ECM (HPA tissue IHC; UniProt), antibody M02861-1, validated IHC image, and IHC protocol steps — Printable **ECM1** IHC protocol sheet — expected localisation Observed cytoplasmic; molecularly secreted to ECM (HPA tissue IHC; UniProt), antibody M02861-1, controls and protocol steps. Open the full ECM1 IHC guide →

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ECM1 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	Observed cytoplasmic; molecularly secreted to ECM (HPA tissue IHC; UniProt)
	Staining pattern	Cytoplasmic staining in squamous and epididymal glandular cells (HPA tissue IHC)
	Antigen retrieval	EDTA pH 8.0 HIER, heat-mediated (datasheet M02861-1)
	Positive control ⓘ	Epididymis+4 more · see all
	Negative control ⓘ	Adipose tissue+4 more · see all

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	Secreted ECM1 may stain away from its source cells (HPA tissue IHC)
	Regulation	Higher expression in breast cancer tissue (UniProt)
	Isoform / epitope	4 isoforms; mature chain 20–540; check epitope coverage (UniProt)

Section 1

Recommended ECM1 IHC & IF Protocols

Compare the catalog antibody’s IHC-P protocol (datasheet M02861-1) with published ECM1 staining in HCC (PMC4962417) and endometrial samples (PMC13231627).

Recommended immunohistochemistry (IHC-P) protocol parameters
Sample	Paraffin-embedded mouse heart tissue; fixative not specified (datasheet M02861-1)
Fixation	Image fixative and duration unreported (datasheet M02861-1); verify before use.
Sectioning	4–5 µm sections on charged slides (standard)
Deparaffinisation	Xylene, graded ethanol series to water (standard)
Antigen retrieval	Heat retrieval: EDTA pH 8.0 (datasheet M02861-1); 20 min, 95–100 °C (standard)
Peroxidase block	3% H2O2, 10 min, room temperature (standard)
Blocking	10% goat serum (datasheet M02861-1)
Primary antibody	Rabbit monoclonal (clone 25E30) anti-ECM1, 1:50 (datasheet M02861-1)
Primary incubation	Overnight at 4 °C (datasheet M02861-1)
Detection	HRP-conjugated secondary, DAB chromogen (datasheet M02861-1)
Counterstain	Hematoxylin, blue, dehydrate and mount (standard)
Expected result	ECM1-positive staining in glandular cells of epididymis (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in squamous epithelia and in glandular cells of epididymis. No signal in the no-primary control.

💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval (datasheet M02861-1). Citrate pH 6.0 is a published comparison for HCC sections (PMC4962417).

Section 2

What Is the Expected ECM1 Staining Pattern?

ECM1 is secreted into the extracellular space and matrix and has no transmembrane segment (UniProt Q16610: location, topology). In tissue IHC, expect chiefly cytoplasmic staining in epididymal glandular cells and squamous epithelia (HPA: tissue IHC profile). HPA rates its tissue IHC reliability Enhanced, while reporting medium agreement between staining and RNA and disregarded presumed off-target binding (HPA: tissue IHC reliability).

What am I looking at on my slide?
Strong cytoplasmic staining in epididymal glandular cells.	This matches the clearest supplied positive tissue: HPA scores these cells High (HPA: epididymis). Compare the stained cells with tissue morphology before scoring; ECM1 is secreted, so the cell producing it and extracellular protein need not occupy identical locations (UniProt Q16610: location; HPA: reliability description).
Moderate cytoplasmic staining in squamous epithelium.	This fits HPA's tissue profile: squamous epithelial cells score Medium in esophagus, tonsil and vagina; cells in the skin corneal layer also score Medium (HPA: tissue IHC). Record the cell layer and distribution as well as intensity, because an entire tissue should not be treated as uniformly positive (HPA: cell-specific tissue scores).
Dominant nuclear-only staining on a tissue section.	This differs from the cytoplasmic tissue IHC profile and warrants review of morphology, detection controls and antibody performance (HPA: tissue IHC profile; standard IHC practice). HPA separately reports approved nucleoplasmic signal in ICC-IF, so a nuclear observation alone does not prove an artefact (HPA: subcellular ICC-IF).
Strong staining in cells scored Not detected by HPA, or widespread colour outside identifiable structures.	Treat this as a specificity or background question, not an ECM1-positive call. Examples of HPA Not detected cell populations include adipocytes in adipose tissue and hematopoietic cells in bone marrow (HPA: tissue IHC). Cross-reactivity or endogenous detection activity can produce unexpected staining (standard IHC practice); HPA also notes presumed off-target binding (HPA: reliability description).
No signal in epididymal glandular cells on a run intended to detect ECM1.	The expected High HPA signal is missing (HPA: epididymis). First assess tissue preservation, the staining run and its positive and detection controls (standard IHC practice). A failed run cannot establish that ECM1 is absent from the sample; HPA's Enhanced rating does not validate every antibody or staining condition (HPA: tissue reliability; standard IHC practice).

💡Expected ECM1 appearanceCall a tissue IHC result concordant when epididymal glandular cells stain strongly or the listed squamous epithelial cells stain moderately, chiefly in the cytoplasm; isolated dominant nuclear staining or broad staining in HPA Not detected cell populations needs control review (HPA: tissue IHC; HPA: subcellular ICC-IF; standard IHC practice).

How each factor affects the staining
Secreted protein and processing	ECM1 has a signal peptide at residues 1–19 and a mature chain at 20–540 (UniProt Q16610: processing). Extracellular signal can therefore be considered alongside cellular staining, but HPA's reported tissue IHC pattern is cytoplasmic (UniProt Q16610: location; HPA: tissue IHC profile).
Cell-specific reference levels	HPA scores epididymal glandular cells High and esophageal, tonsillar and vaginal squamous epithelial cells Medium (HPA: tissue IHC). Use these as pattern references, with the reported cell types and levels kept distinct (HPA: tissue IHC).
Agreement with RNA	HPA reports medium consistency between antibody staining and RNA and expects tissue locations of RNA and secreted protein to differ (HPA: reliability description). An RNA result alone should not determine which structure must stain on the slide (HPA: reliability description; standard IHC interpretation).
Isoforms and antibody recognition	UniProt lists 4 ECM1 isoforms, but the supplied record gives no antibody epitope or isoform coverage (UniProt Q16610: isoforms; supplied antibody data). Do not infer that staining or its absence measures every isoform; consult the chosen antibody's documented validation before interpreting a discrepancy (standard IHC practice).
HPA antibody validation	HPA027241 and HPA076029 each carry an Enhanced IHC status (HPA: antibody validation). That supports their reported IHC patterns, while HPA's tissue summary still notes presumed off-target binding that was disregarded (HPA: reliability description). Interpret unexpected sites against the cell-specific reference pattern and controls (standard IHC practice).
IF/ICC Q: should its compartment match tissue IHC?	A: HPA reports mainly cytosolic and additional nucleoplasmic ECM1 in ICC-IF, both approved, whereas its tissue IHC profile is cytoplasmic (HPA: subcellular ICC-IF; HPA: tissue IHC). Assess each application against its own reported pattern; this is an interpretation note, not an IF protocol.

Why is my staining missing, weak or wrong?
Situation	Likely cause	Next action
Epididymal glandular cells show no or very weak staining.	A run or reagent problem is possible because HPA scores these cells High; the observation alone does not identify which step failed (HPA: epididymis; standard IHC practice).	Check section integrity, positive and detection controls, antibody preparation and the recorded IHC conditions; repeat the run if a control failed (standard IHC practice).
Squamous epithelium stains, but the intensity is lower than in epididymis.	This may fit the reference pattern: HPA scores esophageal, tonsillar and vaginal squamous cells Medium versus High in epididymal glandular cells (HPA: tissue IHC).	Score the correct cell layer and compare like tissue and staining conditions before adjusting the assay (HPA: tissue IHC; standard IHC practice).
Unexpected cells stain strongly.	Possible explanations include cross-reactivity or endogenous detection activity (standard IHC practice). HPA reports presumed off-target binding in its tissue assessment (HPA: reliability description).	Check morphology and a detection control, then compare the exact cell population with HPA's reported positive and Not detected entries (standard IHC practice; HPA: tissue IHC).
Colour covers much of the section without a clear cell or matrix pattern.	Non-specific detection or inadequate background control can obscure cell-specific staining (standard IHC practice). This appearance does not match HPA's defined epithelial and glandular profile (HPA: tissue IHC).	Review the detection control, blocking and washes, then reassess the section for interpretable cell-specific signal (standard IHC practice; HPA: tissue IHC profile).
Predominantly nuclear staining appears in tissue IHC.	The result differs from HPA's cytoplasmic tissue profile, although additional nucleoplasmic signal is approved in ICC-IF (HPA: tissue IHC; HPA: subcellular ICC-IF).	Check controls and staining morphology before calling it specific; keep the ICC-IF observation separate from the tissue IHC interpretation (standard IHC practice; HPA: tissue IHC; HPA: subcellular ICC-IF).
Staining is seen near cells but does not track their RNA-rich compartment.	ECM1 is secreted, and HPA expects the tissue location of RNA and protein to differ (UniProt Q16610: location; HPA: reliability description).	Describe the observed cellular and extracellular distribution separately, then compare with HPA's tissue IHC pattern and assay controls before assigning specificity (HPA: tissue IHC; standard IHC practice).

Source: UniProt Q16610 Human Protein Atlas tissue Human Protein Atlas subcellular

Experiment evidence

IHC & IF images showing ECM1 staining

IHC analysis of ECM1 using anti-ECM1 antibody (M02861-1). ECM1 was detected in a paraffin-embedded section of mouse heart tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1:50 rabbit anti-ECM1 Antibody (M02861-1) overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit (Catalog # SV0002) with DAB as the chromogen.

M02861-1 · IHCIHC analysis of ECM1 using anti-ECM1 antibody (M02861-1). ECM1 was detected in a paraffin-embedded section of mouse heart tissue. Heat media

HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000143369-ECM1/tissue/cerebral+cortex#imid_17896011

HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000143369-ECM1/tissue/colon#imid_17895929

HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000143369-ECM1/tissue/liver#imid_17895938

HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000143369-ECM1/tissue/kidney#imid_17895931

HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA027241; green = ECM1, blue = nucleus, red = microtubules.

← swipe / scroll for more images →

Real IHC / IF data behind the staining-pattern expectations in this section.

Sample controls for ECM1 IHC & IF

🧪Run epididymis first and expect staining in its glandular cells (HPA: Epididymis, glandular cells, High). Use adipose tissue as a negative comparator, focusing on adipocytes (HPA: Adipose tissue, adipocytes, Not detected); cells without specific staining on the positive slide should remain at background level, without assuming every other cell type is ECM1-negative.

Positive control tissue: Epididymis (Glandular cells, HPA High)

Negative control tissue: Adipose tissue (HPA Not detected)

ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show ECM1 in A-431, U-251MG, U2OS, with annotated localisation: Cytosol (approved) (HPA subcellular).

Technical controls: Include a no-primary, secondary-only control and a host- and isotype-matched control IgG; the captioned primary is rabbit anti-ECM1 (selected-SKU tissue-IHC caption: M02861-1). Add ECM1 knockout tissue, if available, or a validated peptide-block control; for chromogenic detection, quench endogenous peroxidase and check for background in the epididymis section (standard IHC practice).

⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected-SKU paraffin-section caption does not state a fixative (selected-SKU tissue-IHC caption: M02861-1). Heat retrieval in EDTA at pH 8.0 was used for mouse heart, but retrieval dependence in epididymis is unreported; whether frozen sections or IF are easier is also unreported (selected-SKU tissue-IHC caption: M02861-1). Because ECM1 is secreted into extracellular space and matrix, distinguish glandular-cell staining from adjacent extracellular or luminal signal when scoring epididymis (UniProt Q16610: subcellular location; HPA: Epididymis, glandular cells, High).

HPA tissue IHC evidence for ECM1

Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data. Secreted protein, tissue location of RNA and protein is expected to differ. Presumed off target binding observed and disregarded.). 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
Epididymis	Glandular cells	High	Protein (IHC)	HPA →
Esophagus	Squamous epithelial cells	Medium	Protein (IHC)	HPA →
Skin	Cells in corneal layer	Medium	Protein (IHC)	HPA →
Tonsil	Squamous epithelial cells	Medium	Protein (IHC)	HPA →
Vagina	Squamous epithelial cells	Medium	Protein (IHC)	HPA →

Undetected expression · recommended negative controls

Tissue	Cell type	Level	Evidence	Source
Adipose tissue	Adipocytes	Not detected	Protein (IHC)	HPA →
Appendix	Glandular cells	Not detected	Protein (IHC)	HPA →
Bone marrow	Hematopoietic cells	Not detected	Protein (IHC)	HPA →
Caudate	Glial cells	Not detected	Protein (IHC)	HPA →
Cerebellum	Cells in granular layer	Not detected	Protein (IHC)	HPA →

Section 3

Advanced ECM1 IHC Tips

Use compartment, tissue controls, and matched processing conditions to troubleshoot ECM1 staining in paraffin sections. The IF question addresses the separate fluorescence workflow.

What retrieval should I try first for weak ECM1 staining in paraffin sections?

Start with heat-mediated antigen retrieval in EDTA, pH 8.0 (datasheet M02861-1). This condition was used for ECM1 staining in a paraffin-embedded mouse heart section with the catalog antibody at 1:50 after overnight incubation at 4°C (datasheet M02861-1). If signal is weak, compare heating durations on adjacent sections while keeping cooling, antibody dilution, and DAB development constant; excessive heating can damage morphology (standard IHC practice). Include a no-primary section to assess detection background and an expected-positive specimen, since secreted ECM1 can be unevenly distributed in extracellular matrix (UniProt Q16610 localisation).

How can I assess whether fixation is causing weak ECM1 IHC staining?

The mouse heart example describes a paraffin-embedded section but gives no fixative, so target-specific ECM1 fixation sensitivity is unknown (datasheet M02861-1). Record fixative, fixation duration, section age, and processing history, then compare matched sections using EDTA at pH 8.0 for retrieval (datasheet M02861-1; standard IHC practice). Hold antibody incubation and DAB development constant so differences can be assessed against tissue morphology and a no-primary control (standard IHC practice). Repeat the comparison within one tissue type before assigning a staining difference to ECM1 abundance, because variable preparation can change IHC signal (standard IHC practice).

Should ECM1 appear in the matrix or inside cells on chromogenic IHC?

ECM1 is a secreted protein assigned to extracellular space and matrix, with no transmembrane segment (UniProt Q16610 localisation and topology). Its signal peptide spans residues 1–19, so extracellular deposits are plausible even when nearby producing cells stain weakly (UniProt Q16610 processing). HPA tissue IHC also reports cytoplasmic staining in squamous epithelia and epididymal glandular cells, while its cell-based IF reports cytosol and nucleoplasm (HPA tissue IHC; HPA subcellular). Score epithelial cytoplasm and adjacent matrix separately, and inspect serial sections for boundaries; do not call isolated nuclear DAB staining ECM1 without corroborating controls (standard IHC practice).

Could an ECM1 epitope or isoform explain discordant tissue staining?

ECM1 has 4 annotated isoforms, but the supplied catalog caption does not identify the antibody epitope or establish isoform selectivity (UniProt Q16610 isoforms; datasheet M02861-1). The precursor has a 1–19 signal peptide and predicted glycosylation sites at 354, 444, and 530 (UniProt Q16610 processing and glycosylation). Ask for epitope mapping before interpreting a negative region as lacking every isoform; do not assume a site alters binding without direct evidence (standard IHC interpretation). Compare matched regions with an independently validated antibody to a known, distinct epitope and retain the same retrieval and scoring criteria (standard IHC practice).

How should I check ECM1 localisation when adding IF to the IHC study?

For separate IF experiments, pair ECM1 with an epithelial marker such as cytokeratin to identify the expected squamous or glandular cells (HPA tissue IHC; standard IF practice). Choose a fluorophore channel after measuring unstained tissue autofluorescence, and use single-label controls to assess bleed-through before multiplexing (standard IF practice). Because mature ECM1 is secreted and has no transmembrane segment, begin without detergent when assessing accessible extracellular signal; compare gentle permeabilisation if testing an intracellular epitope (UniProt Q16610 localisation and topology; standard IF practice). Keep these IF settings separate from the 1:50 paraffin-section IHC example, which establishes no IF dilution or fixation condition (datasheet M02861-1).

How do I reduce diffuse DAB staining without losing ECM1 signal?

Run a no-primary section and inspect whether the brown signal persists after peroxidase blocking, secondary reagent, and DAB development (standard IHC practice). In the documented mouse heart workflow, sections received 10% goat serum blocking, catalog antibody at 1:50 overnight at 4°C, and peroxidase-conjugated secondary for 30 minutes at 37°C (datasheet M02861-1). If background rises with primary antibody, titrate it on adjacent sections while fixing retrieval and DAB time (standard IHC practice). Interpret diffuse matrix staining cautiously because ECM1 is secreted, and compare its spatial pattern with the no-primary section (UniProt Q16610 localisation; standard IHC practice).

What should I measure when ECM1 stains both cells and extracellular matrix? ⚠ ANSWER MARKED FOR VERIFICATION

Define cell and matrix compartments before scoring, since ECM1 is secreted but HPA also reports cytoplasmic tissue staining (UniProt Q16610 localisation; HPA tissue IHC). For cellular signal, record percent positive cells and intensity, then calculate an H-score from the percentages at intensities 0–3 (standard IHC practice). For matrix signal, measure positive area or integrated DAB density per mm² of annotated viable tissue, using consistent illumination and stain separation (standard IHC practice). Normalise each measure to its corresponding cell population or tissue area, and compare sections stained in the same run with identical thresholds (standard IHC practice).

How can I distinguish convincing ECM1 staining from an IHC artefact?

Look for a reproducible cellular or extracellular pattern: ECM1 is secreted, while HPA reports cytoplasmic staining in squamous epithelia and epididymal glandular cells (UniProt Q16610 localisation; HPA tissue IHC). HPA rates its tissue staining Enhanced but notes presumed off-target binding and only medium agreement with RNA expression (HPA tissue IHC). Treat isolated nuclear DAB signal, necrotic areas, and section-edge staining cautiously, and check whether they persist in a no-primary section after peroxidase blocking (standard IHC practice). Compare at least 2 separately processed sections with expected-positive and expected-negative regions before assigning a biological difference (standard IHC practice).

Boster reagents

Best ECM1 / Extracellular matrix protein 1 IHC Antibodies

Anti-ECM1 antibody M02861-1 has IHC images from paraffin-embedded mouse and rat heart sections (IHC image captions). IF/ICC is listed as an application, but no IF image is supplied (catalog applications; catalog IF images).

Real IHC data

Anti-ECM1 Rabbit Monoclonal Antibody

Cat # M02861-1

View product →

The sole card, M02861-1, shows IHC in paraffin-embedded mouse and rat heart sections (IHC image captions). The catalog lists IHC and ICC/IF applications and human, mouse and rat reactivity (catalog applications; catalog reactivity).

Which to pick: Choose M02861-1 for paraffin-section IHC because its own images document mouse and rat heart staining; the fixative is unreported (IHC image captions). The same rabbit monoclonal, clone 25E30, is listed for ICC/IF, although no IF image is supplied (catalog host; catalog clone; catalog applications; catalog IF images). It is listed as reactive with human, mouse and rat, while the supplied IHC images document mouse and rat tissue only (catalog reactivity; IHC image captions).

Each figure is that product's own IHC / IF validation image from its datasheet.

References

UniProt Consortium. UniProt entry Q16610 (ECM1_HUMAN, Extracellular matrix protein 1).
Human Protein Atlas. ECM1 tissue IHC expression (reliability: Enhanced).
Human Protein Atlas. ECM1 subcellular location (ICC-IF): Mainly localized to the cytosol. In addition localized to the nucleoplasm..
Human Protein Atlas. ECM1 antibody validation summary (2 antibodies).
ECM1 promotes migration and invasion of hepatocellular carcinoma by inducing epithelial-mesenchymal transition. World journal of surgical oncology 2016 — PMC4962417.
Endometrial HOXA-10, HOXA-11, ß-1 integrin, ECM-1, FAK, and CD44 immunohistochemical expressions in endometriosis-related recurrent IVF failure: a retrospective case-control study. Journal of ovarian research 2026 — PMC13231627.
Extracellular matrix protein 1 (ECM1) is associated with carcinogenesis potential of human bladder cancer. OncoTargets and therapy 2019 — PMC6389008.
HLA-A2.1-restricted ECM1-derived epitope LA through DC cross-activation priming CD8(+) T and NK cells: a novel therapeutic tumour vaccine. Journal of hematology & oncology 2021 — PMC8082934.
PubMed PMID:9367673 — UniProt-cited evidence.
PubMed PMID:9501329 — UniProt-cited evidence.
PubMed PMID:14702039 — UniProt-cited evidence.