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CD109 / CD109 antigen · IHC design guide
Design Immunohistochemistry for CD109
Plan CD109 IHC-P around membranous staining in parathyroid glandular cells and hair external root sheath cells (HPA tissue IHC). Start the catalog antibody at 1:50 (datasheet: 1:50) and compare staining with negative controls (standard IHC practice).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CD109 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 | Membranous in parathyroid and hair follicle (HPA tissue IHC) | |
| Staining pattern | Membranous glandular and external root sheath cells (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Hair+1 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 | Broad reported expression may not predict tissue staining (UniProt tissue specificity; HPA tissue IHC) | |
| Regulation | Expressed in activated T-lymphoblasts (UniProt) | |
| Isoform / epitope | 4 isoforms; check extracellular epitope coverage (UniProt) |
Section 1
Recommended CD109 IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet) is presented alongside four published CD109 IHC methods (PMC6639710; PMC7926025; PMC5929441; PMC5808841).
| Sample | Paraffin-embedded human liver tissue; fixative not specified (datasheet A03248) |
| Fixation | Image fixative and duration unreported (datasheet A03248); 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-CD109, 1:50 recommended; image 1:200 (datasheet A03248) |
| 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 | CD109-positive staining in cells in external root sheath of hair (HPA tissue IHC: High). HPA tissue profile: Membranous expression in parathyroid gland and hair follicle. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 HIER at 95–98 °C for 20 min (page retrieval). Tris-EDTA pH 9.0 improved staining in one study (PMC5929441).
Section 2
What Is the Expected CD109 Staining Pattern?
Expect CD109 chiefly at the cell surface: HPA reports membranous staining in parathyroid glandular cells and hair follicle external root sheath cells, both scored High (HPA tissue IHC). This agrees with UniProt cell-membrane localization despite the absence of a transmembrane segment (UniProt Q6YHK3 topology). HPA rates the tissue IHC pattern Enhanced, reflecting high agreement between staining and RNA expression (HPA tissue IHC).
| Clear membrane-edge staining in parathyroid glandular cells or hair external root sheath cells. | This matches the reported High, membranous positive pattern (HPA tissue IHC). Judge the signal against adjacent unstained areas and a no-primary control (general IHC practice). |
| Predominantly nuclear staining, with little membrane-edge signal in an expected positive cell. | Nuclear localization is unsupported by the supplied localization records (UniProt Q6YHK3; HPA subcellular). Treat it as a possible staining artefact and review controls before scoring it as CD109 (general IHC practice). |
| Strong staining in adipocytes or adrenal glandular cells. | Those specific cell populations are scored Not detected by HPA tissue IHC. Consider antibody cross-reactivity or endogenous chromogenic activity; a negative primary control helps distinguish detection background (HPA tissue IHC; general IHC practice). |
| Diffuse color coats several cell types and the surrounding tissue without a clear membrane outline. | That distribution is less convincing than the reported membranous pattern (HPA tissue IHC). Check the no-primary control, blocking and wash conditions before interpreting broad color as target signal (general IHC practice). |
| No staining in the expected cells of parathyroid gland or hair follicle. | This conflicts with the supplied High reference patterns (HPA tissue IHC). Verify that the expected cells are present, then review antibody and detection performance on a known-positive control (general IHC practice). |
💡Expected CD109 appearanceA convincing positive is High, predominantly membranous staining of parathyroid glandular or hair external root sheath cells (HPA tissue IHC); widespread nuclear or diffuse color without that pattern is suspect (UniProt Q6YHK3; HPA tissue IHC; general IHC practice).
| Membrane topology | UniProt places CD109 at the cell membrane, lists a GPI-anchor keyword and annotates no transmembrane segment (UniProt Q6YHK3). A membrane-edge IHC pattern is therefore consistent with the record; absence of a transmembrane segment does not require intracellular staining. |
| Tissue-specific reference pattern | HPA's High calls apply to named cell populations in parathyroid gland and hair, while its Not detected calls apply to the named cells in other sampled tissues (HPA tissue IHC). Do not turn those cell-level observations into a claim about every cell in an organ. |
| Antibody validation and assay scope | The supplied tissue IHC profile is Enhanced; one listed antibody has Enhanced IHC validation, while the other has Supported ICC validation (HPA tissue IHC; HPA antibodies). Validation in one application does not by itself establish the same staining pattern in the other. |
| Isoforms and processing | UniProt lists 4 isoforms, a signal peptide, a propeptide and a mature CD109 chain (UniProt Q6YHK3). The supplied IHC record does not resolve isoforms or identify the antibody epitope (HPA tissue IHC); avoid assigning a staining difference to either. |
| IF/ICC Q: where should signal appear? | A: HPA places the main ICC-IF signal at the plasma membrane and approves an additional cytosolic location (HPA subcellular). That cytosolic observation does not make a nuclear-dominant IHC result an expected pattern (HPA subcellular; HPA tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected-positive tissue is blank. | The sampled section may lack the relevant cells, or staining and detection may have failed (HPA tissue IHC; general IHC practice). | Confirm parathyroid glandular or hair external root sheath cells on the section (HPA tissue IHC). Check a known-positive control, antibody application conditions and detection reagents (general IHC practice). |
| Signal appears in the no-primary control. | Color can arise from endogenous detection activity or nonspecific detection reagents (general IHC practice). | Review the detection chemistry, endogenous-activity blocking and wash steps; score CD109 only after control staining is resolved (general IHC practice). |
| Nuclei dominate the stained image. | This does not fit the supplied membrane-centered localization (UniProt Q6YHK3; HPA tissue IHC). Background or detection artefact is possible (general IHC practice). | Compare with the no-primary control and inspect a known-positive section for membrane-edge signal before accepting the result (HPA tissue IHC; general IHC practice). |
| Adipocytes or adrenal glandular cells stain strongly. | HPA scores these cell populations Not detected; cross-reactivity or endogenous chromogenic activity is possible (HPA tissue IHC; general IHC practice). | Check no-primary controls and compare staining with the expected positive cell populations. Investigate persistent cell-specific staining before assigning it to CD109 (HPA tissue IHC; general IHC practice). |
| The whole section has weak, diffuse color. | Broad color without a membrane outline is less consistent with the HPA tissue pattern; incomplete blocking or washing may contribute (HPA tissue IHC; general IHC practice). | Review blocking, washes and chromogen development alongside a no-primary control. Reassess whether membrane staining remains after background is reduced (general IHC practice). |
| A broader tissue shows an unexpected result. | UniProt describes wider tissue expression, whereas the supplied HPA IHC calls document specific sampled cell populations (UniProt Q6YHK3 tissue specificity; HPA tissue IHC). | Record the exact cell type and compartment, then compare with matched controls. Avoid treating expression reported by UniProt alone as a confirmed positive IHC pattern (UniProt Q6YHK3; HPA tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing CD109 staining
A03248 · IHCImmunohistochemical analysis of paraffin-embedded human-liver, antibody was diluted at 1:200
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000156535-CD109/tissue/cerebral+cortex#imid_20368685
HPA · Parathyroid glandHuman Protein Atlas IHC, Parathyroid gland. Source: https://www.proteinatlas.org/ENSG00000156535-CD109/tissue/parathyroid+gland#imid_20368560
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000156535-CD109/tissue/colon#imid_20368595
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000156535-CD109/tissue/liver#imid_20368604
HPA · ICC-IF ASC52teloHuman Protein Atlas immunofluorescence, ASC52telo, antibody HPA008992; green = CD109, blue = nucleus, red = microtubules.
HPA · ICC-IF ASC52teloHuman Protein Atlas immunofluorescence, ASC52telo, antibody HPA008992; green = CD109, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CD109 IHC & IF
🧪Run hair first: cells in the external root sheath should stain (HPA: High). Use adipose tissue adipocytes as the negative comparison (HPA: Not detected); on the hair section, treat cells without specific staining as an internal background reference rather than assuming an unlisted cell type is CD109-negative (standard IHC practice).
Positive control tissue: Hair (Cells in external root sheath, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CD109 in ASC52telo, HaCaT, U2OS, with annotated localisation: Plasma membrane (supported) (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 a CD109 knockout specimen or peptide-block control if the immunizing peptide is available (standard IHC specificity controls). For chromogenic hair sections, quench endogenous peroxidase and compare pigmented areas with the no-primary section because hair pigment can complicate interpretation (standard IHC practice).
⚠️Feasibility: A target-specific fixation window and retrieval dependence are unreported in the supplied evidence; the 1:200 paraffin-section liver caption for A03248 does not report a fixative (datasheet: A03248 liver IHC caption). Optimize antigen retrieval using the hair positive and adipose negative comparisons (HPA: external root sheath High; adipocytes Not detected). The evidence does not establish whether frozen sections or IF are easier; IF interpretation can use plasma-membrane staining as a localization reference and should include an autofluorescence control (HPA subcellular: plasma membrane supported; standard IF practice).
HPA tissue IHC evidence for CD109
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 |
|---|---|---|---|---|
| Hair | Cells in external root sheath | High | Protein (IHC) | HPA → |
| Parathyroid gland | Glandular cells | High | 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 → |
| Appendix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CD109 IHC Tips
Use cell-border staining, appropriate tissue controls and matched processing to troubleshoot CD109 chromogenic IHC; interpret cytoplasmic signal cautiously (HPA tissue IHC; HPA subcellular).
How should I adjust retrieval when CD109 staining is weak or diffuse?
Start with citrate pH 6.0 heat-induced epitope retrieval at 95–98 °C for 20 min (page retrieval rule: membrane antigen). Judge the result by defined cell-border staining, since CD109 is membrane-associated without a transmembrane segment and HPA reports membranous tissue expression (UniProt Q6YHK3 topology; HPA tissue IHC). Compare matched sections processed for the same time and temperature, using the external root sheath or parathyroid glandular cells as positive controls (standard IHC practice; HPA: High in both cell populations). If staining remains weak, test a longer citrate incubation or an alternate retrieval buffer on parallel sections, checking whether background and tissue damage rise alongside signal (standard IHC practice).
Could fixation explain inconsistent CD109 staining between paraffin blocks?
The paraffin-embedded human-liver image reports an antibody dilution of 1:200 but does not state its fixative, so CD109-specific fixation sensitivity is unknown (A03248 tissue-IHC caption). Record each block’s fixative and fixation time before comparing staining, and process control and study sections together where possible (standard IHC practice). If blocks differ, compare matched positive-control sections after the same citrate pH 6.0 retrieval at 95–98 °C for 20 min (standard IHC practice; page retrieval rule: membrane antigen). Treat a difference confined to one processing batch as a reason to investigate processing before assigning it to biological CD109 variation (standard IHC interpretation).
What CD109 staining compartment should I expect in tissue sections?
Prioritize a defined cell-border pattern: CD109 is assigned to the cell membrane, and HPA reports membranous expression in parathyroid gland and hair follicle (UniProt Q6YHK3 subcellular; HPA tissue IHC). HPA cell imaging also supports plasma-membrane localization and records additional cytosolic localization, so cytoplasmic staining deserves assessment alongside its cell-border pattern (HPA subcellular). Compare the stained cells with tissue architecture and a counterstain, especially in the external root sheath and parathyroid glandular cells reported as High (standard IHC practice; HPA tissue IHC). Diffuse signal across unrelated structures or section edges is less persuasive than reproducible staining in the expected cells and compartment (standard IHC interpretation; HPA tissue IHC).
Can this IHC stain distinguish CD109 isoforms or epitope accessibility?
CD109 has 4 annotated isoforms, while isoform 4 is reported in placenta and isoform 1 in keratinocytes and placenta (UniProt Q6YHK3 isoforms; UniProt Q6YHK3 tissue specificity). A positive chromogenic stain alone cannot identify an isoform unless the antibody’s recognized epitope and its presence in each isoform are established (standard antibody interpretation). CD109 is processed from a 1445-residue precursor and has 10 annotated glycosylation sites, factors to consider when checking where an epitope falls (UniProt Q6YHK3 processing; UniProt Q6YHK3 glycosylation). Compare epitope documentation and isoform sequences before making an isoform claim, then verify the proposed specificity with an independent control (standard IHC practice).
How should I investigate CD109 localization by multiplex IF?
For a separate IF experiment, pair CD109 with a validated marker for the cell population under study and inspect whether CD109 outlines those cells (standard IF practice; HPA subcellular: plasma membrane). HPA reports High tissue staining in external root sheath cells and parathyroid glandular cells, which can guide the choice of a relevant cell-type marker (HPA tissue IHC). Select fluorophores after inspecting unstained tissue autofluorescence, and separate emission channels sufficiently to evaluate signal overlap (standard IF practice). Decide whether to permeabilize only after establishing whether the antibody epitope is extracellular or intracellular; compare conditions if its orientation is unknown (standard IF practice; UniProt Q6YHK3: membrane-associated topology).
How can I reduce widespread brown staining without losing CD109 signal?
Compare a no-primary control with the stained section to identify signal from detection reagents or endogenous enzyme activity (standard chromogenic IHC practice). Include a peroxidase-blocking step when using peroxidase and DAB detection, and check that blocking, washes and chromogen development are consistent across sections (standard chromogenic IHC practice). Titrate the catalog antibody around the reported 1:200 dilution on matched sections, scoring cell-border contrast against background rather than overall darkness (A03248 tissue-IHC caption; standard IHC practice). Use HPA High external root sheath or parathyroid glandular cells for context, while checking adipocytes reported as Not detected for unexpected signal (HPA tissue IHC).
How should I score CD109 staining across samples? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, because HPA reports membranous staining in parathyroid gland and hair follicle (HPA tissue IHC). For each comparable region, record the percentage of positive cells and staining intensity, then calculate an H-score with the same thresholds and counterstain criteria across sections (standard IHC quantification practice). If positive cells are sparse, report positive-cell density per mm² of viable tissue alongside the cell count used as its denominator (standard IHC quantification practice). Normalize comparisons to the same eligible cell population or viable tissue area, and exclude folds, necrosis and section edges consistently (standard IHC quantification practice).
When is apparent CD109 positivity likely to be artefact?
Give greatest weight to reproducible cell-border staining in plausible cells: HPA reports membranous expression and High staining in external root sheath and parathyroid glandular cells (HPA tissue IHC). Cytoplasmic signal is possible in cell imaging, but diffuse staining without a coherent cell pattern needs corroboration (HPA subcellular: additional cytosolic localization; standard IHC interpretation). Inspect section edges and necrotic areas separately, since local staining there can mislead scoring, and use a no-primary control to assess detection-related colour (standard IHC practice). Interpret apparent positivity in adipocytes cautiously because HPA reports them as Not detected; confirm any unexpected pattern with appropriate independent controls (HPA tissue IHC; standard IHC practice).
Boster reagents
Best CD109 / CD109 antigen IHC Antibodies
A03248 has a paraffin-section IHC image from human liver (catalog image caption); IF is listed, but no IF image is supplied (catalog applications and images). Human and Mouse reactivity are listed (catalog reactivity).
Real IHC data
Anti-CD109 antigen CD109 Antibody
Cat # A03248
A03248 will render with an IHC image of paraffin-embedded human liver, whose caption reports 1:200 (catalog image caption). The catalog lists IHC and IF applications and Human and Mouse reactivity for A03248, but supplies no IF image or Mouse validation image (catalog applications, reactivity, and images).
Which to pick: For tissue IHC, choose A03248 based on its paraffin-section human-liver image; the fixative is unreported (catalog image caption). For IF, A03248 lists IF at 1:50, but has no IF image; ICC is not listed, and the clone is unreported (catalog applications, dilution, images, and clone). For cross-species work, A03248 lists Human and Mouse reactivity, while its IHC image documents Human tissue only (catalog reactivity and image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q6YHK3 (CD109_HUMAN, CD109 antigen).
- Human Protein Atlas. CD109 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. CD109 subcellular location (ICC-IF): Mainly localized to the plasma membrane. In addition localized to the cytosol..
- Human Protein Atlas. CD109 antibody validation summary (2 antibodies).
- Prognostic Significance of CD109 Expression in Patients with Ovarian Epithelial Cancer. Journal of pathology and translational medicine 2019 — PMC6639710.
- Serum CD109 levels reflect the node metastasis status in head and neck squamous cell carcinoma. Cancer medicine 2021 — PMC7926025.
- Development of novel monoclonal antibodies against CD109 overexpressed in human pancreatic cancer. Oncotarget 2018 — PMC5929441.
- Myxofibrosarcoma primary cultures: molecular and pharmacological profile. Therapeutic advances in medical oncology 2017 — PMC5808841.
- PubMed PMID:11861284 — UniProt-cited evidence.
- PubMed PMID:14980714 — UniProt-cited evidence.
- PubMed PMID:16754747 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CD109 (CD109 antigen). Expected localisation Membranous in parathyroid and hair follicle (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A03248 at 1:50 recommended; image 1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Hair (Cells in external root sheath); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human liver tissue; fixative not specified (datasheet A03248); 2. Fixation: Image fixative and duration unreported (datasheet A03248); 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-CD109, 1:50 recommended; image 1:200 (datasheet A03248); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Broad reported expression may not predict tissue staining (UniProt tissue specificity; HPA tissue IHC); Isoform / epitope: 4 isoforms; check extracellular epitope coverage (UniProt); Regulation: Expressed in activated T-lymphoblasts (UniProt). Evidence: UniProt Q6YHK3, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the plasma membrane. In addition localized to…, Datasheet A03248, PMC6639710, PMC7926025. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CD109/immunohistochemistry-ihc-guide.