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JPH1 / Junctophilin-1 · IHC design guide
Design Immunohistochemistry for JPH1
Plan chromogenic JPH1 IHC-P using skeletal muscle myocytes as a positive tissue reference (HPA tissue IHC). Compare their cytoplasmic staining with the catalog antibody’s 2.5 μg/mL IHC starting concentration (datasheet: mouse skeletal muscle IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
JPH1 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 staining in skeletal myocytes (HPA tissue IHC) | |
| Staining pattern | Strong cytoplasmic staining in skeletal muscle myocytes (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Skeletal muscle+1 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 A08565) | |
| Caveat | Presumed off-target binding can confound staining (HPA tissue IHC) | |
| Regulation | Abundant in skeletal muscle; very low in heart (UniProt) | |
| Isoform / epitope | No isoforms annotated; N-terminal region is cytoplasmic (UniProt) |
Section 1
Recommended JPH1 IHC & IF Protocols
The catalog antibody’s paraffin-section protocol is followed by one published JPH1 chromogenic IHC protocol using mouse muscle cryosections (PMC10889371).
| Sample | Tissue sections; selected-image fixative not specified (standard IHC workflow) |
| Fixation | Image fixative and duration unreported (datasheet A08565); 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-JPH1, 2.5 μg/mL (datasheet A08565) |
| 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 | JPH1-positive staining in myocytes of skeletal muscle (HPA tissue IHC: High). HPA tissue profile: Selective cytoplasmic expression in muscle, most abundant in skeletal muscle. No signal in the no-primary control. |
💡Decision noteFor paraffin sections, start with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page retrieval); the published cryosection protocol reports no retrieval step (PMC10889371).
Section 2
What Is the Expected JPH1 Staining Pattern?
JPH1 links the plasma membrane to the endoplasmic or sarcoplasmic reticulum, with its C-terminal transmembrane segment anchored in the latter membrane (UniProt Q9HDC5 topology). In paraffin-section IHC, expect predominantly cytoplasmic staining in skeletal myocytes; spermatogonia also score high (HPA: tissue IHC). HPA rates the tissue profile Enhanced, while noting medium agreement with RNA and presumed off-target staining that was disregarded (HPA: tissue IHC reliability).
| Strong cytoplasmic staining in skeletal myocytes, with little staining in adjacent negative cells (HPA: High in skeletal myocytes). | This fits the most abundant tissue pattern reported by HPA (HPA: tissue IHC profile). JPH1 is associated with junctions between the cell surface and sarcoplasmic reticulum (UniProt Q9HDC5 function and topology). Judge the signal by its cell distribution and compartment; a sharply outlined plasma-membrane rim is not required by the HPA IHC profile (HPA: selective cytoplasmic expression). |
| Predominant nuclear staining in a paraffin section instead of the expected cytoplasmic pattern (HPA: tissue IHC profile). | Treat this as a discordant IHC result and check controls and staining conditions before assigning it to JPH1 (HPA: selective cytoplasmic IHC expression; general IHC practice). HPA separately reports enhanced nucleoplasmic localization in ICC-IF; that observation concerns a different assay and does not establish a nuclear pattern for tissue IHC (HPA: subcellular ICC-IF). |
| Strong signal in a cell population listed as not detected, such as adipocytes in adipose tissue (HPA: tissue IHC). | Consider cross-reactivity or endogenous detection activity, especially if the signal resembles staining in the positive control (general IHC practice). Do not apply that interpretation to spermatogonia: HPA reports high staining there (HPA: High in spermatogonia). Compare the suspect section with a negative reagent control and the expected distribution before calling it JPH1 (general IHC practice). |
| Broad, diffuse colour across cells and extracellular areas, obscuring cell boundaries (general IHC practice). | This limits compartment and cell-type interpretation; nonspecific reagent binding, incomplete blocking, or excess chromogen development are general possibilities (general IHC practice). Assess the negative reagent control and whether the skeletal-myocyte pattern remains discernible (general IHC practice; HPA: High in skeletal myocytes). Diffuse colour alone does not establish the HPA cytoplasmic pattern (HPA: tissue IHC profile). |
| No detectable signal in skeletal myocytes on a slide intended as a positive control (HPA: High in skeletal myocytes). | The run cannot support a negative interpretation of test tissue until the positive control works (general IHC practice). Check that primary antibody, detection reagents and chromogen were applied and that the chosen retrieval and antibody conditions follow the validated IHC method (general IHC practice). HPA's Enhanced rating describes evidence for its reported pattern, not a guarantee for every staining run (HPA: tissue IHC reliability). |
💡Expected JPH1 appearanceCall the IHC result consistent with JPH1 when skeletal myocytes show strong cytoplasmic staining (HPA: High in skeletal myocytes; HPA: selective cytoplasmic expression); predominant nuclear colour or strong staining in an HPA-listed negative cell type warrants investigation (HPA: tissue IHC; general IHC practice).
| Cellular location and topology (UniProt Q9HDC5 topology). | JPH1 has a cytoplasmic region at residues 1–639 and a transmembrane segment at 640–660 anchored in endoplasmic or sarcoplasmic reticulum membrane (UniProt Q9HDC5 topology). Interpret section staining at the cytoplasmic and membrane-junction level (UniProt Q9HDC5 topology; HPA: cytoplasmic tissue IHC), without assuming that light microscopy will resolve individual triads (general microscopy practice). |
| Tissue and cell context (HPA: tissue IHC). | HPA rates skeletal myocytes and spermatogonia High, cardiomyocytes and smooth muscle cells Low, and several listed cell types Not detected (HPA: tissue IHC). A faint heart or smooth-muscle result has a different expectation from a blank skeletal-muscle control (HPA: tissue IHC; general IHC practice). Use the named cell population when comparing sections (general IHC practice). |
| Evidence strength (HPA: tissue IHC reliability and antibody validation). | HPA labels the tissue profile Enhanced but reports medium agreement between antibody staining and RNA, with presumed off-target staining disregarded (HPA: tissue IHC reliability). HPA lists HPA008996 and HPA009413 as IHC Enhanced, and CAB013461 as IHC Supported (HPA: antibody validation). These labels support comparison with the reported pattern; they do not identify every unexpected stained cell as JPH1 (HPA: tissue IHC reliability). |
| IF/ICC Q: Should nucleoplasmic fluorescence define the expected paraffin-section IHC result? (HPA: subcellular ICC-IF). | A: No. HPA reports enhanced nucleoplasmic localization in ICC-IF images from A-431, U-251MG and U2OS, while its tissue IHC profile describes selective cytoplasmic expression, most abundant in skeletal muscle (HPA: subcellular ICC-IF; HPA: tissue IHC). Interpret each assay against its own reported pattern (general assay practice). |
| Situation | Likely cause | Next action |
|---|---|---|
| Skeletal-muscle positive control is blank (HPA: High in skeletal myocytes). | A failed reagent step or unsuitable retrieval or primary-antibody conditions can remove visible IHC signal (general IHC practice). | Confirm the antibody and detection sequence, then check retrieval and dilution against the IHC-validated antibody's method (general IHC practice). Repeat with a skeletal-muscle positive control before interpreting negative study sections (HPA: High in skeletal myocytes; general IHC practice). |
| Colour is diffuse across the section rather than concentrated in expected cells (HPA: tissue IHC profile). | Nonspecific reagent binding or excessive chromogen development can obscure localization (general IHC practice). | Inspect the negative reagent control, review blocking and washes, and shorten chromogen development if overdevelopment is evident (general IHC practice). Confirm that any remaining positive signal maps to skeletal myocytes or another reported positive cell type (HPA: tissue IHC). |
| Strong nuclear IHC signal dominates the skeletal-muscle section (HPA: cytoplasmic tissue IHC profile). | The compartment conflicts with HPA tissue IHC, even though HPA reports nucleoplasmic localization in ICC-IF (HPA: tissue IHC; HPA: subcellular ICC-IF). | Review control slides and primary-antibody conditions, then score the nuclear result as unresolved for tissue IHC unless independently supported (general IHC practice; HPA: tissue IHC profile). Do not use the ICC-IF localization alone to validate nuclear chromogenic staining (HPA: subcellular ICC-IF). |
| An HPA-listed negative cell type stains strongly (HPA: tissue IHC). | Off-target antibody staining or endogenous detection activity may account for an unexpected chromogenic signal (HPA: reliability note on presumed off-target staining; general IHC practice). | Compare a negative reagent control and a skeletal-muscle positive control, then check whether the unexpected colour tracks the cell type or the detection system (general IHC practice; HPA: High in skeletal myocytes). Keep the finding provisional (general IHC practice). |
| Heart or smooth-muscle staining is faint while skeletal muscle stains strongly (HPA: tissue IHC). | This relative intensity matches HPA's Low levels in cardiomyocytes and smooth muscle cells versus High in skeletal myocytes (HPA: tissue IHC). | Score each named cell population against its reported level and the same run's positive control (HPA: tissue IHC; general IHC practice). Investigate a technical failure if the skeletal-muscle control also loses signal (HPA: High in skeletal myocytes; general IHC practice). |
Experiment evidence
IHC & IF images showing JPH1 staining
A08565 · IHCImmunohistochemistry of JPH1 in mouse skeletal muscle tissue with JPH1 antibody at 2.5 μg/mL.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000104369-JPH1/tissue/cerebral+cortex#imid_3011688
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000104369-JPH1/tissue/colon#imid_3011630
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000104369-JPH1/tissue/liver#imid_3002343
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000104369-JPH1/tissue/kidney#imid_3002344
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA008996; green = JPH1, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA008996; green = JPH1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for JPH1 IHC & IF
🧪Run skeletal muscle first; myocytes should stain (HPA: High in skeletal-muscle myocytes). Use adipose tissue as the negative tissue, with adipocytes expected to lack detectable staining (HPA: Not detected in adipocytes); on the positive slide, nonmyocyte areas should lack staining comparable to myocytes, subject to morphology-based review (HPA: High in myocytes).
Positive control tissue: Skeletal muscle (Myocytes, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show JPH1 in A-431, U-251MG, U2OS, with annotated localisation: Nucleoplasm (enhanced) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and a host-species-matched isotype control for a monoclonal primary, or matched nonimmune IgG for a polyclonal primary (standard IHC practice). If available, use JPH1-knockout tissue as a biological negative; quench endogenous peroxidase for chromogenic IHC and assess skeletal-muscle autofluorescence if using IF (standard IHC/IF practice).
⚠️Feasibility: A target-specific fixation window and antigen-retrieval dependency are unreported in the supplied target and application evidence; the selected A08565 skeletal-muscle IHC caption does not state a fixative (selected-SKU caption). The evidence does not establish whether frozen sections or IF are easier for JPH1 (supplied target/application evidence). For IF interpretation, reconcile any nuclear signal with the reported membrane-associated topology and the HPA nucleoplasm ICC-IF localization using appropriate controls (UniProt Q9HDC5 topology; HPA: Nucleoplasm enhanced).
HPA tissue IHC evidence for JPH1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium 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 |
|---|---|---|---|---|
| Skeletal muscle | Myocytes | High | Protein (IHC) | HPA → |
| Testis | Spermatogonia cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced JPH1 IHC Tips
Troubleshoot JPH1 staining in paraffin sections by checking retrieval, compartment, and muscle cell identity against the available tissue evidence.
What retrieval should I try first when JPH1 staining is weak in paraffin sections?
Start with citrate buffer at pH 6.0, heated to 95–98 °C for 20 minutes (page retrieval setting). Let sections cool in the buffer, then compare them with an otherwise identical section processed without retrieval to judge whether heating improves specific staining (standard IHC practice). Use skeletal muscle as the positive tissue, because JPH1 is abundant there and myocytes stain strongly in tissue IHC (UniProt Q9HDC5 tissue specificity; HPA: High in skeletal muscle myocytes). If signal remains weak, test a different retrieval buffer as a documented fallback on adjacent sections, holding antibody concentration and detection conditions constant (standard IHC practice).
Could fixation explain weak JPH1 staining in my paraffin sections?
The supplied JPH1 tissue image reports antibody A08565 at 2.5 µg/mL, but gives no fixative or fixation time, so target-specific fixation sensitivity is unknown (selected A08565 tissue-IHC caption). Record the fixative, time before fixation, specimen thickness, and processing schedule for each block before comparing staining across specimens (standard IHC practice). On adjacent sections, apply the same citrate pH 6.0, 95–98 °C, 20-minute retrieval to limit procedural differences (page retrieval setting; standard IHC practice). Compare preserved skeletal muscle myocytes within each specimen and assess morphology alongside signal; staining differences alone cannot establish a JPH1-specific fixation effect (HPA: High in skeletal muscle myocytes; standard IHC practice).
Where should convincing JPH1 chromogenic staining appear in skeletal muscle?
Expect staining associated with skeletal muscle myocytes, where tissue IHC reports high JPH1 expression and a selective cytoplasmic pattern (HPA: High in skeletal muscle myocytes; HPA tissue IHC profile). JPH1 links surface and sarcoplasmic reticulum membranes, with residues 640–660 forming a membrane anchor and residues 1–639 on the cytoplasmic side (UniProt Q9HDC5 function and topology). In skeletal muscle, its reported position near the A–I band junction is based on similarity, so routine chromogenic sections need not resolve that pattern (UniProt Q9HDC5 subcellular annotation; standard IHC practice). Assess cytoplasmic distribution and myocyte identity together, and investigate isolated nuclear or diffuse extracellular deposits before assigning them to JPH1 (HPA tissue IHC profile; standard IHC practice).
How should unknown epitope placement affect my JPH1 IHC controls?
The supplied record lists 0 isoforms and places JPH1 residues 1–639 on the cytoplasmic side of its C-terminal membrane anchor (UniProt Q9HDC5 isoforms and topology). It also lists phosphorylation sites, including serines 157, 216, and 220, but supplies no antibody epitope map or evidence that these modifications alter staining (UniProt Q9HDC5 modified residues; supplied antibody evidence). Keep retrieval and antibody concentration fixed while comparing adjacent skeletal muscle sections, so procedural variation does not masquerade as epitope behavior (HPA: High in skeletal muscle myocytes; standard IHC practice). If staining fails, verify the antibody's documented epitope and IHC validation before attributing the result to an isoform or phosphorylation state (standard IHC practice).
How can I assess JPH1 by multiplex IF alongside this IHC workflow?
For the separate IF/ICC workflow, pair JPH1 with an independently validated myocyte marker and compare its pattern with skeletal muscle cells, where tissue IHC reports high expression (HPA: High in skeletal muscle myocytes; standard IF practice). Choose a fluorophore in a channel with low measured tissue autofluorescence, and acquire unstained and single-stained controls at identical exposure settings (standard IF practice). If the antibody epitope lies within JPH1's cytoplasmic residues 1–639, test permeabilisation sufficient to admit antibody to that side of the membrane (UniProt Q9HDC5 topology; standard IF practice). The supplied evidence gives no JPH1-specific IF/ICC fixation condition, so optimise fixation empirically and compare morphology and signal in matched specimens (supplied IF/ICC evidence; standard IF practice).
What should I check when brown signal extends beyond skeletal muscle myocytes?
Compare the staining with a matched no-primary control and inspect section edges, folds, and damaged regions for deposits that do not follow myocyte boundaries (standard IHC practice). Quench endogenous peroxidase before chromogenic detection, and check whether residual enzyme activity contributes to brown signal in the control (standard IHC practice). Titrate the catalog antibody around the reported 2.5 µg/mL tissue-image concentration while holding retrieval and detection constant; the caption does not establish an optimum for your sections (selected A08565 tissue-IHC caption; standard IHC practice). Use skeletal muscle myocytes as a positive reference and an appropriately processed low-expression tissue as a comparison, while accounting for presumed off-target staining noted in the tissue survey (HPA tissue IHC profile and reliability).
How should I quantify JPH1 staining across paraffin muscle sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define skeletal muscle myocytes as the analysis population and exclude folds, tears, and necrotic regions before scoring (HPA: High in skeletal muscle myocytes; standard IHC practice). For chromogenic sections, report an H-score from staining intensity and the percentage of positive myocytes, or report positive-cell percentage if intensity is unreliable (standard IHC practice). Normalise counts to the number of evaluable myocytes; for area-based measurements, report positive area per mm² of viable muscle and retain matched acquisition and threshold settings (standard IHC practice). Record retrieval, antibody concentration, and batch with each result, because the available 2.5 µg/mL caption documents one tissue image rather than a universal scoring condition (selected A08565 tissue-IHC caption; standard IHC practice).
When does a positive JPH1 IHC result need further validation?
A plausible result stains intact skeletal muscle myocytes in a cytoplasmic or membrane-associated pattern, consistent with the tissue survey and JPH1's membrane-linking role (HPA tissue IHC profile; UniProt Q9HDC5 function). Treat isolated nuclear staining cautiously: the separate cell-line IF survey reports nucleoplasmic localisation, while tissue IHC reports selective cytoplasmic muscle expression (HPA subcellular summary; HPA tissue IHC profile). Check unexpected cell types against matched controls, particularly because the tissue survey notes presumed off-target binding despite its Enhanced reliability designation (HPA tissue IHC reliability). Edge staining, necrotic deposits, or signal retained in a no-primary peroxidase control supports a technical explanation that needs resolution before biological interpretation (standard IHC practice).
Boster reagents
Best JPH1 / Junctophilin-1 IHC Antibodies
A08565 has real IHC data from mouse skeletal muscle (A08565 image caption). The catalog lists Human, Mouse, and Rat reactivity, but provides no IF image (catalog: reactivity and images).
Real IHC data
Anti-Junctophilin-1 JPH1 Antibody
Cat # A08565
A08565 renders with an IHC image of mouse skeletal muscle at 2.5 μg/mL (A08565 image caption). Its listed applications include IHC-P, and its listed reactivity covers Human, Mouse, and Rat; the supplied IHC image documents mouse tissue (catalog: applications and reactivity; A08565 image caption).
Which to pick: Choose A08565 for paraffin-section tissue IHC: IHC-P is listed, and its own image shows mouse skeletal muscle at 2.5 μg/mL (catalog: applications; A08565 image caption). The image caption does not report a fixative, and the catalog does not specify clonality (A08565 image caption; catalog: clone). No SKU has listed IF/ICC validation; for Human, Mouse, or Rat IHC, A08565 is the catalog choice by listed reactivity, with direct IHC image evidence limited to mouse skeletal muscle (catalog: applications and reactivity; A08565 image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9HDC5 (JPH1_HUMAN, Junctophilin-1).
- Human Protein Atlas. JPH1 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. JPH1 subcellular location (ICC-IF): Localized to the nucleoplasm..
- Human Protein Atlas. JPH1 antibody validation summary (3 antibodies).
- The Effects of Aging on Sarcoplasmic Reticulum-Related Factors in the Skeletal Muscle of Mice. International journal of molecular sciences 2024 — PMC10889371.
- PubMed PMID:16421571 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
- PubMed PMID:10891348 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for JPH1 (Junctophilin-1). Expected localisation Cytoplasmic staining in skeletal myocytes (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A08565 at 2.5 μg/mL (the product's own IHC validation image is shown). Controls (HPA): positive Skeletal muscle (Myocytes); 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 A08565); 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-JPH1, 2.5 μg/mL (datasheet A08565); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A08565); Caveat: Presumed off-target binding can confound staining (HPA tissue IHC); Isoform / epitope: No isoforms annotated; N-terminal region is cytoplasmic (UniProt); Regulation: Abundant in skeletal muscle; very low in heart (UniProt). Evidence: UniProt Q9HDC5, HPA tissue IHC · Enhanced, HPA ICC-IF · nucleoplasm., Datasheet A08565, PMC10889371. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/JPH1/immunohistochemistry-ihc-guide.