This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
CASQ1 / Calsequestrin-1 · IHC design guide
Design Immunohistochemistry for CASQ1
Plan CASQ1 chromogenic IHC in paraffin sections using skeletal myocytes as a high-staining reference (HPA tissue IHC). This guide covers fixation consistency, the catalog antibody’s 1:50 dilution (datasheet M05235), cytoplasmic scoring, and control selection.
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CASQ1 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 | Myocyte cytoplasm (HPA tissue IHC); SR lumen expected (UniProt) | |
| Staining pattern | Selective cytoplasmic staining in skeletal myocytes (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet M05235) | |
| Positive control | Skeletal muscle | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across samples (standard IHC practice; not target-specific) | |
| Caveat | Low staining in some nonmuscle cells may complicate scoring (HPA tissue IHC) | |
| Regulation | Expression regulation is unreported (UniProt) | |
| Isoform / epitope | No annotated isoforms; mature chain spans residues 35–396 (UniProt) |
Section 1
Recommended CASQ1 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol (datasheet M05235) with one published paraffin-section CASQ1/2 protocol (PMC12955825 methods).
| Sample | Paraffin-embedded mouse skeletal muscle tissue; fixative not specified (datasheet M05235) |
| Fixation | Image fixative and duration unreported (datasheet M05235); 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 M05235); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet M05235) |
| Primary antibody | Rabbit monoclonal (clone AEGE-3) anti-CASQ1, 1:50 (datasheet M05235) |
| Primary incubation | Overnight at 4 °C (datasheet M05235) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet M05235) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | CASQ1-positive staining in myocytes of skeletal muscle (HPA tissue IHC: High). HPA tissue profile: Selective cytoplasmic expression in skeletal muscle. No signal in the no-primary control. |
💡Decision noteStart with EDTA pH 8.0 heat retrieval (datasheet M05235); the published protocol does not specify retrieval (PMC12955825 methods).
Section 2
What Is the Expected CASQ1 Staining Pattern?
CASQ1 is a luminal protein of the sarcoplasmic reticulum, particularly the terminal cisternae of fast skeletal muscle cells; it has no transmembrane segment (UniProt P31415: subcellular location and topology). In paraffin section IHC, expect strong cytoplasmic staining of skeletal myocytes (HPA: High in skeletal muscle myocytes). HPA rates the tissue IHC pattern Enhanced, citing agreement between staining and RNA expression (HPA: tissue IHC reliability).
| Strong cytoplasmic staining in skeletal muscle myocytes, with little staining in nearby cells. | This matches the reported selective cytoplasmic tissue pattern and the sarcoplasmic reticulum location of CASQ1 (HPA: skeletal muscle myocytes High; UniProt P31415: subcellular location). Judge the distribution across myocytes as well as intensity. |
| Predominantly nuclear, extracellular, or surface outlining stain in skeletal myocytes. | These are unexpected patterns for the principal luminal sarcoplasmic reticulum pool (UniProt P31415: subcellular location and topology). Check the negative detection control and examine whether the expected myocyte cytoplasm is also stained before interpreting the signal as CASQ1 (standard IHC practice). |
| Strong staining in adipocytes, hematopoietic cells, or glandular cells while skeletal myocytes stain weakly. | HPA reports CASQ1 as Not detected in adipose adipocytes, bone marrow hematopoietic cells, and several glandular cell populations (HPA: tissue IHC). Consider antibody cross-reactivity or endogenous detection activity; compare antibody omission and detection controls (standard IHC practice). |
| A uniform wash of color across myocytes, connective tissue, and empty spaces. | A field-wide signal cannot establish selective myocyte staining (HPA: selective cytoplasmic expression in skeletal muscle). Review blocking, washing, detection reagent, and counterstain conditions; compare the antibody-omission control on the same tissue (standard IHC practice). |
| No convincing myocyte signal in an adequately preserved skeletal muscle section. | This conflicts with HPA's High myocyte staining (HPA: skeletal muscle myocytes High). First verify that the section contains recognizable myocytes and that the detection run worked; then assess antibody dilution and retrieval conditions using controls (standard IHC practice). |
💡Expected CASQ1 appearanceCall the result positive when skeletal myocytes show strong cytoplasmic staining in the expected sarcoplasmic reticulum distribution (HPA: High myocyte staining; UniProt P31415: subcellular location); isolated nuclear, extracellular, or unrelated-cell staining is suspect (standard IHC interpretation).
| Tissue and cell choice | Skeletal muscle myocytes provide the reported high-signal reference (HPA: High). Adipose adipocytes and bone marrow hematopoietic cells are reported Not detected and can help assess selectivity, subject to comparable processing and detection (HPA: tissue IHC; standard IHC practice). |
| Compartment and topology | CASQ1 occupies sarcoplasmic reticulum luminal spaces and lacks a transmembrane segment (UniProt P31415: subcellular location and topology). Read chromogenic signal as a cytoplasmic distribution at tissue-section resolution; the stain alone does not resolve individual terminal cisternae (standard IHC interpretation). |
| Antibody validation | HPA assigns Enhanced tissue IHC reliability and lists Enhanced IHC status for HPA007845, HPA026823, and CAB015170 (HPA: tissue IHC; HPA: antibody validation). That supports the reported tissue pattern; it does not validate every reagent or staining condition (standard IHC interpretation). |
| Processing and modifications | The annotated precursor has a signal peptide at residues 1–34 and a mature chain at 35–396; UniProt also lists glycosylation at residue 350 and modified residues (UniProt P31415: processing and modifications). Without a supplied antibody epitope, their effect on this IHC assay cannot be predicted. |
| Antigen retrieval | Use the selected antibody's documented IHC-P conditions as the starting point, and evaluate changes against positive and negative controls (standard IHC practice). Neither HPA tissue staining nor UniProt topology establishes CASQ1-specific retrieval requirements or fixation sensitivity. |
| Q: How should an IF/ICC nuclear or mitochondrial result be read? | A: HPA reports approved nucleoplasm and mitochondrial ICC-IF locations, including images from RT-4, while its tissue IHC profile is selective cytoplasmic staining in skeletal muscle (HPA: subcellular ICC-IF; HPA: tissue IHC). Interpret each assay in its own context; the ICC-IF finding does not change the expected paraffin-section IHC pattern. |
| Situation | Likely cause | Next action |
|---|---|---|
| Skeletal myocytes are blank, and the positive control is also blank. | A run-wide staining or detection failure is possible (standard IHC practice). | Confirm reagent delivery and detection steps, then repeat with a working positive control and the antibody's documented IHC-P conditions (standard IHC practice). |
| The positive control stains, but the test section's myocytes remain blank. | Section quality, tissue identification, or assay conditions may differ between sections (standard IHC practice); HPA reports High staining in skeletal myocytes (HPA: tissue IHC). | Confirm myocyte morphology and compare section handling, dilution, and retrieval with the positive control before calling the tissue negative (standard IHC practice). |
| Color appears in unrelated cells and in the antibody-omission control. | Endogenous detection activity or detection-system background may account for the signal (standard IHC practice). | Review the detection chemistry and its blocking step; repeat with appropriate detection controls before assigning that staining to CASQ1 (standard IHC practice). |
| Unrelated cells stain strongly, but the antibody-omission control is clear. | Antibody-associated off-target staining is possible, especially where HPA reports CASQ1 as Not detected (HPA: tissue IHC; standard IHC interpretation). | Compare the cell distribution with a skeletal muscle positive control and, where available, an independently validated antibody (HPA: antibody validation; standard IHC practice). |
| The section has diffuse color that obscures myocyte boundaries. | Excess staining or incomplete background removal may prevent assessment of the selective pattern (standard IHC practice; HPA: selective cytoplasmic tissue IHC). | Compare the antibody-omission control, then review antibody concentration, wash steps, and chromogen development while retaining a skeletal muscle positive control (standard IHC practice). |
| Nuclei dominate the stain and myocyte cytoplasm is weak. | The pattern differs from HPA's selective cytoplasmic skeletal muscle IHC and the luminal sarcoplasmic reticulum location (HPA: tissue IHC; UniProt P31415: subcellular location). | Check counterstain and antibody-omission controls, then reassess localization with a validated IHC antibody before scoring the nuclear signal as CASQ1 (standard IHC practice; HPA: antibody validation). |
Experiment evidence
IHC & IF images showing CASQ1 staining
M05235 · IHCIHC analysis of CASQ1 using anti-CASQ1 antibody (M05235). CASQ1 was detected in a paraffin-embedded section of mouse skeletal muscle tissue.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000143318-CASQ1/tissue/cerebral+cortex#imid_2397800
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000143318-CASQ1/tissue/colon#imid_2397734
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000143318-CASQ1/tissue/liver#imid_2397727
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000143318-CASQ1/tissue/kidney#imid_2397736
HPA · ICC-IF RT-4Human Protein Atlas immunofluorescence, RT-4, antibody HPA007845; green = CASQ1, blue = nucleus, red = microtubules.
HPA · ICC-IF RT-4Human Protein Atlas immunofluorescence, RT-4, antibody HPA007845; green = CASQ1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CASQ1 IHC & IF
🧪Run skeletal muscle first and look for staining in myocytes (HPA: High in skeletal-muscle myocytes). Use adipose tissue adipocytes as the negative tissue (HPA: Not detected in adipocytes); on the positive slide, morphologically distinct nonmyocyte cells should provide a background comparison, checked against the no-primary control (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 CASQ1 in RT-4, with annotated localisation: Nucleoplasm (approved), Mitochondria (approved) (HPA subcellular).
Technical controls: Include no-primary (secondary-only) and concentration-matched rabbit isotype controls for the rabbit primary, plus CASQ1-knockout tissue or a validated immunizing-peptide block if available (caption: rabbit anti-CASQ1 M05235; standard IHC controls). For HRP/DAB detection, quench endogenous peroxidase and inspect the no-primary skeletal-muscle section for residual chromogen background (caption: HRP/DAB detection; standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected M05235 paraffin-section caption does not state the fixative (caption: fixative not stated). Heat retrieval in EDTA at pH 8.0 was used for the mouse skeletal-muscle IHC example; the evidence does not establish that retrieval is required for all specimens (caption: EDTA heat retrieval). The supplied evidence does not establish whether frozen sections or IF are easier; for this skeletal-muscle HRP/DAB assay, assess residual fibre-associated background with the no-primary control (caption: skeletal-muscle HRP/DAB example; standard IHC practice).
HPA tissue IHC evidence for CASQ1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — High consistency between antibody staining and RNA expression data.). 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 → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Epididymis | Glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CASQ1 IHC Tips
Use skeletal muscle myocytes as the reference pattern while checking retrieval, compartment, and background controls before scoring CASQ1 staining (HPA: skeletal muscle myocytes High).
How should I retrieve CASQ1 in paraffin skeletal muscle sections?
Start with heat-mediated retrieval in EDTA pH 8.0 for paraffin skeletal muscle sections (datasheet M05235). The selected antibody produced a chromogenic CASQ1 image after that retrieval, followed by 1:50 primary antibody overnight at 4°C (datasheet M05235). If staining is weak, compare retrieval durations on adjacent sections while keeping antibody dilution, development time, and section thickness constant (standard IHC practice). Assess signal in myocytes against a no-primary control, and reject conditions that increase diffuse background or damage muscle architecture (HPA: skeletal muscle myocytes High; standard IHC practice).
Can fixation explain weak CASQ1 staining in my paraffin sections?
Target-specific sensitivity to fixation is unknown: the selected paraffin-section caption does not state a fixative (datasheet M05235). Record the actual fixative and fixation interval for each specimen, then compare matched sections processed with the same EDTA pH 8.0 retrieval and detection settings (datasheet M05235; standard IHC practice). A weak result after prolonged or uneven fixation warrants a controlled comparison, but it does not establish that CASQ1 itself is fixation sensitive (standard IHC practice). Judge any recovery by preserved myocyte morphology, reduced no-primary background, and reproducible cytoplasmic signal across comparable areas (HPA: selective cytoplasmic expression in skeletal muscle; standard IHC practice).
Where should CASQ1 staining appear within positive skeletal muscle cells?
Expect cytoplasmic staining in skeletal muscle myocytes by chromogenic IHC (HPA: selective cytoplasmic expression in skeletal muscle; HPA: myocytes High). CASQ1 is associated with sarcoplasmic reticulum terminal cisternae luminal spaces in fast skeletal muscle cells, and its record lists no transmembrane segment (UniProt P31415: subcellular location and topology). A granular or reticular cytoplasmic pattern can therefore be assessed against myocyte structure, although routine chromogenic sections cannot resolve the lumen of individual cisternae (UniProt P31415: subcellular location; standard IHC practice). Treat isolated nuclear or surface-only DAB staining cautiously and compare it with a no-primary control before calling it CASQ1 (HPA: tissue IHC pattern; standard IHC practice).
Could CASQ1 processing or epitope accessibility change my IHC result?
The CASQ1 record lists a 1–34 signal sequence and a 35–396 mature chain, with no annotated isoforms (UniProt P31415: processing and isoforms). It also lists glycosylation at residue 350 and several modified residues, but the selected antibody’s epitope is not specified in the supplied caption (UniProt P31415: glycosylation and modified residues; datasheet M05235). Do not assign weak staining to cleavage or a particular modification without epitope mapping and supporting controls (standard IHC practice). For a questionable pattern, compare an independently validated CASQ1 antibody recognizing a documented different epitope on adjacent sections, using the same myocyte regions and background controls (standard IHC practice).
How can IF help check an ambiguous CASQ1 IHC pattern?
Use IF as a separate, validated assay and multiplex CASQ1 with a skeletal muscle myocyte marker to test whether signal lies in the expected cells (HPA: skeletal muscle myocytes High; standard IF practice). Choose fluorophores outside the strongest tissue autofluorescence and inspect single-label and no-primary controls before interpreting overlap (standard IF practice). Because CASQ1 occupies the sarcoplasmic reticulum lumen and has no transmembrane segment, optimize permeabilisation for access to an internal epitope once the antibody’s epitope is known (UniProt P31415: subcellular location and topology; standard IF practice). HPA also reports nucleoplasm and mitochondria in subcellular imaging, so compare unexpected compartments with orthogonal controls rather than transferring an IF pattern directly to chromogenic tissue scoring (HPA: subcellular summary; standard IHC practice).
What should I change if DAB obscures myocyte CASQ1 staining?
The selected paraffin-section image used 10% goat serum blocking, a 1:50 primary antibody overnight at 4°C, and peroxidase-based DAB detection (datasheet M05235). Include a no-primary section, block endogenous peroxidase, and shorten DAB development if background obscures cell boundaries (standard IHC practice). If diffuse staining persists, compare a primary-antibody dilution series while keeping retrieval at EDTA pH 8.0 and using the same exposure to detection reagents (datasheet M05235; standard IHC practice). Score improvement by retention of myocyte cytoplasmic signal with lower interstitial and no-primary staining, rather than by overall darkness alone (HPA: skeletal muscle myocytes High; standard IHC practice).
How should I quantify CASQ1 staining across skeletal muscle sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define myocyte regions before scoring because the reference tissue pattern is selective cytoplasmic expression in skeletal muscle (HPA: tissue IHC profile). Report the percentage of positive myocytes and an intensity-based H-score, using fixed thresholds and a common DAB development interval across specimens (standard IHC practice). For area-based analysis, report positive myocyte area per mm² of evaluable muscle and normalize to the total evaluable myocyte area, rather than whole-section area containing variable nonmuscle space (standard IHC practice). Exclude folds, damaged edges, and necrotic regions by a documented rule, and retain representative images alongside the numerical results (standard IHC practice).
How do I distinguish true CASQ1 staining from IHC artefact?
A credible positive result is reproducible cytoplasmic staining in skeletal muscle myocytes, consistent with the reported tissue IHC profile and sarcoplasmic reticulum location (HPA: selective cytoplasmic expression and myocytes High; UniProt P31415: subcellular location). Staining confined to nuclei, tissue edges, necrotic areas, or cells outside the expected myocyte population needs separate verification before attribution to CASQ1 (HPA: tissue IHC profile; standard IHC practice). Compare adjacent sections with and without primary antibody, and check whether peroxidase blocking removes any misleading DAB signal (standard IHC practice). If compartment findings still conflict, use an independently validated antibody or a separately optimized IF assay to test the same region (standard IHC/IF practice).
Boster reagents
Best CASQ1 / Calsequestrin-1 IHC Antibodies
M05235 has real CASQ1 IHC data from a paraffin-embedded mouse skeletal muscle section (M05235 IHC caption). Human, mouse and rat reactivity is listed; no IF image is supplied (catalog M05235).
Real IHC data
Anti-Calsequestrin 1 Monoclonal Antibody
Cat # M05235
M05235 shows IHC staining in a paraffin-embedded mouse skeletal muscle section using a 1:50 antibody dilution (M05235 IHC caption). The catalog lists IHC application and human, mouse and rat reactivity for M05235, but supplies no IF image (catalog M05235).
Which to pick: For tissue IHC, choose M05235, a rabbit monoclonal whose caption documents heat retrieval in EDTA at pH 8.0 and overnight incubation at 1:50 in a paraffin-embedded mouse skeletal muscle section; the fixative is unreported (catalog M05235; M05235 IHC caption). For IF/ICC, M05235 has neither a listed IF application nor an IF image, so its suitability is unverified (catalog M05235). For cross-species IHC, M05235 lists human, mouse and rat reactivity, while its shown tissue IHC evidence is limited to mouse (catalog M05235; M05235 IHC caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P31415 (CASQ1_HUMAN, Calsequestrin-1).
- Human Protein Atlas. CASQ1 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. CASQ1 subcellular location (ICC-IF): Localized to the nucleoplasm and mitochondria..
- Human Protein Atlas. CASQ1 antibody validation summary (3 antibodies).
- Conformational ligand-directed targeting of calcium-dependent receptors in acute trauma. Med (New York, N.Y.) 2025 — PMC12955825.
- CCDC78: Unveiling the Function of a Novel Gene Associated with Hereditary Myopathy. Cells 2024 — PMC11394131.
- PubMed PMID:2321095 — UniProt-cited evidence.
- PubMed PMID:7945294 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CASQ1 (Calsequestrin-1). Expected localisation Myocyte cytoplasm (HPA tissue IHC); SR lumen expected (UniProt). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet M05235). Validated IHC antibody: M05235 at 1:50 (the product's own IHC validation image is shown). Controls (HPA): positive Skeletal muscle (Myocytes); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded mouse skeletal muscle tissue; fixative not specified (datasheet M05235); 2. Fixation: Image fixative and duration unreported (datasheet M05235); 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 retrieval: EDTA pH 8.0 (datasheet M05235); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet M05235); 8. Primary antibody: Rabbit monoclonal (clone AEGE-3) anti-CASQ1, 1:50 (datasheet M05235); 9. Primary incubation: Overnight at 4 °C (datasheet M05235). Watch for: Fixation: Keep fixation consistent across samples (standard IHC practice; not target-specific); Caveat: Low staining in some nonmuscle cells may complicate scoring (HPA tissue IHC); Isoform / epitope: No annotated isoforms; mature chain spans residues 35–396 (UniProt); Regulation: Expression regulation is unreported (UniProt). Evidence: UniProt P31415, HPA tissue IHC · Enhanced, HPA ICC-IF · nucleoplasm and mitochondria., Datasheet M05235, PMC12955825. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CASQ1/immunohistochemistry-ihc-guide.