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BCAS4 / Breast carcinoma-amplified sequence 4 · IHC design guide
Design Immunohistochemistry for BCAS4
For BCAS4 paraffin IHC, expect general cytoplasmic staining and consider appendix glandular cells as a high-staining reference (HPA tissue IHC). Start the catalog antibody at 5 μg/mL (datasheet), and interpret scoring in light of the uncertain tissue profile (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
BCAS4 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 | General cytoplasmic tissue staining (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining in glandular and other cells (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Appendix+4 more · see all | |
| Negative control | None in HPA (detected in all 45 tissues); use no-primary + isotype controls |
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 A13963-1) | |
| Caveat | Low staining–RNA concordance; tissue profile uncertain (HPA tissue IHC) | |
| Regulation | Lymphoid tissue–enhanced RNA (HPA tissue RNA) | |
| Isoform / epitope | 3 isoforms; cytoplasmic epitope coverage unknown (UniProt) |
Section 1
Recommended BCAS4 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is paired with one published BCAS4 chromogenic IHC protocol (PMC13520631).
| Sample | Tissue sections; selected-image fixative not specified (standard IHC workflow) |
| Fixation | Image fixative and duration unreported (datasheet A13963-1); 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-BCAS4, 5 μg/mL (datasheet A13963-1) |
| 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 | BCAS4-positive staining in glandular cells of appendix (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 HIER at 95–98 °C for 20 min (page retrieval); the article does not specify retrieval conditions (PMC13520631).
Section 2
What Is the Expected BCAS4 Staining Pattern?
In paraffin-section IHC, expect BCAS4 staining mainly in the cytoplasm (UniProt Q8TDM0: cytoplasm; no transmembrane segment). HPA reports high staining in selected glandular, squamous, respiratory, follicle and cardiac cells (HPA: tissue IHC). Treat the pattern as provisional: HPA rates tissue staining Uncertain because antibody staining and RNA expression show low consistency (HPA: tissue IHC reliability).
| Cytoplasmic chromogen in appendix glandular cells or esophageal squamous epithelial cells. | This fits the reported compartment and cell types (UniProt Q8TDM0: cytoplasm; HPA: High in both). Compare stained cells with tissue morphology and controls before scoring; the HPA tissue IHC result is Uncertain (HPA: reliability). |
| Predominantly nuclear or sharply membranous staining, with little cytoplasmic signal. | This differs from the reported cytoplasmic location and lacks support from the topology record (UniProt Q8TDM0: cytoplasm; no transmembrane segment). Consider nonspecific staining or a detection artefact; review controls before calling it BCAS4. |
| Strong staining in fibroblasts, lymph-node germinal center cells or bone-marrow hematopoietic cells. | HPA reports Low staining in these cells (HPA: tissue IHC). Unexpected strong signal raises concern for cross-reactivity or endogenous detection activity; compare it with the negative control and local background. |
| Uniform chromogen across cells and extracellular spaces, obscuring cell boundaries. | A diffuse deposit is difficult to assign to the reported cytoplasmic pattern (UniProt Q8TDM0: cytoplasm). In standard IHC practice, inspect reagent-only controls and washing, blocking and detection conditions before interpreting intensity. |
| No detectable signal in appendix glandular cells or another HPA high-staining site. | That result conflicts with the reported tissue pattern (HPA: High in appendix glandular cells and other listed cells). First check whether the assay and control slide developed; because HPA rates tissue IHC Uncertain, absence alone does not establish biological loss (HPA: reliability). |
💡Expected BCAS4 appearanceCall a positive result when discrete cytoplasmic chromogen is visible in a reported high-staining cell type, such as appendix glandular cells (UniProt Q8TDM0: cytoplasm; HPA: High); isolated nuclear, membranous or diffuse background signal warrants control review (HPA: tissue IHC Uncertain).
| Compartment and topology | BCAS4 is annotated as cytoplasmic, with no transmembrane segment (UniProt Q8TDM0). Use cell boundaries and counterstain to judge whether chromogen lies in cytoplasm; topology alone does not validate antibody specificity. |
| Tissue and cell selection | HPA reports High staining in several specified cell types and Low staining in fibroblasts, germinal center cells and hematopoietic cells (HPA: tissue IHC). Select reference areas by cell type, not tissue name alone. |
| Isoforms and epitope coverage | UniProt lists 3 isoforms (UniProt Q8TDM0: isoforms 1–3). The supplied record does not map the antibody epitope, so staining cannot be assigned to an isoform or assumed to detect every isoform. |
| Evidence strength | HPA rates tissue IHC Uncertain and HPA038812 IHC Uncertain (HPA: tissue reliability; antibody validation). Report the observed pattern and controls; avoid treating intensity as a validated measure of BCAS4 abundance. |
| IF/ICC cross-check — Q&A | Q: What localisation is reported in IF/ICC? A: Cytosol and intermediate filaments, both approved (HPA: subcellular ICC-IF). This supports a localisation comparison, but does not resolve the uncertain tissue IHC assessment (HPA: tissue reliability). |
| Situation | Likely cause | Next action |
|---|---|---|
| No chromogen in a reported high-staining cell population. | The run may have failed, or signal may be below the chosen assay's detection threshold; biological absence is also possible. HPA tissue IHC remains Uncertain (HPA: reliability). | Check a run control, section integrity and the reagent sequence; then review retrieval, antibody dilution and development as general IHC variables. Do not infer BCAS4-specific fixation sensitivity from this record. |
| Nuclear staining dominates the section. | Nuclear predominance conflicts with the cytoplasmic annotation (UniProt Q8TDM0: cytoplasm). Background or antibody cross-reactivity is possible, but appearance alone cannot identify the cause. | Compare reagent-only and tissue controls, inspect cytoplasmic signal in the same cells, and record nuclear staining separately from any BCAS4-compatible pattern (UniProt Q8TDM0: cytoplasm). |
| Strong signal appears in fibroblasts or germinal center cells. | These are Low-staining populations in the HPA tissue record (HPA: fibroblasts; germinal center cells). Cross-reactivity or endogenous detection activity should be considered. | Confirm cell identity by morphology, compare an appropriate detection control, and reassess blocking or detection conditions as general IHC practice before scoring those cells positive. |
| Chromogen is widespread and obscures cellular detail. | Diffuse background prevents a reliable cytoplasmic call (UniProt Q8TDM0: cytoplasm). Excess detection signal or inadequate washing is a general IHC possibility, not a BCAS4-specific finding. | Inspect the reagent-only control, adjust general washing or development conditions if indicated, and score only signal that can be assigned to intact cells. |
| Adjacent high-staining candidate tissues differ markedly in intensity. | HPA lists several High-staining cell types but rates the aggregate tissue IHC evidence Uncertain (HPA: tissue IHC). Differences cannot be attributed to BCAS4 abundance from staining alone. | Compare equivalent cell populations and matched run controls; document distribution and intensity separately, and avoid ranking tissues by chromogen strength without independent validation. |
| IHC looks membranous while ICC-IF looks filamentous or cytosolic. | ICC-IF reports approved cytosol and intermediate-filament localisation, whereas UniProt annotates cytoplasm and no transmembrane segment (HPA: subcellular ICC-IF; UniProt Q8TDM0: topology). | Review IHC morphology and detection controls, then describe the discordance. The ICC-IF result does not upgrade tissue IHC reliability, which HPA calls Uncertain (HPA: tissue reliability). |
Experiment evidence
IHC & IF images showing BCAS4 staining
A13963-1 · IHCImmunohistochemistry of BCAS4 in human breast carcinoma with BCAS4 antibody at 5 μg/mL.
A13963-1 · IFImmunofluorescence of BCAS4 in human breast carcinoma tissue with BCAS4 antibody at 20 μg/mL.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000124243-BCAS4/tissue/cerebral+cortex#imid_10873571
HPA · StomachHuman Protein Atlas IHC, Stomach. Source: https://www.proteinatlas.org/ENSG00000124243-BCAS4/tissue/stomach+2#imid_10873431
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000124243-BCAS4/tissue/colon#imid_10873481
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000124243-BCAS4/tissue/liver#imid_10873498
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA038812; green = BCAS4, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA038812; green = BCAS4, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for BCAS4 IHC & IF
🧪Run appendix first and score its glandular cells for staining (HPA: High in appendix glandular cells). HPA detects BCAS4 in all 45 scored tissues, so use no-primary and isotype controls in place of a negative tissue; on the positive slide, any cells treated as internal negatives should show only background staining, and their BCAS4-negative status needs independent validation (HPA: no negative tissue listed).
Positive control tissue: Appendix (Glandular cells, HPA High)
Negative control tissue: None in HPA: BCAS4 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead.
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show BCAS4 in A-431, MCF-7, U2OS, with annotated localisation: Intermediate filaments (approved), Cytosol (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control, an isotype control matched to the primary antibody’s host species and clonality, and a matched BCAS4 knockout sample or immunizing-peptide block when available (standard IHC practice). For chromogenic appendix sections, quench endogenous peroxidase and inspect glandular lumina and nearby inflammatory cells for background before scoring (standard IHC practice).
⚠️Feasibility: A target-specific fixation window and retrieval dependency are unreported in the supplied evidence; optimize antigen retrieval empirically for paraffin IHC (standard IHC practice). The catalog antibody caption shows breast carcinoma IHC at 5 μg/mL, but its fixative is unreported (catalog antibody: A13963-1 tissue-IHC caption). HPA provides ICC-IF images and reports cytosol and intermediate filament localization, but the supplied evidence does not establish whether frozen sections or IF are easier than paraffin IHC (HPA: subcellular localization and ICC-IF images).
HPA tissue IHC evidence for BCAS4
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Uncertain — Low 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 |
|---|---|---|---|---|
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Cervix | Glandular cells | High | Protein (IHC) | HPA → |
| Epididymis | Glandular cells | High | Protein (IHC) | HPA → |
| Esophagus | Squamous epithelial cells | High | Protein (IHC) | HPA → |
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: BCAS4 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced BCAS4 IHC Tips
Troubleshoot BCAS4 staining in paraffin sections by checking cytoplasmic localisation, tissue morphology, controls, and the limits of the available antibody evidence.
What retrieval should I try first when BCAS4 staining is weak?
Use citrate buffer at pH 6.0 for heat-induced retrieval at 95–98 °C for 20 min (page retrieval rule). Let sections cool in the buffer, then compare staining with a section processed identically except for retrieval (standard IHC practice). The selected antibody image shows BCAS4 staining in human breast carcinoma at 5 µg/mL, but its caption does not report retrieval conditions (A13963-1 caption). If staining remains weak, test an alternative retrieval buffer on matched sections while keeping detection and exposure to chromogen constant; inspect both signal and tissue integrity before adopting the change (standard IHC practice).
Could fixation explain inconsistent BCAS4 staining between paraffin blocks?
BCAS4-specific sensitivity to fixation is unknown: the selected tissue-IHC caption does not state a fixative, and the supplied record provides no fixation experiment (A13963-1 caption; UniProt Q8TDM0). Record each block’s fixative and fixation interval, then compare sections with the same pH 6.0, 20 min retrieval and staining run (page retrieval rule; standard IHC practice). For prospective specimens, a documented 10% neutral buffered formalin workflow provides a consistent starting condition, without implying that this duration or reagent is optimal for BCAS4 (standard IHC practice). Interpret differences alongside morphology and control staining before assigning them to fixation (standard IHC practice).
Where should convincing BCAS4 staining appear in tissue sections?
Look chiefly for cytoplasmic staining: BCAS4 is annotated as cytoplasmic, and the tissue-IHC profile describes general cytoplasmic expression (UniProt Q8TDM0; HPA tissue IHC). Approved cell-image locations include cytosol and intermediate filaments, so diffuse or filament-like cytoplasmic patterns can be examined against cell morphology (HPA subcellular). Score cells individually with a nuclear counterstain rather than treating pigment over nuclei or tissue edges as a localisation result (standard IHC practice). A predominantly nuclear or sharply membrane-confined DAB pattern needs additional controls before attribution to BCAS4, which has no annotated transmembrane segment (UniProt Q8TDM0 topology).
Can a negative section rule out every BCAS4 isoform?
No: BCAS4 has 3 annotated isoforms, while the supplied antibody caption does not identify its immunogen or epitope (UniProt Q8TDM0; A13963-1 caption). Check the catalog antibody’s epitope information, if available, against the isoform sequences before making an isoform-wide claim (standard IHC practice). The record annotates no transmembrane segment, glycosylation sites, modified residues, or domains; those annotations do not establish antibody recognition after processing (UniProt Q8TDM0). Compare a positive-control section and the test section in the same run, and describe a negative result as absence of detectable staining under the tested retrieval and detection conditions (standard IHC practice).
How can IF help check an ambiguous chromogenic BCAS4 pattern?
Use IF on a matched specimen as an orthogonal localisation check while treating the selected 5 µg/mL breast-carcinoma image as IHC evidence only (A13963-1 caption). Multiplex BCAS4 with a cell-type marker, such as cytokeratin for glandular epithelial cells, and inspect overlap within individual cells; HPA reports high staining in glandular cells in several tissues (HPA tissue IHC; standard IF practice). Choose a far-red fluorophore and include an unstained control to assess tissue autofluorescence (standard IF practice). Because BCAS4 is cytoplasmic and lacks a transmembrane segment, test gentle permeabilisation, such as 0.1% Triton X-100 for 5 min, with an unpermeabilised control (UniProt Q8TDM0; standard IF practice).
How should I reduce diffuse brown background without losing BCAS4 signal?
Run a primary-antibody omission control to locate staining from detection reagents or endogenous tissue activity (standard IHC practice). In a peroxidase and DAB workflow, block endogenous peroxidase with, for example, 3% hydrogen peroxide for 10 min, then wash thoroughly; this is a general workflow step, not BCAS4-specific evidence (standard IHC practice). Add an appropriate protein block for about 30 min and compare DAB development times across matched sections (standard IHC practice). The image caption reports 5 µg/mL in human breast carcinoma, but does not establish that concentration as optimal for other specimens (A13963-1 caption).
How should I score BCAS4 across heterogeneous paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and analysis region before scoring, and exclude folds, necrosis, and section edges (standard IHC practice). For viable cells, record the percentage at each cytoplasmic intensity grade 0–3 and calculate an H-score from 0–300; alternatively, report percentage positive with a fixed threshold (standard IHC practice; UniProt Q8TDM0 localisation). Normalise cell counts to the number of evaluable cells, or positive-cell density to viable tissue area in mm², using the same rule for every section (standard IHC practice). Keep retrieval, antibody concentration, chromogen development, and image settings consistent, and report compartments separately when cell composition varies (standard IHC practice).
When is BCAS4 positivity more likely to be artefact?
Give greatest weight to reproducible cytoplasmic staining in intact cells, consistent with the BCAS4 annotation and tissue-IHC profile (UniProt Q8TDM0; HPA tissue IHC). Investigate signal confined to nuclei, section edges, necrotic areas, or the primary-antibody omission control before calling cells positive (standard IHC practice). A sharply membrane-only pattern also merits scrutiny because BCAS4 has no annotated transmembrane segment (UniProt Q8TDM0 topology). Compare the apparent positive cell type with morphology and a matched control section; HPA reports low consistency between antibody staining and RNA expression, so its tissue pattern alone cannot confirm specificity (HPA tissue IHC).
Boster reagents
Best BCAS4 / Breast carcinoma-amplified sequence 4 IHC Antibodies
A13963-1 has IHC and IF images from human breast carcinoma tissue (IHC and IF image captions); the catalog lists Human and Mouse reactivity (catalog: reactivity).
Real IHC data
Anti-BCAS4 Antibody
Cat # A13963-1
A13963-1 is listed for IHC-P and IF, with Human and Mouse reactivity (catalog: applications and reactivity). Its IHC image shows human breast carcinoma at 5 μg/mL, and its IF image shows human breast carcinoma tissue at 20 μg/mL (IHC and IF image captions).
Which to pick: Choose A13963-1 for paraffin-section IHC (catalog: IHC-P application); its human breast carcinoma IHC caption does not report the fixative (IHC image caption). For tissue IF, choose A13963-1 based on its human breast carcinoma IF image; ICC validation is unreported (IF image caption; catalog: applications). For work involving Mouse samples, A13963-1 lists Mouse reactivity, but the supplied IHC and IF images show Human samples only; its clone is unreported (catalog: reactivity and clone; IHC and IF image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q8TDM0 (BCAS4_HUMAN, Breast carcinoma-amplified sequence 4).
- Human Protein Atlas. BCAS4 tissue IHC expression (reliability: Uncertain).
- Human Protein Atlas. BCAS4 subcellular location (ICC-IF): Localized to the cytosol and intermediate filaments..
- Human Protein Atlas. BCAS4 antibody validation summary (1 antibodies).
- RNF180 loss stabilizes BCAS4 to enhance PD-L1 expression and immune evasion in hepatocellular carcinoma. iScience 2026 — PMC13520631.
- PubMed PMID:12378525 — UniProt-cited evidence.
- PubMed PMID:11780052 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for BCAS4 (Breast carcinoma-amplified sequence 4). Expected localisation General cytoplasmic tissue staining (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A13963-1 at 5 μg/mL (the product's own IHC validation image is shown). Controls (HPA): positive Appendix (Glandular cells); negative None in HPA (detected in all 45 tissues); use no-primary + isotype controls. 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 A13963-1); 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-BCAS4, 5 μg/mL (datasheet A13963-1); 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 A13963-1); Caveat: Low staining–RNA concordance; tissue profile uncertain (HPA tissue IHC); Isoform / epitope: 3 isoforms; cytoplasmic epitope coverage unknown (UniProt); Regulation: Lymphoid tissue–enhanced RNA (HPA tissue RNA). Evidence: UniProt Q8TDM0, HPA tissue IHC · Uncertain, HPA ICC-IF · cytosol and intermediate filaments., Datasheet A13963-1, PMC13520631. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/BCAS4/immunohistochemistry-ihc-guide.