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CACNA1H / Voltage-dependent T-type calcium channel subunit alpha-1H · IHC design guide
Design Immunohistochemistry for CACNA1H
Plan chromogenic CACNA1H IHC-P with the catalog antibody starting at 5 μg/mL (datasheet: IHC-P). Evaluate Leydig-cell staining and cytoplasmic/nuclear signal cautiously because presumed off-target binding was observed (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
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 | Tissue: cytoplasm/nuclei (HPA tissue IHC); expected: cell membrane (UniProt) | |
| Staining pattern | Highest in Leydig cells; cytoplasmic/nuclear across tissues (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 | Keep fixation consistent (standard IHC practice; not target-specific); 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 staining and low RNA concordance (HPA tissue IHC) | |
| Regulation | STAC increases cell-membrane expression (UniProt) | |
| Isoform / epitope | 2 isoforms; check if the epitope is cytoplasmic or extracellular (UniProt) |
Section 1
Recommended CACNA1H IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by published chromogenic IHC protocols for PPGL and breast tumor specimens (PMC11301417; PMC13271060).
| 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 for this membrane antigen (page antigen retrieval; UniProt O95180 topology).
Section 2
What Is the Expected CACNA1H Staining Pattern?
CACNA1H is a 24-pass cell-membrane channel (UniProt O95180 topology). In paraffin IHC, HPA reports the strongest staining in testicular Leydig cells, with moderate staining in several other cell types (HPA tissue IHC). HPA also describes cytoplasmic and nuclear staining, but rates tissue IHC Approved with low consistency against RNA data and presumed off-target binding (HPA tissue IHC). Interpret compartment and cell identity together.
| Strong staining in testicular Leydig cells, with cell-associated signal. | This matches the strongest listed tissue result: High in Leydig cells (HPA tissue IHC). Compare adjacent cell types and tissue structure before scoring a slide; intensity alone cannot resolve the off-target concern in the HPA reliability note (HPA tissue IHC). |
| Moderate staining in adrenal glandular cells or ciliary rootlets of bronchial or fallopian-tube ciliated cells. | These are listed at Medium intensity (HPA tissue IHC). The ciliary-rootlet pattern is a specific reported observation; score the stained cell and structure, rather than treating every stained area in those tissues as equivalent (HPA tissue IHC). |
| Predominantly nuclear staining without a convincing cell-associated pattern. | Treat this as uncertain: a membrane location is annotated (UniProt O95180 subcellular location), while nuclear tissue staining is also reported (HPA tissue IHC). HPA notes presumed off-target binding, so nuclear signal alone does not establish specific CACNA1H detection (HPA tissue IHC). |
| Strong staining in adipocytes or pancreatic exocrine glandular cells. | Both are listed as Not detected (HPA tissue IHC). Recheck cell identification, slide background and detection controls; unexpected staining may reflect cross-reactivity or endogenous detection activity (general IHC practice). It cannot be assigned to CACNA1H from intensity alone. |
| Uniform haze across cells and extracellular areas, or no staining in Leydig cells. | Haze obscures the reported cell-selective pattern; consider background from staining or detection steps (general IHC practice). A blank Leydig-cell section conflicts with HPA’s High observation, but may reflect assay performance or sample variation (HPA tissue IHC; general IHC practice). |
💡Expected CACNA1H appearanceA convincing positive is strong Leydig-cell staining with identifiable cellular boundaries (HPA tissue IHC); diffuse field-wide color or strong signal in HPA-listed negative cells warrants control review (HPA tissue IHC; general IHC practice).
| Membrane topology and epitope access | CACNA1H has 24 transmembrane segments and intracellular and extracellular regions (UniProt O95180 topology). Epitope location is not supplied; assess the antibody’s documented epitope before inferring whether staining should outline a membrane or require permeabilisation (general IHC practice). |
| Reported intracellular patterns | HPA tissue IHC describes cytoplasmic and nuclear staining, while UniProt annotates the cell membrane (HPA tissue IHC; UniProt O95180 subcellular location). This disagreement limits compartment-only calls; HPA also reports low RNA–staining consistency and presumed off-target binding (HPA tissue IHC). |
| Isoforms and processing | UniProt lists 2 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 an antibody epitope to either isoform, so isoform-specific IHC predictions are unavailable. |
| Retrieval and detection controls | Antigen retrieval, blocking and detection controls are general paraffin-IHC workflow choices (general IHC practice). The supplied HPA and UniProt records give no CACNA1H-specific retrieval condition or fixation sensitivity; do not infer one from topology or staining intensity (HPA tissue IHC; UniProt O95180 topology). |
| Situation | Likely cause | Next action |
|---|---|---|
| Leydig cells lack signal in a section expected to be positive. | The result differs from HPA’s High Leydig-cell observation; tissue preservation, retrieval, antibody or detection performance may need checking (HPA tissue IHC; general IHC practice). | Confirm Leydig-cell identity and tissue integrity, then review the validated IHC procedure and a suitable positive control (general IHC practice). Do not infer a CACNA1H-specific fixation effect from this failure. |
| Strong nuclear signal dominates the section. | HPA reports nuclear staining but also presumed off-target binding; UniProt locates CACNA1H at the cell membrane (HPA tissue IHC; UniProt O95180 subcellular location). | Record nuclear and nonnuclear patterns separately. Compare cell types with HPA’s tissue observations and inspect controls before calling the nuclear signal specific (HPA tissue IHC; general IHC practice). |
| Adipocytes, pancreatic exocrine cells or other listed negative cells stain strongly. | These examples conflict with HPA’s Not detected entries (HPA tissue IHC). Cross-reactivity or endogenous detection activity is possible (general IHC practice). | Check cell identification and a no-primary control; review blocking and detection steps for the assay used (general IHC practice). Keep the unexpected staining separate from a positive CACNA1H score. |
| Color is diffuse, including extracellular spaces. | Field-wide haze can arise from background staining or detection steps (general IHC practice); it does not reproduce HPA’s cell-specific observations (HPA tissue IHC). | Compare the section with a no-primary control, inspect wash and blocking steps, and score only resolvable cells after background is addressed (general IHC practice). |
| A tissue shows moderate staining, but the stained cell type is unclear. | HPA assigns levels to particular cells, including adrenal glandular cells and bronchial ciliary rootlets, rather than to every cell in a tissue (HPA tissue IHC). | Use morphology and counterstain to identify the stained cells; document compartment and intensity with the cell name (general IHC practice). Mark uncertain identities as unresolved. |
| Can HPA’s IF/ICC images define the paraffin-IHC positive pattern? | HPA IF/ICC lists approved nucleoplasm and vesicles in A-549 and MCF-7 images; tissue IHC has a separate staining profile and reliability note (HPA subcellular ICC-IF; HPA tissue IHC). | Use those IF/ICC locations as a modality-specific comparison only. Base paraffin-IHC cell selection and scoring on tissue-IHC observations, with their low-consistency caveat (HPA tissue IHC). |
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.
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 Leydig cells). Use adipose tissue as the negative tissue (HPA: Not detected in adipocytes); on the testis slide, morphologically identified non-Leydig cells should show background-level staining, but their negative status is not established by the supplied HPA rows.
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 control; an irrelevant isotype control matched to the primary antibody’s host species, antibody class, and clonality; and a knockout sample or validated peptide-block control. For chromogenic IHC in testis, quench endogenous peroxidase and use the no-primary slide to assess residual background.
⚠️Feasibility: The selected A01406-1 mouse brain IHC caption reports 5 μg/ml but does not report a fixative (selected tissue-IHC caption). A target-specific fixation window or fixation effect and an antigen-retrieval dependency are unreported in the supplied evidence; it also does not establish whether frozen sections or IF are easier. In testis, blood-associated endogenous peroxidase can produce chromogenic background, so assess it with the no-primary control (standard IHC practice).
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
Troubleshoot CACNA1H chromogenic IHC by testing retrieval and detection conditions, then judging staining against membrane topology and cell-level controls.
What retrieval conditions should I try when CACNA1H staining is weak?
Start with citrate buffer at pH 6.0, heated to 95–98 °C for 20 min (page retrieval setting). Allow slides to cool in the buffer, then compare staining with a no-primary control processed through the same retrieval and chromogenic detection steps (standard IHC practice). If staining remains weak, test a second retrieval condition on adjacent sections while keeping antibody concentration and detection time constant (standard IHC practice). Score cellular detail alongside signal, because excessive heating can damage morphology and make apparent staining harder to assign to a compartment (standard IHC practice).
Could fixation explain weak or uneven staining in paraffin sections?
Target-specific fixation sensitivity for CACNA1H is unknown from the supplied evidence; the mouse brain IHC caption for A01406-1 does not state a fixative (selected IHC caption: A01406-1). Record the fixative, fixation duration, processing schedule and section thickness for each specimen before comparing staining (standard IHC practice). If specimens were processed differently, stain adjacent sections together and include a reference specimen processed by a consistent method (standard IHC practice). Test retrieval changes one at a time and assess morphology as well as signal, since fixation and processing differences can alter antigen accessibility in paraffin sections (standard IHC practice).
How should I assess cytoplasmic or nuclear staining for this membrane channel?
CACNA1H is annotated at the cell membrane and has 24 transmembrane segments (UniProt O95180 localisation and topology). Examine whether chromogenic signal outlines cells, follows intracellular structures, or instead fills nuclei, using the counterstain to assign compartments (standard IHC practice). Cytoplasmic and nuclear tissue staining has been reported, but its consistency with RNA expression was low and presumed off-target binding was noted (HPA tissue IHC reliability and profile). Approved nucleoplasm and vesicle localisation in cell imaging provides a reason to investigate those patterns, while remaining separate evidence from paraffin-section IHC (HPA subcellular). Compare each pattern with no-primary controls before assigning it to CACNA1H (standard IHC practice).
Why might different CACNA1H antibodies stain different compartments?
CACNA1H has 2 annotated isoforms and spans the membrane 24 times, with both extracellular and cytoplasmic regions (UniProt O95180 isoforms and topology). Check the immunogen or mapped epitope before interpreting discordant staining, because its position determines which side of the membrane the antibody must access (standard IHC practice; UniProt O95180 topology). The record lists glycosylation sites at residues 192, 271 and 1466, which may matter when comparing epitopes near those positions; no effect on staining is established here (UniProt O95180 glycosylation). Compare antibodies on adjacent sections with identical retrieval and detection, and report the observed compartment for each (standard IHC practice).
How can IF help check the cellular source of an IHC signal?
On the separate IF/ICC guide workflow, pair CACNA1H staining with a marker for the cell type being assessed, such as a validated Leydig-cell marker when examining testis (HPA tissue IHC: High in Leydig cells; standard IF practice). Choose spectrally separated fluorophores and inspect unstained tissue for autofluorescence before assigning faint signal, especially in the channel selected for CACNA1H (standard IF practice). Match permeabilisation to the mapped epitope: a cytoplasmic epitope needs access through the plasma membrane, whereas extracellular-epitope staining can be assessed without that access (UniProt O95180 topology; standard IF practice). Include single-label and no-primary controls to check bleed-through and background; do not equate IF patterns with validated chromogenic IHC staining (standard IF practice).
How do I distinguish CACNA1H staining from chromogenic background?
Run a no-primary slide through the same secondary reagent and chromogen, and inspect it beside the test slide at matched development time (standard IHC practice). For peroxidase-based detection, include a peroxidase block and check whether residual color appears in the control; use an appropriate protein block to assess nonspecific reagent binding (standard IHC practice). Compare tissue edges, folds and damaged areas with intact interior regions before calling a diffuse brown signal positive (standard IHC practice). This check matters particularly because presumed off-target tissue staining was observed in the reference profile (HPA tissue IHC reliability). Titrate the primary antibody against a consistent reference section if background persists (standard IHC practice).
What should I score when comparing CACNA1H staining across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, then record either an H-score from staining intensity and percentage of positive cells or the percentage positive at a fixed threshold (standard IHC practice). For spatial comparisons, count positive cells per mm² of intact, evaluable tissue and report the analyzed area (standard IHC practice). Normalize counts to the number of eligible cells or tissue area, and compare only sections stained with the same retrieval, antibody and chromogen development conditions (standard IHC practice). Keep membrane-associated and nuclear signals in separate scoring fields because CACNA1H is membrane-annotated while nuclear staining has also been reported (UniProt O95180 localisation; HPA tissue IHC profile).
What evidence makes a CACNA1H-positive cell convincing?
A convincing call requires reproducible staining in an anatomically identified cell, an interpretable compartment and little matching signal in the no-primary control (standard IHC practice). Membrane-associated staining fits the annotated channel location; nuclear-only staining needs additional scrutiny because tissue IHC showed low consistency with RNA and presumed off-target binding (UniProt O95180 localisation; HPA tissue IHC reliability). Assess cell identity against the reported high Leydig-cell staining and medium adrenal glandular-cell staining where those tissues are examined, without treating either pattern as proof of specificity (HPA tissue IHC). Exclude section edges, necrotic regions and residual endogenous peroxidase signal from positive calls (standard IHC practice).
Boster reagents
Best CACNA1H / Voltage-dependent T-type calcium channel subunit alpha-1H IHC Antibodies
A01406-1 has mouse brain IHC and IF images (catalog: image captions) and listed reactivity with human, mouse, and rat (catalog: reactivity).
Real IHC data
Anti-CACNA1H Antibody
Cat # A01406-1
A01406-1 is listed for IHC-P and IF (catalog: applications), with mouse brain images at 5 μg/mL for IHC and 20 μg/mL for IF (catalog: image captions). The supplied images document mouse brain only (catalog: image captions); human and rat are listed as reactive species (catalog: reactivity).
Which to pick: For paraffin-section IHC, choose A01406-1 (catalog: IHC-P); its IHC image shows mouse brain at 5 μg/mL (catalog: IHC image caption), but the fixative is unreported (catalog: IHC image caption). For IF, choose A01406-1 based on its mouse brain image at 20 μg/mL (catalog: IF image caption); ICC is not separately listed (catalog: applications). For work across species, A01406-1 lists human, mouse, and rat reactivity (catalog: reactivity), although its supplied IHC and IF images show mouse brain only (catalog: image captions); the host is rabbit and clonality is unspecified (catalog: host; clone).
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.
Immunohistochemistry protocol sheet for CACNA1H (Voltage-dependent T-type calcium channel subunit alpha-1H). Expected localisation Tissue: cytoplasm/nuclei (HPA tissue IHC); expected: cell membrane (UniProt). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A01406-1 at 5 μg/mL (the product's own IHC validation image is shown). Controls (HPA): positive Testis (Leydig cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Tissue sections; selected-image fixative not specified (standard IHC workflow); 2. Fixation: Image fixative and duration unreported (datasheet A01406-1); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat-induced epitope retrieval in citrate buffer, pH 6.0 , 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 anti-CACNA1H, 5 μg/mL (datasheet A01406-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent (standard IHC practice; not target-specific); 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 staining and low RNA concordance (HPA tissue IHC); Isoform / epitope: 2 isoforms; check if the epitope is cytoplasmic or extracellular (UniProt); Regulation: STAC increases cell-membrane expression (UniProt). Evidence: UniProt O95180, HPA tissue IHC · Approved, HPA ICC-IF · nucleoplasm and vesicles., Datasheet A01406-1, PMC11301417, PMC13271060. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CACNA1H/immunohistochemistry-ihc-guide.