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COL14A1 / Collagen alpha-1(XIV) chain · IHC design guide
Design Immunohistochemistry for COL14A1
Plan COL14A1 paraffin-section IHC using reported cytoplasmic staining, including high signal in appendix glandular cells (HPA tissue IHC). Start the catalog antibody at 1:50 and optimize with controls (datasheet: 1:50); interpret staining cautiously because the tissue IHC evidence is uncertain (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 staining in most tissues (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic signal; high in glandular and squamous 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 | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across paraffin sections (standard IHC practice; not target-specific) | |
| Caveat | Tissue staining is uncertain for this secreted protein (HPA tissue IHC) | |
| Regulation | Tissue-enhanced RNA in blood vessels (HPA tissue RNA) | |
| Isoform / epitope | 3 isoforms; signal peptide removed; epitope unknown (UniProt) |
Section 1
Recommended COL14A1 IHC & IF Protocols
The catalog antibody's IHC-P protocol is accompanied by 3 published COL14A1 paraffin IHC protocols (PMC9977988; PMC10401928; 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 HIER at 95–98 °C for 20 min (page retrieval); consider alkaline retrieval if staining is weak (PMC9977988).
Section 2
What Is the Expected COL14A1 Staining Pattern?
COL14A1 is a secreted, extracellular matrix protein with no transmembrane segment (UniProt Q05707 topology). For paraffin-section IHC, assess matrix-associated staining alongside the cytoplasmic staining reported in glandular cells of appendix, colon and rectum and squamous epithelial cells of esophagus (HPA tissue IHC). Treat either pattern cautiously: HPA rates its tissue IHC reliability Uncertain, noting that secreted protein can complicate the match between RNA and protein location (HPA tissue IHC).
| Signal outlines interstitial structures, with staining near appendix or colon glands (UniProt Q05707 subcellular location; HPA tissue IHC). | An extracellular distribution is consistent with COL14A1 secretion and its proposed association with collagen fibrils (UniProt Q05707 function). Judge it alongside the documented glandular-cell signal; HPA has not verified a definitive tissue pattern (HPA tissue IHC: Uncertain). |
| The strongest signal is confined to nuclei, without a comparable extracellular or reported cytoplasmic pattern. | Nuclear-only staining conflicts with the extracellular location assigned by UniProt and the cytoplasmic tissue profile reported by HPA (UniProt Q05707 subcellular location; HPA tissue IHC). Check counterstain overlap and repeat with appropriate controls before calling it COL14A1-specific signal (standard IHC practice). |
| Strong staining appears in adipocytes or bone-marrow hematopoietic cells instead of the expected comparison cells. | Those cell populations are reported as not detected by HPA (HPA tissue IHC). Unexpected chromogen may reflect cross-reactivity or endogenous detection activity (standard IHC practice); the finding alone does not establish which cause applies. |
| Chromogen covers much of the section without a discernible cell or matrix distribution. | Diffuse staining cannot be assigned confidently to the reported HPA cell pattern or UniProt extracellular location (HPA tissue IHC; UniProt Q05707 subcellular location). Review the no-primary control and detection background before scoring intensity (standard IHC practice). |
| Appendix glandular cells show no staining in a run that includes that tissue. | Appendix glandular cells are reported High by HPA, so their absence calls for a run-level check (HPA tissue IHC). HPA also rates the IHC evidence Uncertain; a blank section alone cannot prove target absence or antibody failure (HPA tissue IHC). |
💡Expected COL14A1 appearanceA plausible positive IHC result shows localized matrix-associated signal consistent with secretion (UniProt Q05707 subcellular location), potentially alongside High glandular-cell staining in appendix, colon or rectum (HPA tissue IHC); nuclear-only or uniform diffuse chromogen is suspect (standard IHC interpretation), and HPA rates its tissue IHC reliability Uncertain (HPA tissue IHC).
| Extracellular location and processing | COL14A1 has a signal peptide at residues 1–28 and a mature chain beginning at residue 29 (UniProt Q05707 processing). Secreted protein can appear away from its producing cells; HPA explicitly warns that tissue RNA and protein locations may differ (HPA tissue IHC). |
| Choice of comparison cells | Appendix, colon and rectum glandular cells are High; esophageal squamous cells are also High (HPA tissue IHC). Adipocytes and bone-marrow hematopoietic cells are reported not detected (HPA tissue IHC). Compare the named cell populations, not entire tissues. |
| Confidence in the IHC pattern | HPA rates tissue staining Uncertain and lists antibody HPA023781 as IHC Uncertain (HPA tissue IHC; HPA antibodies). Interpret a convincing distribution as provisional and use run controls when judging specificity (standard IHC practice). |
| IF/ICC Q&A: What signal is reported? | HPA reports vesicles as the approved ICC-IF location and lists HPA023781 as ICC Approved (HPA subcellular; HPA antibodies). This describes cell imaging; it does not establish a paraffin-section IHC pattern (HPA tissue IHC: Uncertain). |
| Situation | Likely cause | Next action |
|---|---|---|
| Positive comparison tissue is blank. | The run may lack detectable signal, or the chosen field may miss the reported cells (standard IHC practice). Appendix glandular cells are reported High (HPA tissue IHC). | Inspect glandular areas and confirm the run's positive and no-primary controls; then review the validated antibody and detection workflow (standard IHC practice). Do not infer a COL14A1-specific fixation effect from the blank result. |
| Nuclei stain prominently while matrix and reported cell compartments do not. | A nuclear-only pattern conflicts with UniProt's extracellular assignment and HPA's cytoplasmic tissue profile (UniProt Q05707 subcellular location; HPA tissue IHC). | Check whether chromogen is being confused with counterstain, compare the no-primary control, and reassess the compartment in an independently stained section (standard IHC practice). |
| Adipocytes or marrow hematopoietic cells stain strongly. | HPA reports these cells as not detected (HPA tissue IHC). Cross-reactivity or endogenous detection activity can produce unexpected staining (standard IHC practice). | Compare a no-primary control and an appropriate endogenous-enzyme control; score the named cells separately from nearby stroma (standard IHC practice). |
| Brown signal is widespread with little structural contrast. | Diffuse chromogen may arise from background in blocking, washing or detection steps (standard IHC practice); it cannot resolve the uncertain HPA tissue pattern (HPA tissue IHC). | Compare the no-primary control, inspect wash and detection steps, and repeat under the validated IHC workflow before assigning a positive score (standard IHC practice). |
| Only extracellular staining appears; reported glandular-cell staining is absent. | COL14A1 is secreted, and HPA notes that RNA and protein tissue locations may differ (UniProt Q05707 subcellular location; HPA tissue IHC). The isolated matrix signal remains unverified as specific. | Document the matrix compartment and named cell populations separately; compare controls and another HPA-reported positive tissue before interpreting the result (standard IHC practice; HPA tissue IHC). |
| IHC looks cytoplasmic, but an IF/ICC image looks vesicular. | HPA reports cytoplasmic tissue staining and an approved vesicular ICC-IF location in separate assays (HPA tissue IHC; HPA subcellular). Neither observation validates the other. | Report each assay's compartment and validation status separately: tissue IHC is Uncertain, while the listed antibody's ICC result is Approved (HPA tissue IHC; HPA antibodies). |
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 appendix first and assess its glandular cells (HPA: appendix glandular cells, High); use adrenal gland as the negative tissue (HPA: adrenal gland glandular cells, Not detected). On the appendix slide, compare glandular staining with nearby cells that lack signal, while scoring extracellular staining separately because COL14A1 is an extracellular matrix protein (UniProt Q05707: subcellular location).
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 and immunoglobulin class; and a biological negative such as knockout tissue or a peptide-blocked primary, if available (standard IHC practice). For chromogenic IHC, block endogenous peroxidase and, if using avidin–biotin detection, assess endogenous biotin (standard IHC practice).
⚠️Feasibility: The selected paraffin-section skin caption reports a 1:200 dilution, but its fixative is unreported (selected A06685 tissue-IHC caption). A target-specific fixation window, retrieval dependence, and whether frozen sections or IF are easier are unreported in the supplied evidence. No appendix-specific artefact is reported; for IF/ICC interpretation, distinguish extracellular matrix localization from the vesicular signal reported in cell-line images (UniProt Q05707: subcellular location; HPA: subcellular localization, vesicles).
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
Use extracellular matrix distribution and matched controls to troubleshoot COL14A1 staining in paraffin sections; interpret cellular signals cautiously (UniProt Q05707 localisation; HPA tissue IHC reliability: Uncertain).
How should I adjust retrieval when COL14A1 staining is weak in paraffin sections?
Start with citrate buffer at pH 6.0, heated to 95–98 °C for 20 min (page retrieval setting). Compare the same tissue across a short retrieval time course while keeping section thickness, antibody concentration, and chromogen development constant (standard IHC practice). If staining remains weak, test a higher pH buffer as a fallback on adjacent sections, watching for tissue damage and increased background (standard IHC practice). Score extracellular matrix separately from cellular staining because COL14A1 is secreted into the extracellular matrix, while the available tissue IHC profile reports widespread cytoplasmic staining with uncertain reliability (UniProt Q05707 localisation; HPA tissue IHC).
Could fixation explain weak or uneven COL14A1 staining?
Target-specific fixation sensitivity is unknown: the catalog image shows paraffin-embedded human skin, but its caption does not state a fixative (A06685 tissue IHC caption). Record the actual fixative, fixation duration, and tissue thickness for each block before comparing staining intensity (standard IHC practice). On adjacent sections, keep retrieval at citrate pH 6.0, 95–98 °C for 20 min, then vary only the staining condition under investigation (page retrieval setting; standard IHC practice). If signal tracks processing batches, compare matched blocks and controls before attributing the difference to COL14A1 expression; its secreted topology does not establish fixation sensitivity (UniProt Q05707 localisation).
How should I investigate staining that appears mainly inside cells?
Map chromogenic signal against cell borders and collagen-rich extracellular areas before assigning a compartment (standard IHC practice). COL14A1 is annotated as secreted and extracellular, with no transmembrane segment, so convincing matrix staining deserves separate documentation from intracellular signal (UniProt Q05707 localisation and topology). Intracellular staining is still plausible during synthesis or trafficking: tissue IHC reports cytoplasmic expression in most tissues, and subcellular IF reports vesicles (HPA tissue IHC; HPA subcellular). Because tissue IHC reliability is Uncertain, repeat the observation with an independent epitope antibody or another specific control before interpreting cell-associated brown staining as deposited COL14A1 (HPA tissue IHC reliability; standard IHC practice).
Could epitope choice account for discordant COL14A1 staining patterns?
Check the immunogen coordinates against the 3 annotated isoforms before treating differing staining patterns as biological disagreement (UniProt Q05707 isoforms). COL14A1 contains fibronectin type-III, von Willebrand factor A, laminin G-like, and collagen-like regions, so antibodies against different regions may sample different accessible epitopes (UniProt Q05707 domains; standard IHC practice). The record also lists glycosylation and hydroxylation sites, but those annotations alone do not establish that a particular antibody loses binding in paraffin sections (UniProt Q05707 modifications). Compare antibodies on adjacent sections using the same retrieval and detection conditions, and report each reagent’s mapped epitope or its unknown status (standard IHC practice).
How can IF help resolve ambiguous chromogenic COL14A1 staining?
On a matched section, multiplex COL14A1 with an epithelial marker when investigating the glandular-cell signal reported in appendix or colon (HPA tissue IHC; standard IF practice). Choose spectrally separated fluorophores and inspect unstained tissue in each channel before assigning signal, since tissue autofluorescence can overlap the label (standard IF practice). For extracellular deposits, begin without detergent permeabilisation; for an intracellular vesicle epitope, compare a permeabilised condition because subcellular IF localises COL14A1 to vesicles (UniProt Q05707 localisation; HPA subcellular; standard IF practice). Compare matrix and cellular signal independently, and do not treat this IF check as validation of the catalog antibody for IF/ICC (standard IF practice).
What should I check when extracellular regions stain diffusely brown?
Inspect a no-primary control and the counterstained section to distinguish diffuse chromogen deposition from structured extracellular staining (standard IHC practice). Apply a peroxidase block before DAB development and compare shorter development times if both test and control sections accumulate brown signal (standard chromogenic IHC practice). Recheck blocking, washes, and the primary concentration; the selected paraffin-skin image used 1:200, which is a documented starting reference rather than proof of optimal dilution in every specimen (A06685 tissue IHC caption; standard IHC practice). Preserve plausible matrix staining during optimization because COL14A1 is an extracellular protein associated with collagen fibrils (UniProt Q05707 localisation and function).
What should I quantify when COL14A1 labels both matrix and cells? ⚠ ANSWER MARKED FOR VERIFICATION
Predefine separate regions for extracellular matrix and cells, then hold section area, threshold, and chromogen development constant across specimens (standard image-analysis practice). Report matrix-positive area or integrated staining density per mm² of annotated viable tissue, normalized to the area assessed (standard image-analysis practice). If a cellular signal is the study endpoint, report the percentage of positive cells or an H-score within a named cell compartment, with the number of evaluable cells recorded (standard IHC scoring practice). Keep these endpoints separate because COL14A1 is annotated as extracellular, whereas the available tissue IHC profile describes cytoplasmic expression with Uncertain reliability (UniProt Q05707 localisation; HPA tissue IHC).
When does a COL14A1-positive section support a biological conclusion?
Treat a reproducible extracellular pattern in intact tissue as more consistent with COL14A1’s annotated location than isolated nuclear or sharply edge-restricted staining (UniProt Q05707 localisation; standard IHC interpretation). Examine glandular-cell signals in context: HPA reports high staining in appendix and colon glandular cells, but rates its tissue IHC reliability Uncertain (HPA tissue IHC). Exclude necrotic areas, section edges, and brown signal reproduced in no-primary controls before calling cells or matrix positive (standard IHC practice). A clean control and agreement across adjacent sections strengthen the interpretation, while the paraffin-skin caption documents staining at 1:200 without specifying fixation (A06685 tissue IHC caption; standard IHC practice).
Boster reagents
Best COL14A1 / Collagen alpha-1(XIV) chain IHC Antibodies
A06685 has an IHC image from paraffin-embedded human skin and lists IF as an application; its stated reactivity covers human, mouse and rat (catalog: A06685 applications, reactivity and IHC caption).
Real IHC data
Anti-Collagen alpha-1(XIV) chain COL14A1 Antibody
Cat # A06685
A06685 has a human-skin paraffin-section IHC image with antibody diluted at 1:200 (A06685 IHC caption). IF is listed as an application, but no IF image is supplied; human, mouse and rat are listed as reactive species (catalog: A06685 applications, IF image alts and reactivity).
Which to pick: For tissue IHC, choose A06685: its image shows paraffin-embedded human skin at 1:200, with the fixative unreported (A06685 IHC caption). For IF/ICC, A06685 lists IF and a 1:50 dilution, though no IF image or ICC validation is supplied (catalog: A06685 applications, IF dilution and IF image alts). For cross-species work, A06685 lists human, mouse and rat reactivity; its pictured IHC result is human only, and the antibody host is rabbit with clonality unreported (catalog: A06685 reactivity, IHC caption, host and clone).
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 staining in most tissues (HPA tissue IHC). 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 across paraffin sections (standard IHC practice; not target-specific); Caveat: Tissue staining is uncertain for this secreted protein (HPA tissue IHC); Isoform / epitope: 3 isoforms; signal peptide removed; epitope unknown (UniProt); Regulation: Tissue-enhanced RNA in blood vessels (HPA tissue RNA). Evidence: UniProt Q05707, HPA tissue IHC · Uncertain, HPA ICC-IF · vesicles., Datasheet A06685, PMC9977988, PMC10401928. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/COL14A1/immunohistochemistry-ihc-guide.