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CASQ1 / Calsequestrin-1 · IHC design guide
Design Immunohistochemistry for CASQ1
Plan CASQ1 paraffin IHC around selective cytoplasmic staining in skeletal myocytes (HPA tissue IHC). The catalog antibody was used at 1:50 with chromogenic detection in a mouse skeletal muscle section (datasheet M05235).
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 skeletal myocyte cytoplasmic staining (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 sections (standard IHC practice; not target-specific) | |
| Caveat | Low neuronal staining may complicate interpretation (HPA tissue IHC) | |
| Regulation | Expression regulation unreported (UniProt) | |
| Isoform / epitope | No annotated isoforms; mature chain starts at residue 35 (UniProt) |
Section 1
Recommended CASQ1 IHC & IF Protocols
The catalog antibody protocol is accompanied by one published paraffin-section IHC protocol for CASQ1/2 staining in tissue biopsies (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 heat-mediated EDTA pH 8.0 retrieval (datasheet M05235); the published biopsy protocol does not specify retrieval (PMC12955825 methods).
Section 2
What Is the Expected CASQ1 Staining Pattern?
CASQ1 should give strong cytoplasmic staining in skeletal-muscle myocytes on IHC sections (HPA: High in skeletal-muscle myocytes; selective cytoplasmic expression). Its principal muscle location is the sarcoplasmic-reticulum lumen, especially terminal cisternae of fast skeletal-muscle cells (UniProt P31415: subcellular location). It has no transmembrane segment (UniProt P31415: topology). HPA rates the tissue IHC profile Enhanced, citing high consistency between antibody staining and RNA expression data (HPA: tissue IHC reliability).
| Strong cytoplasmic staining in skeletal-muscle myocytes, with little staining in neighboring nonmyocytes. | This matches the reported positive tissue and cell type (HPA: High in skeletal-muscle myocytes). At light-microscope resolution, a cytoplasmic result is compatible with CASQ1 in the sarcoplasmic-reticulum lumen; the IHC pattern alone does not resolve terminal cisternae (UniProt P31415: subcellular location). |
| Predominantly nuclear staining, or a membrane-outline pattern, in skeletal-muscle myocytes. | Treat this as a compartment mismatch requiring validation: tissue IHC is reported as cytoplasmic (HPA: tissue IHC profile), while UniProt places muscle CASQ1 chiefly in the sarcoplasmic-reticulum lumen and reports no transmembrane segment (UniProt P31415: location and topology). |
| Strong staining in adipocytes, hematopoietic cells, or glandular cells while myocytes are weak. | Review specificity and detection controls. Those cell populations were reported as Not detected in the listed HPA tissues (HPA: adipose tissue, bone marrow, cervix). Cross-reactivity or endogenous detection activity are possible explanations under general IHC practice; this pattern does not establish CASQ1 expression. |
| Diffuse color across the section, including cells expected to be negative. | The loss of cell-type contrast makes CASQ1 interpretation unreliable. HPA reports selective cytoplasmic expression in skeletal muscle and Not detected staining in several other cell types (HPA: tissue IHC profile). Under general IHC practice, assess nonspecific antibody binding and detection-system background. |
| No staining in skeletal-muscle myocytes on an otherwise interpretable section. | This conflicts with the HPA High result for those cells (HPA: skeletal-muscle myocytes). Check the positive control, antibody application, retrieval, and chromogenic detection as general IHC troubleshooting steps. The supplied sources do not establish a CASQ1-specific retrieval requirement or fixation sensitivity. |
💡Expected CASQ1 appearanceCall a section positive when skeletal-muscle myocytes show high cytoplasmic signal (HPA: skeletal-muscle myocytes); strong nuclear, membrane-outline, or HPA-negative-cell staining warrants investigation rather than a CASQ1-positive call (HPA: tissue IHC profile; UniProt P31415: location and topology).
| Cell-type distribution | Skeletal-muscle myocytes are a positive reference (HPA: High); adipocytes and bone-marrow hematopoietic cells are reported Not detected (HPA: tissue IHC). HPA also lists low staining in some neuronal, glandular, and respiratory epithelial cells, so every nonmuscle signal should not automatically be called false. |
| Subcellular interpretation | UniProt places CASQ1 in the sarcoplasmic-reticulum lumen of fast skeletal-muscle cells and also lists ER and mitochondrial-matrix locations (UniProt P31415: subcellular location). HPA's tissue IHC summary supports a cytoplasmic reading, without resolving those organelles (HPA: tissue IHC profile). |
| IHC antibody evidence | The listed antibodies HPA007845, HPA026823, and CAB015170 each have Enhanced IHC status (HPA: antibody validation). This supports the reported tissue pattern; it does not make an unexpected compartment or cell type positive without controls. |
| Protein processing and epitope | UniProt annotates a signal peptide at residues 1–34 and the mature chain at 35–396 (UniProt P31415: processing). No antibody epitope is supplied, so these annotations cannot predict recognition after retrieval, or explain a weak section by themselves. |
| IF/ICC Q&A: should nuclear or mitochondrial IF be expected in this IHC guide? | HPA approves nucleoplasm and mitochondria as ICC-IF locations in RT-4 images (HPA: subcellular ICC-IF), but tissue IHC reports selective cytoplasmic skeletal-muscle staining (HPA: tissue IHC). HPA007845 has Uncertain ICC status (HPA: antibody validation). Interpret IF on its own guide page; no IF protocol is established here. |
| Situation | Likely cause | Next action |
|---|---|---|
| Skeletal-muscle myocytes show no chromogenic signal. | The result disagrees with HPA's High myocyte staining (HPA: tissue IHC); a failed staining or detection step is possible under general IHC practice. | Run a known-positive skeletal-muscle section alongside the sample. Verify primary-antibody application and chromogen development, then review retrieval against the antibody's IHC-P instructions. Do not assume CASQ1-specific fixation sensitivity: none is supplied. |
| Myocytes are weak, but the entire section has a faint wash of color. | Low contrast may reflect general IHC background or insufficient usable signal; the supplied sources do not identify a CASQ1-specific chemical cause. | Compare with a no-primary control, inspect blocking and wash steps, and adjust antibody or detection conditions only within validated IHC-P guidance. Score the myocyte pattern against the HPA High reference after background is controlled (HPA: skeletal-muscle myocytes). |
| Nuclei dominate the stain in a skeletal-muscle section. | The compartment conflicts with HPA's cytoplasmic tissue pattern (HPA: tissue IHC profile). HPA's approved nucleoplasmic observation comes from ICC-IF, a different application (HPA: subcellular ICC-IF). | Check the tissue positive control and a no-primary control. Reassess specificity before scoring nuclear IHC as CASQ1; the ICC-IF observation alone cannot validate that tissue IHC pattern. |
| Adipocytes or bone-marrow hematopoietic cells stain as strongly as myocytes. | Those cells are reported Not detected by HPA, whereas skeletal-muscle myocytes are High (HPA: tissue IHC). Cross-reactivity or endogenous detection activity remain possibilities under general IHC practice. | Compare matched control sections, including a no-primary control, and check whether color appears without primary antibody. Recheck the antibody's IHC validation before attributing the unexpected cell staining to CASQ1. |
| An apparent membrane outline replaces the myocyte cytoplasmic pattern. | A membrane-dominant result does not fit the reported tissue IHC profile (HPA: selective cytoplasmic expression) or the luminal muscle location and absent transmembrane segment (UniProt P31415: location and topology). | Inspect section morphology and control staining, then repeat with the IHC-validated antibody under its documented conditions. Record the compartment as discordant until a reproducible, specifically controlled result supports interpretation. |
| A nonmuscle section shows a few weakly stained cells. | HPA lists Low staining in several nonmuscle cell populations, including neuronal and respiratory epithelial cells (HPA: tissue IHC). Weak signal therefore needs cell-specific review rather than an automatic false-positive label. | Identify the cell type and compare its HPA level with the observed intensity. Check background controls and require the expected strong skeletal-muscle myocyte signal on the positive control before interpreting weak nonmuscle staining. |
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; myocytes should show strong CASQ1 staining (HPA: High in skeletal muscle myocytes). Use adipose tissue as a negative tissue (HPA: Not detected in adipocytes), and assess non-myocyte cells on the skeletal muscle slide for background rather than scoring them as positive myocytes (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 a no-primary, secondary-only control; a concentration-matched rabbit IgG isotype control; and, if available, CASQ1 knockout tissue or an immunizing-peptide block as a biological specificity control (caption: rabbit primary antibody; standard IHC practice). Block endogenous peroxidase and check for residual myofiber background before interpreting DAB staining (caption: peroxidase secondary and DAB; standard IHC practice).
⚠️Feasibility: The selected paraffin-section caption reports heat retrieval in EDTA at pH 8.0, followed by a 1:50 primary dilution overnight at 4°C; retrieval dependence beyond those reported conditions is unreported (M05235 tissue-IHC caption). The fixative and any target-specific fixation window or fixation effect are unreported; the caption’s paraffin section does not establish fixation (M05235 tissue-IHC caption). No matched comparison establishes whether frozen sections or IF are easier; for skeletal muscle IHC, assess myofiber background when interpreting the DAB signal (supplied evidence: no matched comparison; caption: DAB detection; standard IHC practice). The selected M05235 tissue-IHC caption documents paraffin sections, but does not specify the fixative (selected-SKU IHC image M05235).
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 the paraffin-section IHC example as the starting workflow, then judge CASQ1 staining against skeletal muscle expression and sarcoplasmic reticulum localisation (datasheet M05235; HPA tissue IHC; UniProt P31415).
How should I retrieve CASQ1 in paraffin sections when skeletal muscle staining is weak?
Start with heat-mediated retrieval in EDTA buffer at pH 8.0 for paraffin sections (datasheet M05235). The documented mouse skeletal muscle example then used 10% goat serum and antibody at 1:50 overnight at 4°C (datasheet M05235). Run a known skeletal muscle section alongside the test section, and hold detection conditions constant while adjusting retrieval time within the laboratory’s validated range (HPA: high in skeletal muscle myocytes; standard IHC practice). If staining remains weak, compare a second validated retrieval condition on adjacent sections and assess both muscle signal and tissue preservation before adopting it (standard IHC practice).
Can I choose a fixative based on the published CASQ1 staining pattern?
The selected paraffin-section caption does not state a fixative, so CASQ1-specific sensitivity to fixation is unknown for this antibody (datasheet M05235). Establish a consistent fixation and processing schedule for test and control sections, then record those conditions when comparing staining runs (standard IHC practice). If signal varies, compare sections with documented processing histories using the same EDTA pH 8.0 retrieval and 1:50 antibody conditions (datasheet M05235; standard IHC practice). Evaluate staining together with morphology and a skeletal muscle positive control; neither the reported tissue pattern nor CASQ1 topology establishes a preferred fixative (HPA tissue IHC; UniProt P31415 topology).
Which staining compartment should count as convincing CASQ1 expression in skeletal muscle?
In skeletal muscle sections, expect cytoplasmic staining in myocytes, consistent with the selective tissue IHC profile (HPA: high in skeletal muscle myocytes). CASQ1 occupies the sarcoplasmic reticulum lumen, especially terminal cisternae of fast skeletal muscle cells, and has no transmembrane segment (UniProt P31415 localisation and topology). At chromogenic light-microscope resolution, score staining within intact myocyte cytoplasm without claiming that DAB alone resolves individual terminal cisternae (UniProt P31415 localisation; standard IHC practice). Compare questionable signal with a skeletal muscle control processed using EDTA pH 8.0 retrieval, and document any predominantly nuclear pattern separately (datasheet M05235; HPA tissue IHC; standard IHC practice).
How do CASQ1 processing and epitope uncertainty affect antibody interpretation?
CASQ1 has a signal peptide at residues 1–34 and a mature chain spanning 35–396; no alternative isoforms are annotated in the supplied record (UniProt P31415 processing and isoforms). The record also lists glycosylation at residue 350 and phosphorylation sites, but the antibody’s epitope is unspecified (UniProt P31415 modifications; datasheet M05235). Check the antibody’s documented immunogen or epitope before attributing a staining change to processing or modification, and keep claims limited to what that information supports (standard IHC practice). For this antibody, assess specificity through the reported skeletal muscle distribution and appropriate negative controls rather than assigning a particular epitope from the image (HPA tissue IHC; datasheet M05235; standard IHC practice).
How should I plan a confirmatory CASQ1 fluorescence experiment alongside tissue IHC?
Treat fluorescence as a separate assay: the selected antibody example documents chromogenic paraffin-section IHC, while the HPA subcellular record reports ICC/IF images in RT-4 cells (datasheet M05235; HPA subcellular). For skeletal muscle tissue, multiplex CASQ1 with a validated myocyte marker and compare its distribution with the IHC pattern in myocytes (HPA: high in skeletal muscle myocytes; standard IF practice). Select spectrally separated fluorophores after checking tissue autofluorescence, and include single-label and no-primary controls (standard IF practice). Because CASQ1 is luminal and lacks a transmembrane segment, optimise permeabilisation for antibody access to that compartment; the epitope itself is unspecified (UniProt P31415 topology and localisation; datasheet M05235; standard IF practice).
What should I check when DAB staining obscures CASQ1 in muscle sections?
The documented IHC example blocked with 10% goat serum, used antibody at 1:50 overnight at 4°C, and developed an HRP/DAB signal (datasheet M05235). If diffuse colour masks myocytes, compare a no-primary control and shorten DAB development or adjust antibody concentration using matched sections (standard IHC practice). Include an endogenous peroxidase block as part of the general HRP workflow, and inspect folds, section edges and damaged areas before interpreting colour as antigen signal (standard IHC practice). Judge remaining staining against the expected selective cytoplasmic expression in skeletal muscle myocytes, using similarly processed control tissue (HPA tissue IHC; standard IHC practice).
How can I score CASQ1 IHC without confusing muscle content with expression? ⚠ ANSWER MARKED FOR VERIFICATION
Define intact myocytes as the scoring population and use a consistent region-selection rule across sections (HPA: high in skeletal muscle myocytes; standard IHC practice). Record the percentage of positive myocytes and staining intensity, or calculate an H-score as the sum of each intensity category’s percentage multiplied by its category value (standard IHC practice). Normalise area-based positive staining to viable skeletal muscle area, reporting density per mm² when counts or areas are used (standard IHC practice). Keep retrieval, 1:50 antibody incubation and DAB development comparable across batches, and exclude folds and necrotic regions by a prespecified rule (datasheet M05235; standard IHC practice).
When does a CASQ1-positive DAB pattern warrant further validation?
A persuasive tissue result shows cytoplasmic signal in intact skeletal muscle myocytes, where the tissue IHC profile reports high expression (HPA tissue IHC). CASQ1 is associated with the sarcoplasmic reticulum lumen, so dominant nuclear staining in tissue warrants review, although the separate HPA ICC/IF record lists approved nucleoplasm and mitochondria locations (UniProt P31415 localisation; HPA subcellular). Signal concentrated at section edges, folds or necrotic areas, or persisting without primary antibody, should prompt checks for staining artefact and endogenous enzyme activity (standard IHC practice). Reassess uncertain regions against morphology, a matched skeletal muscle control and the documented EDTA pH 8.0 retrieval condition (HPA tissue IHC; datasheet M05235; standard IHC practice).
Boster reagents
Best CASQ1 / Calsequestrin-1 IHC Antibodies
M05235 has real IHC data from mouse skeletal muscle paraffin sections (M05235 IHC caption). Listed reactivity covers human, mouse and rat (catalog: M05235 reactivity).
Real IHC data
Anti-Calsequestrin 1 Monoclonal Antibody
Cat # M05235
M05235 is a rabbit monoclonal antibody listed for IHC, with an image from mouse skeletal muscle paraffin sections (catalog: M05235 host and clone; M05235 applications; M05235 IHC caption). Its listed reactivity is human, mouse and rat; no IF image is supplied (catalog: M05235 reactivity and IF images).
Which to pick: For tissue IHC, choose M05235: its own image caption documents paraffin sections, EDTA retrieval at pH 8.0 and a 1:50 primary dilution; the fixative is unreported (M05235 IHC caption). There is no supported IF/ICC choice among the supplied products because M05235 has no listed IF/ICC application or IF image (catalog: M05235 applications and IF images). For cross-species planning, M05235 lists human, mouse and rat reactivity, while its IHC image documents mouse tissue only (catalog: M05235 reactivity; 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 sections (standard IHC practice; not target-specific); Caveat: Low neuronal staining may complicate interpretation (HPA tissue IHC); Isoform / epitope: No annotated isoforms; mature chain starts at residue 35 (UniProt); Regulation: Expression regulation 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.