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COL14A1 / Collagen alpha-1(XIV) chain · IHC design guide
Design Immunohistochemistry for COL14A1
Plan paraffin-section COL14A1 IHC using the reported cytoplasmic tissue pattern (HPA tissue IHC). Compare positive and negative cell populations while accounting for the protein’s extracellular matrix location (UniProt) and the tissue staining report’s uncertain reliability (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
COL14A1 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 tissue staining (HPA tissue IHC); extracellular matrix protein (UniProt) | |
| Staining pattern | Cytoplasmic staining in most tissues (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 | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent (standard IHC practice; not target-specific); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A06685) | |
| Caveat | Secreted protein may show weak RNA–protein correlation (HPA tissue IHC) | |
| Regulation | Blood-vessel-enhanced RNA; staining impact unknown (HPA tissue RNA + IHC) | |
| Isoform / epitope | 3 isoforms; mature chain begins at aa 29 (UniProt) |
Section 1
Recommended COL14A1 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by reported COL14A1 staining methods for mouse maxillary sections (PMC10401928) and a bladder cancer tissue microarray (PMC12372714).
| Sample | Paraffin-embedded human skin tissue; fixative not specified (datasheet A06685) |
| Fixation | Image fixative and duration unreported (datasheet A06685); 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-COL14A1, 1:50 recommended; image 1:200 (datasheet A06685) |
| 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 | COL14A1-positive staining in glandular cells of appendix (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page retrieval); neither article specifies retrieval conditions (PMC10401928; PMC12372714).
Section 2
What Is the Expected COL14A1 Staining Pattern?
COL14A1 is secreted into the extracellular matrix and has no transmembrane segment (UniProt Q05707: subcellular location and topology). In paraffin tissue sections, HPA reports predominantly cytoplasmic staining, including high staining in selected glandular and squamous epithelial cells (HPA: tissue IHC). Treat that observed pattern as provisional: HPA rates its tissue IHC reliability Uncertain because secretion complicates the relationship between cellular protein expression and tissue location (HPA: reliability).
| Strong cytoplasmic staining in appendix, colon, or rectum glandular cells, or esophageal squamous epithelial cells (HPA: High in those cells). | This matches the reported tissue IHC distribution (HPA: tissue IHC). Compare stained cells with adjacent unstained structures and controls; the HPA reliability rating remains Uncertain, so intensity alone does not confirm specificity (HPA: reliability; general IHC practice). |
| Predominantly nuclear staining, with little of the reported cytoplasmic epithelial pattern (HPA: tissue IHC). | Nuclear dominance conflicts with the secreted, extracellular annotation and the reported cytoplasmic IHC profile (UniProt Q05707: subcellular location; HPA: tissue IHC). Check controls and staining conditions before interpreting it as COL14A1; an unexpected compartment can reflect nonspecific signal (general IHC practice). |
| Strong signal mainly in a cell population reported as undetected, such as bone marrow hematopoietic cells (HPA: Not detected). | That distribution conflicts with the supplied HPA observation, though an undetected result is not proof of biological absence (HPA: tissue IHC). Cross-reactivity or endogenous detection activity is possible; assess antibody-specific and detection controls before assigning the signal (general IHC practice). |
| Uniform haze across tissue, lumina, and blank areas obscures cell boundaries (general IHC practice). | This pattern cannot be scored against HPA's cell-specific observations (HPA: tissue IHC). Background may arise from insufficient blocking, overly concentrated reagents, or incomplete washing; compare a no-primary control and inspect the detection chemistry (general IHC practice). |
| No signal in appendix, colon, rectum, or esophagus despite intact morphology (HPA: High in the listed epithelial cells). | A negative run in a reported high-staining tissue calls for a technical review before a biological conclusion (HPA: tissue IHC; general IHC practice). Check section quality, antibody and detection controls, and the validated IHC-P workflow; HPA's Uncertain rating limits the tissue's value as a definitive positive control (HPA: reliability). |
💡Expected COL14A1 appearanceA plausible positive IHC result is predominantly cytoplasmic staining, strongest in the specified glandular or squamous epithelial cells (HPA: tissue IHC); dominant nuclear or indiscriminate tissue-wide color is suspect (UniProt Q05707: secreted extracellular protein; general IHC practice).
| Secreted protein versus tissue staining | UniProt places COL14A1 in extracellular space and matrix, whereas HPA describes cytoplasmic staining in most tissues (UniProt Q05707: subcellular location; HPA: tissue IHC). Do not require a continuous matrix outline or infer a producing cell solely from extracellular signal; secretion makes tissue location harder to interpret (HPA: reliability). |
| Tissue and cell selection | Appendix, colon, rectum, and esophagus provide reported high-staining epithelial populations (HPA: High). Adipocytes in adipose tissue and hematopoietic cells in bone marrow are reported undetected (HPA: Not detected). Score the named cell population, since a tissue-level label does not describe every structure on the slide (general IHC practice). |
| Protein forms and epitope uncertainty | COL14A1 has three isoforms, an N-terminal signal peptide at residues 1–28, and multiple glycosylation sites (UniProt Q05707: isoforms, processing, glycosylation). The supplied record does not identify this antibody's epitope, so these features do not predict which isoforms or processed forms its IHC stain detects. |
| Antigen retrieval and detection | No COL14A1-specific retrieval condition or fixation sensitivity is supplied. Follow the IHC-validated antibody's documented IHC-P procedure where available; vary retrieval or detection conditions only as a controlled optimization, comparing positive tissue and no-primary controls (general IHC practice). Do not infer a retrieval requirement from UniProt modifications or HPA intensity. |
| IF/ICC Q&A: should vesicles define the IHC-P result? | No: HPA reports approved vesicular ICC-IF localization, while its tissue IHC profile is cytoplasmic and rated Uncertain (HPA: subcellular ICC-IF; HPA: tissue IHC reliability). The two preparations answer different localization questions; vesicles in cultured cells do not establish a required paraffin-section staining pattern. |
| Situation | Likely cause | Next action |
|---|---|---|
| Reported high-staining epithelium is blank (HPA: tissue IHC). | The stain may have failed, or the chosen section may lack evaluable target cells (general IHC practice); HPA's Uncertain rating also limits expectations for an individual specimen (HPA: reliability). | Confirm the named cells on the section, review antibody and detection controls, and repeat using the documented IHC-P workflow (general IHC practice). Compare another reported high-staining tissue if available (HPA: tissue IHC). |
| Diffuse brown color makes cytoplasmic scoring impossible (general IHC practice). | Excess detection signal, incomplete washing, or endogenous activity can obscure the reported cell-specific pattern (general IHC practice; HPA: tissue IHC). | Inspect a no-primary control, increase washes, and adjust blocking or reagent concentration within the validated workflow (general IHC practice). Score COL14A1 only when cell boundaries and control behavior are interpretable. |
| Nuclei dominate the stain (general IHC practice). | The pattern conflicts with HPA's cytoplasmic tissue profile and UniProt's secreted extracellular location (HPA: tissue IHC; UniProt Q05707: subcellular location). | Review counterstain and chromogen separately, then compare no-primary and positive-tissue controls (general IHC practice). If nuclear color persists as antibody-dependent signal, treat specificity as unresolved. |
| Strong signal appears in HPA-undetected hematopoietic cells or adipocytes (HPA: tissue IHC). | Possible cross-reactivity or endogenous detection activity; HPA's undetected classification is an observation, not a universal absence claim (general IHC practice; HPA: tissue IHC). | Compare the same run with reported high-staining epithelial cells and no-primary controls (HPA: tissue IHC; general IHC practice). Withhold a COL14A1 assignment until the unexpected cell-type signal is independently resolved. |
| Extracellular deposits and cytoplasmic cells stain together (general IHC practice). | An extracellular component is biologically plausible for secreted COL14A1, but the supplied HPA tissue profile reports cytoplasmic expression and has Uncertain reliability (UniProt Q05707: subcellular location; HPA: tissue IHC reliability). | Record extracellular and cellular staining separately, examine morphology, and compare controls (general IHC practice). Do not score every matrix deposit as confirmed COL14A1 or use it alone to identify its cell of origin. |
| Staining varies after changing retrieval conditions (general IHC practice). | Protocol changes can alter IHC signal generally; no target-specific fixation or retrieval effect is established in the supplied sources. | Return to the documented IHC-P conditions and compare one change at a time on adjacent sections with the same controls (general IHC practice). Report the condition used and avoid claiming that COL14A1 is fixation sensitive. |
Experiment evidence
IHC & IF images showing COL14A1 staining
A06685 · IHCImmunohistochemical analysis of paraffin-embedded human-skin, antibody was diluted at 1:200.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000187955-COL14A1/tissue/cerebral+cortex#imid_6124680
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000187955-COL14A1/tissue/colon#imid_6124606
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000187955-COL14A1/tissue/liver#imid_6124607
HPA · GallbladderHuman Protein Atlas IHC, Gallbladder. Source: https://www.proteinatlas.org/ENSG00000187955-COL14A1/tissue/gallbladder#imid_6124599
HPA · ICC-IF Rh30Human Protein Atlas immunofluorescence, Rh30, antibody HPA023781; green = COL14A1, blue = nucleus, red = microtubules.
HPA · ICC-IF Rh30Human Protein Atlas immunofluorescence, Rh30, antibody HPA023781; green = COL14A1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for COL14A1 IHC & IF
🧪Run colon first and look for staining associated with glandular cells (HPA: High in colon glandular cells). Run adrenal gland glandular cells as the negative tissue (HPA: Not detected in adrenal gland glandular cells); on the colon slide, glandular nuclei should remain unstained while extracellular matrix staining may be present (UniProt Q05707: secreted, extracellular matrix).
Positive control tissue: Appendix (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show COL14A1 in Rh30, U-251MG, U2OS, with annotated localisation: Vesicles (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control; an isotype control matched to the primary antibody’s host species, immunoglobulin class and concentration; and COL14A1 knockout tissue or a validated antigen-peptide competition control (standard IHC practice). For chromogenic detection, quench endogenous peroxidase and block endogenous biotin if using a biotin-based detection system (standard IHC practice).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the 1:200 paraffin-section caption for A06685 does not state a fixative (A06685 tissue-IHC caption). Antigen-retrieval dependency is unreported, so retrieval conditions need empirical optimization for paraffin IHC (supplied evidence; standard IHC practice). The supplied evidence does not establish whether frozen sections or IF are easier; if testing IF in colon, assess mucosal autofluorescence with a no-primary control and interpret vesicular signal separately from extracellular matrix staining (standard IF practice; HPA subcellular: vesicles; UniProt Q05707: extracellular matrix).
HPA tissue IHC evidence for COL14A1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Uncertain — At least one protein variant secreted, tissue location of RNA and protein might differ and correlation is complex. Pending external verification.). 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 → |
| Colon | Glandular cells | High | Protein (IHC) | HPA → |
| Esophagus | Squamous epithelial cells | High | Protein (IHC) | HPA → |
| Rectum | Glandular cells | High | Protein (IHC) | HPA → |
| Stomach | Glandular cells | Medium | 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 → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced COL14A1 IHC Tips
Troubleshoot COL14A1 staining in paraffin sections by checking retrieval, compartment, matrix morphology, and controls before comparing chromogenic signal across samples.
How should I retrieve COL14A1 when paraffin sections stain weakly?
Start with citrate buffer at pH 6.0 for heat-induced retrieval at 95–98 °C for 20 min (page retrieval rule). Let sections cool in buffer, then compare matched sections with and without retrieval to detect loss of morphology or excess background (standard IHC practice). If staining remains weak, test a shorter heating time before increasing retrieval intensity, keeping antibody concentration and detection conditions constant (standard IHC practice). Record matrix staining separately from cellular staining because COL14A1 is secreted into extracellular matrix, while HPA reports cytoplasmic tissue staining (UniProt Q05707 subcellular; HPA tissue IHC profile).
How can I assess whether fixation is limiting COL14A1 IHC?
Target-specific fixation sensitivity is unknown; the catalog image identifies paraffin-embedded human skin and a 1:200 dilution but does not state its fixative (A06685 tissue-IHC caption). Record the fixative, fixation duration, processing schedule, and section thickness for each specimen before comparing staining intensity (standard IHC practice). Compare sections processed under documented conditions using identical citrate pH 6.0 retrieval, antibody incubation, and chromogen development (page retrieval rule; standard IHC practice). If signal differs, check tissue preservation and internal staining patterns before attributing the difference to fixation; no supplied evidence establishes a COL14A1-specific fixation effect (A06685 tissue-IHC caption).
Should COL14A1 staining appear in cells or extracellular matrix?
Assess extracellular matrix staining because COL14A1 is secreted, has no transmembrane segment, and is annotated in extracellular space and matrix (UniProt Q05707 subcellular; UniProt Q05707 topology). Its proposed association with interstitial collagen fibrils supports examining fibrillar regions, although that function is annotated by similarity (UniProt Q05707 function). Also document cellular signal rather than automatically rejecting it: HPA describes cytoplasmic staining in most tissues, with uncertain tissue-IHC reliability (HPA tissue IHC profile; HPA tissue IHC reliability). Compare the pattern with section morphology and a no-primary control, scoring cellular and matrix compartments separately (standard IHC practice).
Could epitope location explain inconsistent COL14A1 staining?
Check the catalog antibody’s documented immunogen or epitope before interpreting differences between samples; its binding region is not supplied here (A06685 tissue-IHC caption). COL14A1 has 3 annotated isoforms, so an epitope outside a shared sequence could change which forms are detected (UniProt Q05707 isoforms; standard antibody interpretation). Its domains span an amino-terminal globular region and carboxyl-terminal collagen-like segments, with annotated glycosylation and hydroxylation sites that make epitope context relevant (UniProt Q05707 domains; UniProt Q05707 PTMs). Compare staining after the same retrieval and processing, then use an independently mapped antibody or orthogonal evidence if isoform specificity matters (standard IHC practice).
How should I investigate COL14A1 localisation by multiplex IF?
Use this as a separate IF assay: the catalog caption establishes paraffin-section chromogenic IHC at 1:200, without stating an IF dilution or fixation method (A06685 tissue-IHC caption). Pair COL14A1 with a validated marker for the cell population being evaluated, and inspect matrix signal independently of marker overlap (standard IF practice; UniProt Q05707 subcellular). Choose fluorophores and exposure settings using unstained tissue controls, particularly where tissue autofluorescence could obscure extracellular signal (standard IF practice). For a surface or extracellular epitope, compare staining without permeabilisation; permeabilise only when testing an intracellular epitope or vesicular signal (UniProt Q05707 topology; HPA subcellular; standard IF practice).
What controls help distinguish COL14A1 signal from chromogenic background?
Run a no-primary control with the same secondary reagent, peroxidase block, and DAB development to reveal detection-system background (standard chromogenic IHC practice). Inspect folds, cut edges, damaged areas, and precipitate before assigning brown deposits to COL14A1 (standard IHC practice). If background is diffuse, optimise blocking and washes, then compare antibody dilutions around the caption’s 1:200 condition while holding development time constant (A06685 tissue-IHC caption; standard IHC practice). A plausible extracellular pattern supports review, but it cannot establish specificity alone because HPA rates its tissue-IHC profile uncertain (UniProt Q05707 subcellular; HPA tissue IHC reliability).
How should I quantify COL14A1 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the compartment before scoring: measure extracellular staining as positive area or optical density per mm² of viable tissue, and score cellular staining separately (UniProt Q05707 subcellular; standard IHC quantification). For cellular signal, report the percentage of positive cells or an H-score using prespecified intensity categories (standard IHC quantification). Keep section thickness, retrieval, the caption’s 1:200 antibody dilution, and chromogen development consistent across compared sections (A06685 tissue-IHC caption; page retrieval rule; standard IHC practice). Normalise matrix measurements to the analysed viable tissue area and cellular measurements to the counted eligible cells, excluding folds and necrosis by fixed criteria (standard IHC quantification).
When is a COL14A1-positive IHC pattern convincing?
A convincing result follows preserved tissue architecture, survives comparison with a no-primary control, and is reproducible across similarly processed sections (standard IHC practice). Assess extracellular staining against COL14A1’s secreted matrix annotation, while recording cellular staining separately because HPA reports cytoplasmic expression with uncertain reliability (UniProt Q05707 subcellular; HPA tissue IHC profile; HPA tissue IHC reliability). Treat isolated nuclear deposits, section-edge staining, necrotic regions, and residual endogenous peroxidase signal as potential artefacts requiring control review (UniProt Q05707 topology; standard chromogenic IHC practice). Do not equate an HPA-positive cell type or RNA enrichment with confirmed protein production in that cell: secreted protein can reside away from its source (HPA tissue IHC reliability; HPA tissue RNA specificity).
Boster reagents
Best COL14A1 / Collagen alpha-1(XIV) chain IHC Antibodies
A06685 has an IHC image of paraffin-embedded human skin (IHC image caption) and lists IF use and Human, Mouse, and Rat reactivity (catalog applications/reactivity); no IF image is supplied (catalog IF images).
Real IHC data
Anti-Collagen alpha-1(XIV) chain COL14A1 Antibody
Cat # A06685
A06685 will render with its paraffin-embedded human-skin IHC figure at 1:200 (IHC image caption). It lists IHC and IF applications and Human, Mouse, and Rat reactivity (catalog applications/reactivity); no IF figure is supplied (catalog IF images).
Which to pick: For tissue IHC, choose A06685: its own figure shows paraffin-embedded human skin (IHC image caption); the fixative is unreported (IHC image caption). For IF, A06685 lists an IF dilution of 1:50 (catalog IF dilution), but IF imaging and ICC validation are unreported (catalog IF images/applications). A06685 also lists Mouse and Rat reactivity (catalog reactivity), although its supplied IHC figure shows only human skin (IHC image caption); clonality is unreported (catalog clone field).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q05707 (COEA1_HUMAN, Collagen alpha-1(XIV) chain).
- Human Protein Atlas. COL14A1 tissue IHC expression (reliability: Uncertain).
- Human Protein Atlas. COL14A1 subcellular location (ICC-IF): Localized to vesicles..
- Human Protein Atlas. COL14A1 antibody validation summary (1 antibodies).
- Novel Biomarkers of Health and Degeneration in Human Intervertebral Discs: In-depth Proteomic Analysis of Collagen Framework of Fetal, Healthy, Scoliotic, Degenerate, and Herniated Discs. Asian spine journal 2023 — PMC9977988.
- Mouse gingival single-cell transcriptomic atlas: An activated fibroblast subpopulation guides oral barrier immunity in periodontitis. bioRxiv : the preprint server for biology 2023 — PMC10401928.
- Innovative logic "AND" gate gene circuits for bladder cancer treatment: preclinical study. International journal of surgery (London, England) 2025 — PMC12372714.
- Part 1: profiling extra cellular matrix core proteome of human fetal nucleus pulposus in search for regenerative targets. Scientific reports 2020 — PMC7519061.
- PubMed PMID:16421571 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
- PubMed PMID:9427527 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for COL14A1 (Collagen alpha-1(XIV) chain). Expected localisation Cytoplasmic tissue staining (HPA tissue IHC); extracellular matrix protein (UniProt). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A06685 at 1:50 recommended; image 1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Appendix (Glandular cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human skin tissue; fixative not specified (datasheet A06685); 2. Fixation: Image fixative and duration unreported (datasheet A06685); 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-COL14A1, 1:50 recommended; image 1:200 (datasheet A06685); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent (standard IHC practice; not target-specific); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A06685); Caveat: Secreted protein may show weak RNA–protein correlation (HPA tissue IHC); Isoform / epitope: 3 isoforms; mature chain begins at aa 29 (UniProt); Regulation: Blood-vessel-enhanced RNA; staining impact unknown (HPA tissue RNA + IHC). Evidence: UniProt Q05707, HPA tissue IHC · Uncertain, HPA ICC-IF · vesicles., Datasheet A06685, PMC10401928, PMC12372714. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/COL14A1/immunohistochemistry-ihc-guide.