MICAL2 / [F-actin]-monooxygenase MICAL2 · IHC design guide

Design Immunohistochemistry for MICAL2

Plan MICAL2 paraffin-section IHC using adrenal or rectal glandular cells as strong positive references (HPA tissue IHC). This guide pairs the catalog antibody’s IHC evidence (datasheet A08683-1) with cytoplasmic tissue-staining expectations and the reported low consistency between staining and RNA expression (HPA tissue IHC).

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

Immunohistochemistry protocol sheet for MICAL2 (IHC for MICAL2): expected localisation Cytoplasmic tissue staining; nuclear location annotated (HPA tissue IHC; UniProt), antibody A08683-1, validated IHC image, and IHC protocol steps — Printable **MICAL2** IHC protocol sheet — expected localisation Cytoplasmic tissue staining; nuclear location annotated (HPA tissue IHC; UniProt), antibody A08683-1, controls and protocol steps. Open the full MICAL2 IHC guide →

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MICAL2 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 tissue staining; nuclear location annotated (HPA tissue IHC; UniProt)
	Staining pattern	Glandular and other cells show cytoplasmic staining (HPA tissue IHC)
	Antigen retrieval	EDTA pH 8.0 HIER, heat-mediated (datasheet A08683-1)
	Positive control ⓘ	Adrenal gland+4 more · see all
	Negative control ⓘ	Caudate+3 more · see all

Important caveats

Reasons your staining may differ from the expected pattern.

	Fixation	Keep fixation consistent across sections (standard IHC practice; not target-specific)
	Caveat	Staining shows low consistency with RNA expression (HPA tissue IHC)
	Regulation	Abundance regulation is not annotated (UniProt)
	Isoform / epitope	7 isoforms; antibody epitope coverage is unspecified (UniProt; datasheet A08683-1)

Section 1

Recommended MICAL2 IHC & IF Protocols

Compare the catalog antibody’s IHC protocol (datasheet A08683-1) with four published MICAL2 IHC methods (PMC13182836; PMC10850094; PMC6276083; PMC4811499).

Recommended immunohistochemistry (IHC-P) protocol parameters
Sample	Paraffin-embedded human pancreas cancer tissue; fixative not specified (datasheet A08683-1)
Fixation	Image fixative and duration unreported (datasheet A08683-1); 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 A08683-1); 20 min, 95–100 °C (standard)
Peroxidase block	3% H2O2, 10 min, room temperature (standard)
Blocking	10% goat serum (datasheet A08683-1)
Primary antibody	Rabbit anti-MICAL2, 1:50 recommended; image 1:100 (datasheet A08683-1)
Primary incubation	Overnight at 4 °C (datasheet A08683-1)
Detection	HRP-conjugated secondary, DAB chromogen (datasheet A08683-1)
Counterstain	Hematoxylin, blue, dehydrate and mount (standard)
Expected result	MICAL2-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most tissues. No signal in the no-primary control.

💡Decision noteStart with heat retrieval in EDTA pH 8.0 (datasheet A08683-1). For tendon sections, citrate pH 6.0 pressure retrieval is a published alternative (PMC13182836).

Section 2

What Is the Expected MICAL2 Staining Pattern?

MICAL2 is a non-transmembrane protein annotated in the cytoplasm and nucleus (UniProt O94851: topology and subcellular location). In paraffin-section IHC, expect predominantly cytoplasmic staining in glandular cells and other cell populations documented by HPA (HPA: tissue IHC). HPA rates the tissue staining “Approved” but reports low consistency with RNA expression and pending external verification (HPA: tissue IHC reliability).

What am I looking at on my slide?
Cytoplasmic staining in adrenal or rectal glandular cells.	This matches HPA’s reported High staining in those cells and its predominantly cytoplasmic tissue profile (HPA: tissue IHC). Score the stained cell population and compartment together; intensity alone cannot establish specificity because HPA reports low consistency with RNA expression (HPA: tissue IHC reliability).
Predominantly nuclear staining with little cytoplasmic staining.	A nuclear signal is biologically plausible because UniProt also annotates MICAL2 in the nucleus (UniProt O94851: subcellular location). Nuclear-only tissue staining, however, differs from HPA’s predominantly cytoplasmic IHC profile; check localization and controls before interpreting it as MICAL2 (HPA: tissue IHC; general IHC practice).
Strong staining in cardiomyocytes or oral squamous epithelial cells.	HPA reports MICAL2 as Not detected in these cell populations (HPA: heart muscle and oral mucosa tissue IHC). Unexpected signal warrants checks for antibody cross-reactivity and endogenous chromogen-generating activity; those are possible explanations, not findings established for MICAL2 (general IHC practice).
Uniform color over cells, stroma and blank areas.	A broad deposit that does not follow cell boundaries or the expected cytoplasmic distribution is difficult to assign to MICAL2 (HPA: tissue IHC; general IHC practice). Compare it with a negative detection control and inspect washing, blocking and detection conditions (general IHC practice).
No detectable staining in adrenal or rectal glandular cells.	Absence conflicts with HPA’s High staining calls for those cells (HPA: adrenal gland and rectum tissue IHC). Check that the relevant glandular cells are present, then review the antibody’s IHC validation, staining run and detection controls before calling the sample negative (HPA: antibodies; general IHC practice).

💡Expected MICAL2 appearanceA convincing positive is predominantly cytoplasmic staining in glandular cells, potentially High in adrenal gland or rectum; strong staining in a cell population HPA calls Not detected is a warning sign requiring control-based review (HPA: tissue IHC; general IHC practice).

How each factor affects the staining
Cell population and tissue	HPA reports High staining in adrenal and rectal glandular cells, Medium staining in several other listed populations, and Not detected in selected cells including cardiomyocytes (HPA: tissue IHC). Interpret the named cell population rather than assigning one expected intensity to an entire tissue (general IHC practice).
Compartment	UniProt annotates nucleus and cytoplasm, while HPA describes predominantly cytoplasmic tissue staining (UniProt O94851: subcellular location; HPA: tissue IHC). Thus, nuclear signal can be plausible, but a nuclear-only IHC pattern needs separate evaluation against the reported tissue pattern (general IHC practice).
Evidence strength and antibody choice	HPA labels the tissue IHC profile “Approved” while noting low agreement with RNA expression and pending external verification (HPA: tissue IHC reliability). The three listed antibodies, HPA030437, HPA039895 and HPA040438, have “Approved” IHC status; this does not remove the tissue-profile caveat (HPA: antibodies; HPA: tissue IHC reliability).
Isoforms and antigen processing	UniProt lists 7 MICAL2 isoforms, no signal peptide or propeptide, and one annotated chain spanning residues 1–1957 (UniProt O94851: isoforms and processing). The supplied record gives no antibody epitope, so it cannot establish which isoforms any listed antibody detects or predict an epitope-specific IHC pattern (UniProt O94851; HPA: antibodies).
IF/ICC: should it match tissue IHC?	HPA ICC-IF places MICAL2 mainly in cytosol, with supported nucleoplasmic and uncertain plasma-membrane localization; the main cytosol assignment is also uncertain (HPA: subcellular ICC-IF). Treat those localization details as IF context, not as a paraffin-section IHC protocol or a requirement for membrane staining in tissue (HPA: subcellular ICC-IF; HPA: tissue IHC).

Why is my staining missing, weak or wrong?
Situation	Likely cause	Next action
Positive-control glandular cells show no signal.	The run may have failed, or the expected cells may be absent from the section; HPA reports High staining in adrenal and rectal glandular cells (HPA: tissue IHC; general IHC practice).	Confirm the cells on the counterstained section, then review the IHC-validated antibody, detection reagents and run controls (HPA: antibodies; general IHC practice).
Signal is mainly nuclear in tissue.	UniProt permits nuclear localization, but HPA describes tissue staining as predominantly cytoplasmic (UniProt O94851: subcellular location; HPA: tissue IHC).	Score nuclear and cytoplasmic staining separately and compare matched controls before accepting a nuclear-only result (general IHC practice).
Cardiomyocytes or oral squamous cells stain strongly.	These populations are Not detected in HPA tissue IHC; cross-reactivity or endogenous detection activity is possible (HPA: tissue IHC; general IHC practice).	Compare an appropriate negative detection control and an HPA-reported positive cell population; investigate persistent unexpected staining (HPA: tissue IHC; general IHC practice).
Diffuse deposit obscures cell boundaries.	Background from blocking, washing or detection conditions can prevent cellular interpretation (general IHC practice).	Inspect the negative control, washing and blocking steps; score MICAL2 only where staining can be assigned to cells and a compartment (general IHC practice).
A negative population is used to reject a positive result elsewhere.	HPA reports different levels by cell population, from High in some glandular cells to Not detected in others (HPA: tissue IHC).	Identify and score each cell population separately, with its own HPA reference where available (HPA: tissue IHC; general IHC practice).
An IF membrane signal is treated as the expected IHC pattern.	HPA calls plasma-membrane ICC-IF localization uncertain, while its tissue IHC profile is cytoplasmic (HPA: subcellular ICC-IF; HPA: tissue IHC).	Interpret the paraffin-section result against the tissue IHC pattern; use IF/ICC localization only as context and keep the HPA reliability caveat visible (HPA: tissue IHC reliability; general IHC practice).

Source: UniProt O94851 Human Protein Atlas tissue Human Protein Atlas subcellular

Experiment evidence

IHC & IF images showing MICAL2 staining

IHC analysis of MICAL2 using anti-MICAL2 antibody (A08683-1). MICAL2 was detected in a paraffin-embedded section of human pancreas cancer 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 1:100 rabbit anti-MICAL2 Antibody (A08683-1) 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.

A08683-1 · IHCIHC analysis of MICAL2 using anti-MICAL2 antibody (A08683-1). MICAL2 was detected in a paraffin-embedded section of human pancreas cancer ti

IHC analysis of MICAL2 using anti-MICAL2 antibody (A08683-1). MICAL2 was detected in a paraffin-embedded section of human prostate cancer 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 1:100 rabbit anti-MICAL2 Antibody (A08683-1) 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.

A08683-1 · IHCIHC analysis of MICAL2 using anti-MICAL2 antibody (A08683-1). MICAL2 was detected in a paraffin-embedded section of human prostate cancer ti

HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000133816-MICAL2/tissue/cerebral+cortex#imid_7540032

HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000133816-MICAL2/tissue/colon#imid_7539974

HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000133816-MICAL2/tissue/liver#imid_7539967

HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000133816-MICAL2/tissue/kidney#imid_7539968

HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA039895; green = MICAL2, blue = nucleus, red = microtubules.

← swipe / scroll for more images →

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

Sample controls for MICAL2 IHC & IF

🧪Run rectum first: glandular cells should stain (HPA: High in rectal glandular cells). Use oral mucosa squamous epithelial cells as the negative tissue (HPA: Not detected); on the rectal slide, compare staining with adjacent nonglandular stromal cells, confirming their background level on that slide rather than assuming they are MICAL2-negative.

Positive control tissue: Adrenal gland (Glandular cells, HPA High)

Negative control tissue: Caudate (HPA Not detected)

ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show MICAL2 in A-431, U-251MG, U2OS, with annotated localisation: Cytosol (uncertain) (HPA subcellular).

Technical controls: Include a no-primary, secondary-only control, a concentration-matched rabbit IgG isotype control, and MICAL2 knockout material processed alongside the tissue (caption: rabbit primary antibody; standard IHC controls). For chromogenic detection, quench endogenous peroxidase and check inflammatory cells and blood for background signal (caption: peroxidase secondary and DAB; standard IHC practice).

⚠️Feasibility: The selected SKU caption describes heat-mediated EDTA retrieval at pH 8.0 and 1:100 primary antibody incubation overnight at 4°C in a paraffin section of pancreas cancer; it does not establish that retrieval is required for rectum (selected SKU tissue-IHC caption). A target-specific fixation window or fixation effect is unreported, and the caption does not report a fixative (selected SKU tissue-IHC caption). The supplied evidence does not establish whether frozen sections or IF would be easier; assess rectal mucin and inflammatory-cell background when scoring glandular staining (HPA: High in rectal glandular cells; standard IHC practice).

HPA tissue IHC evidence for MICAL2

Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Low consistency between antibody staining and RNA expression data. Pending external verification.). 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 →
Rectum	Glandular cells	High	Protein (IHC)	HPA →
Appendix	Glandular cells	Medium	Protein (IHC)	HPA →
Bone marrow	Hematopoietic cells	Medium	Protein (IHC)	HPA →
Breast	Glandular cells	Medium	Protein (IHC)	HPA →

Undetected expression · recommended negative controls

Tissue	Cell type	Level	Evidence	Source
Caudate	Glial cells	Not detected	Protein (IHC)	HPA →
Heart muscle	Cardiomyocytes	Not detected	Protein (IHC)	HPA →
Oral mucosa	Squamous epithelial cells	Not detected	Protein (IHC)	HPA →
Spleen	Cells in red pulp	Not detected	Protein (IHC)	HPA →

Section 3

Advanced MICAL2 IHC Tips

Troubleshoot MICAL2 staining in paraffin sections by checking retrieval, compartment, cell identity, and controls before interpreting chromogenic signal.

What retrieval should I try first if MICAL2 staining is weak?

Start with heat-mediated retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A08683-1). The selected image used this retrieval before staining human pancreas cancer tissue with the catalog antibody at 1:100 overnight at 4°C (datasheet A08683-1). If signal remains weak, vary heating time on serial sections while keeping antibody concentration, detection, and development constant; include an established positive control in every run (standard IHC practice). Judge improvement by staining in intact cells rather than stronger signal at folds or section edges, and reject conditions that increase diffuse background (standard IHC practice).

How should I troubleshoot possible fixation effects on MICAL2 staining?

Target-specific fixation sensitivity is unknown: the selected paraffin-section caption does not state a fixative, and its staining cannot establish a fixation recommendation (datasheet A08683-1). Record the actual fixative and fixation interval for each specimen, then compare sections processed together using EDTA retrieval at pH 8.0 (datasheet A08683-1; standard IHC practice). Keep the catalog antibody at the documented 1:100 starting dilution during that comparison so fixation is the main variable (datasheet A08683-1; standard IHC practice). If archival blocks differ in handling, interpret a staining difference alongside tissue preservation and internal controls before assigning it to MICAL2 expression (standard IHC practice).

Which cellular compartments should show credible MICAL2 staining?

Assess cytoplasmic staining first: tissue IHC describes cytoplasmic expression in most tissues, although its antibody staining has low consistency with RNA data (HPA tissue IHC). Nuclear signal is biologically plausible because MICAL2 is annotated in both nucleus and cytoplasm, while nucleoplasmic localisation has support from ICC/IF evidence (UniProt O94851 subcellular; HPA subcellular). Score nuclear and cytoplasmic staining separately on serial sections, using the same counterstain and development endpoint across specimens (standard IHC practice). Treat isolated surface staining cautiously: MICAL2 has no transmembrane segment, and plasma-membrane localisation is marked uncertain (UniProt O94851 topology; HPA subcellular).

Could isoforms or epitope accessibility explain discordant MICAL2 staining?

MICAL2 has 7 annotated isoforms, so check whether the antibody's documented immunogen overlaps the isoforms relevant to the specimen before interpreting absent staining (UniProt O94851 isoforms; standard IHC practice). The record places a CH domain at residues 516–619, a LIM domain at 1000–1062, and a bMERB domain at 1796–1945; these positions help map a disclosed epitope (UniProt O94851 domains). Phosphoserines at 631 and 1688 are annotated, but their effects on this antibody's IHC staining are unreported (UniProt O94851 modified residues; datasheet A08683-1). If the epitope is undisclosed, compare staining with an independently validated antibody recognizing another region and use matching controls (standard IHC practice).

How can IF help verify the pattern seen by chromogenic IHC?

For a separate IF/ICC experiment, multiplex MICAL2 with a marker identifying the cell population being assessed; glandular cells in adrenal gland or rectum are documented tissue-IHC reference populations (HPA tissue IHC). Choose a fluorophore channel after imaging an unstained section for tissue autofluorescence, and include single-label controls to assess bleed-through (standard IF practice). Select permeabilisation according to a disclosed epitope's accessibility: intracellular epitopes require membrane access, while MICAL2 has no transmembrane segment and is annotated in nucleus and cytoplasm (standard IF practice; UniProt O94851 topology and subcellular). Compare compartment patterns cautiously because the paraffin IHC caption supplies no IF fixation or permeabilisation conditions (datasheet A08683-1).

How do I reduce diffuse or misleading MICAL2 DAB background?

The selected tissue caption used 10% goat serum blocking, a peroxidase-conjugated secondary, and DAB development after 1:100 primary incubation overnight at 4°C (datasheet A08683-1). For diffuse brown signal, compare a no-primary section and an established positive section, then adjust blocking, washes, primary concentration, or DAB development one variable at a time (standard IHC practice). Include a peroxidase-blocking step in the chromogenic workflow and check whether residual endogenous activity persists in the no-primary control (standard IHC practice). Treat staining concentrated at folds, damaged edges, or necrotic areas as suspect until intact-cell staining is reproducible (standard IHC practice).

What scoring method makes MICAL2 IHC comparisons defensible? ⚠ ANSWER MARKED FOR VERIFICATION

Define the cell population and compartment before scoring; report cytoplasmic and nuclear staining separately because both locations are annotated for MICAL2 (UniProt O94851 subcellular; standard IHC practice). For DAB sections, use an H-score from intensity and percentage of positive cells, or report the percentage positive within a prespecified cell population (standard IHC practice). Normalise counts or positive area to the number of evaluable cells or evaluable tissue area, and exclude folds, necrosis, and poorly preserved regions by the same rule in every section (standard IHC practice). Keep retrieval, imaging, counterstain, and DAB development consistent, then document thresholds and observer agreement before comparing groups (standard IHC practice).

When is an apparent MICAL2-positive section likely an artefact?

Credible staining should occur in intact cells and fit the expected cytoplasmic pattern, with possible nuclear staining assessed separately (HPA tissue IHC; UniProt O94851 subcellular). Compare the cell type with tissue references: adrenal and rectal glandular cells were scored high, whereas heart cardiomyocytes and oral squamous epithelial cells were scored not detected (HPA tissue IHC). Suspect a technical cause when signal is restricted to section edges, folds, or necrosis, or appears in a no-primary control after peroxidase detection (standard IHC practice). Interpret any mismatch conservatively because the tissue-IHC profile is approved but has low consistency with RNA expression and awaits external verification (HPA tissue IHC).

Boster reagents

Best MICAL2 / [F-actin]-monooxygenase MICAL2 IHC Antibodies

A08683-1 has real IHC images from paraffin-embedded human cancer and mouse intestine (catalog IHC captions). IF/ICC and rat reactivity are listed, but no IF image is supplied (catalog applications/reactivity/images).

Real IHC data

Anti-MICAL2 Antibody

Cat # A08683-1

View product →

A08683-1 is the sole card; its IHC captions show human pancreas cancer, human prostate cancer and mouse small intestine in paraffin sections (A08683-1 IHC captions). IF/ICC is listed for this rabbit polyclonal antibody, with human, mouse and rat reactivity, but no IF image is supplied (catalog applications/host/clonality/reactivity/IF images).

Which to pick: Choose A08683-1 for tissue IHC: its own captions document EDTA retrieval at pH 8.0, goat-serum blocking, 1:100 overnight incubation and peroxidase/DAB detection in paraffin sections; the fixative is unreported (A08683-1 IHC captions). For IF/ICC, A08683-1 is listed at 1:50, without an IF figure (catalog applications/IF dilution/IF images). For cross-species work, A08683-1 lists human, mouse and rat reactivity, while its IHC images document human and mouse samples only (catalog reactivity/A08683-1 IHC captions).

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

References

UniProt Consortium. UniProt entry O94851 (MICA2_HUMAN, [F-actin]-monooxygenase MICAL2).
Human Protein Atlas. MICAL2 tissue IHC expression (reliability: Approved).
Human Protein Atlas. MICAL2 subcellular location (ICC-IF): Mainly localized to the cytosol. In addition localized to the nucleoplasm and plasma membrane..
Human Protein Atlas. MICAL2 antibody validation summary (3 antibodies).
Identification of Key Genes and Exploration of Therapeutic Targets for Chronic Tendon Injury Based on Bioinformatics and Machine Learning. International journal of general medicine 2026 — PMC13182836.
MICAL2 implies immunosuppressive features and acts as an independent and adverse prognostic biomarker in pancreatic cancer. Scientific reports 2024 — PMC10850094.
MICAL2 Mediates p53 Ubiquitin Degradation through Oxidating p53 Methionine 40 and 160 and Promotes Colorectal Cancer Malignance. Theranostics 2018 — PMC6276083.
MICAL2 is a novel human cancer gene controlling mesenchymal to epithelial transition involved in cancer growth and invasion. Oncotarget 2016 — PMC4811499.
PubMed PMID:16675569 — UniProt-cited evidence.
PubMed PMID:9872452 — UniProt-cited evidence.
PubMed PMID:14702039 — UniProt-cited evidence.