CACNA1H / Voltage-dependent T-type calcium channel subunit alpha-1H · IHC design guide

Design Immunohistochemistry for CACNA1H

Start chromogenic IHC-P with antibody A01406-1 at 5 μg/mL (datasheet). Use high staining in testis Leydig cells as a reference (HPA tissue IHC), while interpreting cytoplasmic or nuclear signal in light of the annotated membrane location (UniProt).

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

Immunohistochemistry protocol sheet for CACNA1H (IHC for CACNA1H): expected localisation Cytoplasmic/nuclear staining observed (HPA tissue IHC); membrane expected (UniProt), antibody A01406-1, validated IHC image, and IHC protocol steps — Printable **CACNA1H** IHC protocol sheet — expected localisation Cytoplasmic/nuclear staining observed (HPA tissue IHC); membrane expected (UniProt), antibody A01406-1, controls and protocol steps. Open the full CACNA1H IHC guide →

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CACNA1H 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/nuclear staining observed (HPA tissue IHC); membrane expected (UniProt)
	Staining pattern	Cytoplasmic/nuclear staining across tissues; strongest in Leydig cells (HPA tissue IHC)
	Antigen retrieval	Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen)
	Positive control ⓘ	Testis+4 more · see all
	Negative control ⓘ	Adipose tissue+4 more · see all

Important caveats

Reasons your staining may differ from the expected pattern.

	Fixation	Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A01406-1)
	Caveat	Presumed off-target binding can confound staining (HPA tissue IHC)
	Regulation	STAC increases cell-membrane expression (UniProt)
	Isoform / epitope	2 isoforms; check epitope conservation and membrane side (UniProt)

Section 1

Recommended CACNA1H IHC & IF Protocols

Compare the catalog antibody’s IHC-P protocol (catalog datasheet) with a published CACNA1H protocol for FFPE breast tissue (PMC13271060).

Recommended immunohistochemistry (IHC-P) protocol parameters
Sample	Tissue sections; selected-image fixative not specified (standard IHC workflow)
Fixation	Image fixative and duration unreported (datasheet A01406-1); 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 citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane 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 anti-CACNA1H, 5 μg/mL (datasheet A01406-1)
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	CACNA1H-positive staining in leydig cells of testis (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic and nuclear expression in several tissues, Most abundant in Leydig cells of testis, stromal cells in ovary and glandular cells. No signal in the no-primary control.

💡Decision noteStart with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page antigen retrieval rule); the published protocol also uses citrate pH 6.0 (PMC13271060).

Section 2

What Is the Expected CACNA1H Staining Pattern?

CACNA1H is a 24-pass cell-membrane calcium channel (UniProt O95180 topology). In paraffin-section IHC, assess cell-associated staining in Leydig cells of testis, where HPA reports high staining, and adrenal glandular cells, where it reports medium staining (HPA tissue IHC). HPA rates the tissue antibody Approved but reports low agreement with RNA and presumed off-target binding; compartment and cell identity therefore need cautious interpretation (HPA tissue IHC).

What am I looking at on my slide?
Strong, cell-associated staining in testicular Leydig cells, with a discernible cell-edge component.	This fits the strongest supplied tissue observation (HPA: High in Leydig cells) and the predicted membrane location (UniProt O95180 subcellular location). An edge component supports the expected location, but chromogenic IHC may not resolve the plasma membrane sharply; compare cell identity and distribution with the HPA observation before calling the result specific.
Predominantly nuclear staining, with no convincing cell-associated edge signal.	Treat a nuclear-only pattern as inconclusive for the membrane-channel expectation (UniProt O95180 topology). HPA does report cytoplasmic and nuclear tissue staining and approved nucleoplasm staining by ICC-IF, while also noting presumed off-target tissue binding (HPA tissue IHC; HPA subcellular). Nuclear signal alone cannot establish either true CACNA1H localisation or an artefact.
Similar staining across many cell types, including adipocytes or pancreatic exocrine cells.	This distribution conflicts with the supplied tissue comparison: those cells were not detected, while Leydig cells were High (HPA tissue IHC). Consider antibody cross-reactivity or endogenous chromogen-producing activity as general IHC possibilities. The HPA negative observations are comparison points, not proof that every specimen of those tissues must be blank.
Diffuse colour over cells, stroma, and the surrounding section, obscuring cell boundaries.	Interpret this as background until a restricted cellular pattern can be distinguished (standard IHC practice). Nonspecific antibody binding, insufficient washing, or endogenous detection activity are general workflow possibilities; the supplied sources do not identify which applies to CACNA1H. Check a matched control before assigning the haze to target expression.
No visible staining in a testis section containing identifiable Leydig cells.	A blank candidate positive control conflicts with the reported High Leydig-cell observation (HPA tissue IHC). First check tissue preservation and the IHC run, including retrieval, primary-antibody application, detection reagents, and chromogen development (standard IHC practice). A negative result alone cannot distinguish a failed run from specimen variation or an antibody-specific limitation.

💡Expected CACNA1H appearanceCall a result supportive when identifiable testicular Leydig cells show high, cell-associated staining with a plausible membrane component (HPA: High in Leydig cells; UniProt O95180: cell membrane); widespread uniform colour or staining equally strong in HPA-undetected cell types is suspect, especially given HPA’s off-target caveat (HPA tissue IHC).

How each factor affects the staining
Membrane topology and unknown antibody epitope	CACNA1H has 24 transmembrane segments and both cytoplasmic and extracellular regions (UniProt O95180 topology). Epitope position is absent from the supplied antibody record, so topology cannot justify a particular antigen-retrieval condition, permeabilisation setting, or claim that a weak result reflects inaccessible epitope.
Tissue evidence and validation limit	HPA labels the tissue antibody Approved, yet reports low staining–RNA consistency and presumed off-target binding (HPA tissue IHC). Compare signal across the named positive and undetected cell populations; the approval label alone cannot validate an unexpected compartment or a new tissue pattern.
Isoforms and processing	UniProt lists two isoforms and one chain spanning residues 1–2353, with no signal peptide or propeptide annotated (UniProt O95180 isoforms and processing). The supplied record does not map the antibody epitope to either isoform, so a difference in staining cannot be assigned to isoform selection or cleavage.
Does IF/ICC establish the IHC compartment?	HPA reports approved nucleoplasm and vesicle localisation in ICC-IF images from A-549 and MCF-7 (HPA subcellular). This is a separate application and does not validate nuclear chromogenic staining in paraffin tissue, particularly given the tissue IHC off-target caveat (HPA tissue IHC). Use the separate IF/ICC guide for that application.

Why is my staining missing, weak or wrong?
Situation	Likely cause	Next action
Leydig cells are blank in the proposed positive control.	The result conflicts with HPA’s High Leydig-cell observation; the specific cause is undetermined (HPA tissue IHC).	Verify Leydig-cell identity and section quality, then check retrieval, antibody application, detection reagents, and chromogen development using the established IHC workflow (standard IHC practice).
Every tissue compartment develops similar colour.	General IHC possibilities include nonspecific antibody binding or endogenous detection activity; HPA also flags presumed off-target binding for tissue IHC (standard IHC practice; HPA tissue IHC).	Inspect a matched no-primary control and the spatial pattern. Apply the detection system’s appropriate endogenous-activity blocking step if the control indicates it, then reassess the named positive and undetected cells (standard IHC practice; HPA tissue IHC).
Only nuclei stain in testis or adrenal tissue.	Nuclear tissue staining is reported, but the result does not by itself match UniProt’s cell-membrane assignment; HPA’s tissue specificity caveat remains (HPA tissue IHC; UniProt O95180 subcellular location).	Score nuclear and cell-edge signals separately and compare their distribution across identified cell types. Report nuclear-only staining as unresolved unless independently supported; do not treat the ICC-IF result as tissue-IHC validation (HPA subcellular; HPA tissue IHC).
Pancreatic exocrine cells or adipocytes stain as strongly as Leydig cells.	Those comparison populations were not detected in HPA tissue IHC; broad reactivity can reflect cross-reactivity or background (HPA tissue IHC; standard IHC practice).	Compare sections processed in the same run, inspect no-primary controls, and reassess whether the colour follows specific cells rather than section-wide background (standard IHC practice). Avoid declaring the antibody specific from intensity alone.
Adrenal glandular staining is weaker than testis staining.	This can agree with the supplied observations: adrenal glandular cells are Medium and testicular Leydig cells are High (HPA tissue IHC).	Identify the relevant cells and score each tissue against its own reported level before adjusting the assay. UniProt also reports protein in adrenal glomerulosa, but that statement does not prescribe an IHC intensity (UniProt O95180 tissue specificity).
A diffuse cytoplasmic haze obscures a possible cell-edge signal.	HPA describes cytoplasmic tissue staining, while diffuse field-wide colour can also be background; appearance alone cannot resolve the source (HPA tissue IHC; standard IHC practice).	Check whether the haze persists in a no-primary control, review blocking and washes, and score only interpretable cell-associated signal. Record cytoplasmic staining separately from a discernible edge pattern (standard IHC practice; UniProt O95180 subcellular location).

Source: UniProt O95180 Human Protein Atlas tissue Human Protein Atlas subcellular

Experiment evidence

IHC & IF images showing CACNA1H staining

A01406-1 · IHCImmunohistochemistry of CACNA1H in mouse brain tissue with CACNA1H antibody at 5 μg/ml.

A01406-1 · IFImmunofluorescence of CACNA1H in mouse brain tissue with CACNA1H antibody at 20 μg/ml.

HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000196557-CACNA1H/tissue/cerebral+cortex#imid_11067639

HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000196557-CACNA1H/tissue/colon#imid_11067557

HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000196557-CACNA1H/tissue/liver#imid_11067566

HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000196557-CACNA1H/tissue/kidney#imid_11067567

HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA039125; green = CACNA1H, blue = nucleus, red = microtubules.

← swipe / scroll for more images →

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

Sample controls for CACNA1H IHC & IF

🧪Run testis first and look for staining in Leydig cells (HPA: High in testis Leydig cells). Use adipose tissue as the negative control, where adipocytes are listed as not detected (HPA: Not detected in adipocytes); on the testis slide, expect adjacent non-Leydig cells to remain near background, but do not treat them as a validated negative population (HPA: Leydig cells are the annotated positive population).

Positive control tissue: Testis (Leydig cells, HPA High)

Negative control tissue: Adipose tissue (HPA Not detected)

ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CACNA1H in A-549, MCF-7, with annotated localisation: Nucleoplasm (approved), Vesicles (approved) (HPA subcellular).

Technical controls: Include a no-primary (secondary-only) slide, an isotype control matched to the primary antibody’s host species and clonality, and a knockout specimen or peptide-block control if available (standard IHC practice). On testis sections, block endogenous peroxidase before chromogenic detection and check for background staining (standard chromogenic IHC practice).

⚠️Feasibility: A target-specific fixation window, fixation effect, and antigen-retrieval dependency are unreported in the supplied evidence; optimize retrieval empirically for paraffin sections (standard IHC practice). The selected A01406-1 mouse-brain IHC caption reports 5 µg/ml but leaves the fixative unreported (selected-SKU tissue-IHC caption). Frozen sections or IF cannot be judged easier from these data: UniProt places CACNA1H at the cell membrane, while HPA ICC-IF reports nucleoplasm and vesicles, so assess localization and background in each preparation (UniProt O95180 subcellular location; HPA subcellular).

HPA tissue IHC evidence for CACNA1H

Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Low consistency between antibody staining and RNA expression data. 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
Testis	Leydig cells	High	Protein (IHC)	HPA →
Adrenal gland	Glandular cells	Medium	Protein (IHC)	HPA →
Bronchus	Ciliated cells (ciliary rootlets)	Medium	Protein (IHC)	HPA →
Esophagus	Squamous epithelial cells	Medium	Protein (IHC)	HPA →
Fallopian tube	Ciliated cells (ciliary rootlets)	Medium	Protein (IHC)	HPA →

Undetected expression · recommended negative controls

Tissue	Cell type	Level	Evidence	Source
Adipose tissue	Adipocytes	Not detected	Protein (IHC)	HPA →
Pancreas	Exocrine glandular cells	Not detected	Protein (IHC)	HPA →
Seminal vesicle	Glandular cells	Not detected	Protein (IHC)	HPA →
Smooth muscle	Smooth muscle cells	Not detected	Protein (IHC)	HPA →
Spleen	Cells in red pulp	Not detected	Protein (IHC)	HPA →

Section 3

Advanced CACNA1H IHC Tips

For CACNA1H chromogenic IHC in paraffin sections, troubleshoot retrieval and staining alongside compartment, cell type, and antibody specificity evidence.

How should I adjust retrieval when CACNA1H staining is weak in paraffin sections?

Use citrate pH 6.0 heat-induced epitope retrieval at 95–98 °C for 20 min (page retrieval setting). Cool sections consistently, then compare staining in matched sections with the same antibody concentration and detection conditions so retrieval is the main variable (standard IHC practice). If staining remains weak, test a more stringent retrieval condition on a separate section while monitoring tissue integrity and background; this is an optimization step, not a demonstrated CACNA1H requirement (standard IHC practice). Record the location of any recovered signal because CACNA1H has 24 transmembrane segments and a long cytoplasmic C-terminal region, while reported tissue staining includes cytoplasmic and nuclear signal (UniProt O95180 topology; HPA tissue IHC).

Could fixation explain inconsistent CACNA1H staining across paraffin samples?

CACNA1H-specific fixation sensitivity is unknown from the supplied evidence, and the catalog antibody’s mouse brain IHC caption does not state a fixative (selected A01406-1 tissue-IHC caption). For paraffin IHC, document fixative, fixation duration, processing, and section age before comparing samples, because these variables can change epitope accessibility and morphology (standard IHC practice). Apply the same citrate pH 6.0, 95–98 °C, 20 min retrieval to matched sections when investigating variation (page retrieval setting; standard IHC practice). Do not infer a CACNA1H fixation effect from its membrane topology, glycosylation sites, or HPA staining pattern; evaluate the samples empirically with matched controls (UniProt O95180 topology and glycosylation; HPA tissue IHC; standard IHC practice).

How should I assess membrane, cytoplasmic, and nuclear CACNA1H staining?

Prioritize staining that follows cell boundaries or plausible intracellular membrane profiles, since UniProt places CACNA1H at the cell membrane and annotates 24 transmembrane segments (UniProt O95180 subcellular location and topology). Cytoplasmic or nuclear chromogenic signal needs extra scrutiny: HPA reports both in tissues but also reports low consistency with RNA and presumed off-target binding (HPA tissue IHC). HPA separately reports approved nucleoplasm and vesicle locations from cell imaging, which does not by itself validate a nuclear pattern in paraffin tissue (HPA subcellular; standard IHC interpretation). Compare compartments in morphologically intact positive and negative control cells under identical retrieval and detection conditions before assigning a localisation (standard IHC practice).

What should I check when an antibody stains only some CACNA1H-positive cells?

Check the catalog antibody’s stated immunogen and epitope before interpreting selective staining; the supplied evidence does not locate this antibody’s epitope (standard IHC practice; supplied antibody evidence). UniProt lists 2 CACNA1H isoforms, 24 transmembrane segments, extensive cytoplasmic regions, and glycosylation sites at residues 192, 271, and 1466 (UniProt O95180 isoforms, topology, and glycosylation). An epitope in a variable or poorly accessible region could affect recognition, but the specific effect cannot be predicted without an epitope map and validation data (standard IHC interpretation). Compare staining with an independently validated antibody to a distinct epitope and assess matched tissue controls before attributing differences to isoforms (standard IHC practice).

How can IF help evaluate an ambiguous CACNA1H IHC pattern?

On the separate IF/ICC guide, multiplex CACNA1H with an independently validated marker of the expected cell type, such as an adrenal zona glomerulosa cell marker when examining adrenal tissue (UniProt O95180 tissue specificity; standard IF practice). Choose a fluorophore channel after checking tissue autofluorescence with an unstained section, and use single-stain controls to evaluate spectral spillover (standard IF practice). Set permeabilisation according to the antibody epitope: a cytoplasmic epitope requires intracellular access, whereas an extracellular epitope may be assessed without that access if the assay permits (UniProt O95180 topology; standard IF practice). Interpret any nuclear or vesicular signal alongside controls because HPA cell imaging reports those locations, while tissue IHC has a low staining-to-RNA consistency warning (HPA subcellular; HPA tissue IHC).

How can I reduce diffuse brown staining without losing CACNA1H signal?

First compare no-primary controls and intact tissue morphology to separate nonspecific chromogen deposition from an antibody-dependent pattern (standard chromogenic IHC practice). Block endogenous peroxidase before peroxidase-based DAB detection, use an appropriate protein block, and titrate the primary antibody and detection reagents on matched sections (standard chromogenic IHC practice). The selected catalog antibody caption reports staining in mouse brain at 5 µg/ml, but supplies no fixative or general-purpose dilution recommendation (selected A01406-1 tissue-IHC caption). Judge improvements against the same retrieval condition and cell-level controls, since HPA reports presumed off-target tissue staining for CACNA1H (page retrieval setting; HPA tissue IHC; standard IHC practice).

What scoring method best captures heterogeneous CACNA1H staining? ⚠ ANSWER MARKED FOR VERIFICATION

Define the cell population and compartment before scoring, then report the percentage of positive cells and an intensity-based H-score for the same population (standard IHC quantification practice). An H-score weights the percentages at intensity scores 0–3 and spans 0–300; use one prespecified threshold and exposure-independent brightfield assessment across samples (standard IHC quantification practice). For spatial questions, report positive-cell density per mm² of viable, annotated tissue and normalize each measure to the eligible cells or area actually assessed (standard IHC quantification practice). Keep nuclear, cytoplasmic, and membrane scores separate because UniProt places CACNA1H at the cell membrane while HPA reports additional patterns with a specificity warning (UniProt O95180 subcellular location; HPA tissue IHC).

Which findings make a CACNA1H-positive IHC result convincing?

Look for reproducible staining in intact, anatomically appropriate cells with a consistent compartment pattern, supported by matched positive and negative controls (standard IHC interpretation). UniProt reports CACNA1H protein in adrenal glomerulosa, whereas HPA reports medium staining in adrenal glandular cells and high staining in testicular Leydig cells; these are useful context, not proof of antibody specificity (UniProt O95180 tissue specificity; HPA tissue IHC). Scrutinize dominant nuclear staining, unexpected cell populations, section-edge enhancement, and necrotic regions, especially because HPA flags presumed off-target binding and low staining-to-RNA consistency (HPA tissue IHC; standard IHC interpretation). Check no-primary and peroxidase controls when brown signal could reflect detection background or endogenous enzyme activity (standard chromogenic IHC practice).

Boster reagents

Best CACNA1H / Voltage-dependent T-type calcium channel subunit alpha-1H IHC Antibodies

The IHC-validated anti-CACNA1H antibody has mouse brain IHC and IF images (catalog image captions); listed reactivity covers human, mouse, and rat (catalog reactivity).

Real IHC data Immunohistochemistry of CACNA1H in mouse brain tissue with CACNA1H antibody at 5 μg/ml.

Anti-CACNA1H Antibody

Cat # A01406-1

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A01406-1 is listed for IHC-P, with a mouse brain tissue image at 5 μg/mL (catalog applications; IHC image caption). A01406-1 is also listed for IF, with a mouse brain tissue image at 20 μg/mL (catalog applications; IF image caption).

Which to pick: For tissue IHC, choose A01406-1: it is listed for IHC-P and has its own mouse brain IHC image; the fixative is unreported (catalog applications; IHC image caption). For IF, choose A01406-1 based on its mouse brain IF image; ICC validation is unreported (IF image caption; catalog applications). For cross-species studies, A01406-1 lists human, mouse, and rat reactivity, although the supplied IHC and IF images show mouse tissue only; its host is rabbit and clonality is unreported (catalog reactivity; image captions; catalog host and clone fields).

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

References

UniProt Consortium. UniProt entry O95180 (CAC1H_HUMAN, Voltage-dependent T-type calcium channel subunit alpha-1H).
Human Protein Atlas. CACNA1H tissue IHC expression (reliability: Approved).
Human Protein Atlas. CACNA1H subcellular location (ICC-IF): Localized to the nucleoplasm and vesicles..
Human Protein Atlas. CACNA1H antibody validation summary (1 antibodies).
Genetic variants and down-regulation of CACNA1H in pheochromocytoma. Endocrine-related cancer 2024 — PMC11301417.
Exploring CACNA1H as a potential candidate biomarker for calcific aortic valve disease. iScience 2026 — PMC13137072.
Pan-Cancer Analysis of a Depression-Related Gene Signature Identifies an Immunosuppressive Tumor Microenvironment and Links to Immunotherapy Response in Breast Cancer. Breast cancer (Dove Medical Press) 2026 — PMC13271060.
PubMed PMID:9670923 — UniProt-cited evidence.
PubMed PMID:9930755 — UniProt-cited evidence.
PubMed PMID:11751928 — UniProt-cited evidence.