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B9D1 / B9 domain-containing protein 1 · IHC design guide
Design Immunohistochemistry for B9D1
Plan chromogenic B9D1 IHC in paraffin sections using bronchial ciliated cell bodies as a high-staining reference and smooth muscle cells as a reported unstained comparison (HPA tissue IHC). Start the IHC-validated antibody within its 1:50–1:200 IHC dilution range (datasheet: A10722-1).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
B9D1 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, including bronchial ciliated cell bodies (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Caudate+2 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep tissue fixation consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Staining has medium agreement with RNA expression (HPA tissue IHC) | |
| Regulation | Expression regulator not annotated (UniProt) | |
| Isoform / epitope | 2 isoforms; epitope coverage is unspecified (UniProt) |
Section 1
Recommended B9D1 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol (datasheet) with one published B9d1 paraffin-section protocol (PMC4395494).
| Sample | Paraffin-embedded human stomach tissue; fixative not specified (datasheet A10722-1) |
| Fixation | Image fixative and duration unreported (datasheet A10722-1); 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-B9D1, 1:50-1:200 (datasheet A10722-1) |
| 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 | B9D1-positive staining in glandular cells of adrenal gland (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 rule); the published protocol uses SDS retrieval (PMC4395494).
Section 2
What Is the Expected B9D1 Staining Pattern?
B9D1 localizes to the ciliary basal body, axoneme and transition zone (UniProt Q9UPM9 subcellular location); it has no transmembrane segment (UniProt Q9UPM9 topology). In paraffin tissue sections, expect chiefly cytoplasmic staining, including high staining in bronchial ciliated cell bodies and several glandular cell populations (HPA tissue IHC). Treat this as a reference pattern with limited certainty: HPA rates its tissue IHC reliability Approved, with medium staining–RNA consistency and external verification pending (HPA tissue IHC).
| Cytoplasmic staining in bronchial ciliated cell bodies, with high staining in the same cell population across the section (HPA tissue IHC). | This fits an observed tissue pattern (HPA: High in bronchial ciliated cell bodies). The basal body and transition zone are more precise localization expectations (UniProt Q9UPM9 subcellular location); a diffuse chromogenic cell-body signal alone cannot resolve those small structures (general IHC interpretation). |
| Predominantly nuclear staining in a paraffin section, without the expected cytoplasmic pattern (HPA tissue IHC). | Investigate compartment-specific background or antibody specificity before assigning a tissue IHC result (general IHC interpretation). HPA describes general cytoplasmic tissue expression (HPA tissue IHC); its uncertain nucleoplasm assignment comes from ICC-IF and does not establish a nuclear paraffin-section pattern (HPA subcellular ICC-IF). |
| Strong staining confined to ovarian stroma cells, caudate glial cells or smooth muscle cells (HPA tissue IHC). | These populations are listed as Not detected (HPA tissue IHC). Check for cross-reactivity, endogenous detection activity or misidentified cell types (general IHC troubleshooting). A discrepant result needs validation; the HPA tissue reference remains pending external verification (HPA tissue IHC reliability). |
| Diffuse staining across cells and extracellular spaces, with little distinction between cells expected to differ in signal (general IHC interpretation). | Treat uniform haze as possible background rather than evidence of widespread B9D1 expression (general IHC interpretation). Compare the same run with a no-primary control and with documented high and Not detected populations (general IHC controls; HPA tissue IHC). |
| No visible staining in bronchial ciliated cell bodies or another documented high population (HPA tissue IHC). | First suspect a failed staining run or inadequate assay sensitivity (general IHC troubleshooting). Confirm tissue identity and preservation, then check retrieval, primary-antibody use and detection against the validated IHC-P instructions if available (general IHC practice). Absence alone cannot overturn an Approved reference pattern that awaits external verification (HPA tissue IHC reliability). |
💡Expected B9D1 appearanceCall positive when cytoplasmic signal is clear in documented high cell populations, especially bronchial ciliated cell bodies (HPA tissue IHC); isolated strong staining in HPA Not detected cell types or an unsupported nuclear-only pattern needs investigation (HPA tissue IHC; general IHC interpretation).
| Subcellular scale and topology | B9D1 is reported at the basal body, axoneme and transition zone, with no transmembrane segment (UniProt Q9UPM9). Score chromogenic tissue IHC at the cell and compartment level; do not demand that a punctum resolve the transition zone (general IHC interpretation). |
| Alternative splicing | Two isoforms are listed (UniProt Q9UPM9). The supplied record gives no antibody epitope or isoform-specific tissue pattern, so do not attribute a difference in staining to one isoform without separate validation (UniProt Q9UPM9; evidence limit). |
| Processing and annotated modifications | The record lists a single chain spanning residues 1–204, no signal peptide or propeptide, and no annotated glycosylation or modified residues (UniProt Q9UPM9). These annotations provide no basis to predict a processed, shed or modification-dependent IHC pattern (evidence limit). |
| Reference-pattern certainty | HPA rates the tissue IHC pattern Approved but reports medium consistency with RNA data and pending external verification; antibody HPA022957 is Approved for IHC and Uncertain for ICC (HPA tissue IHC; HPA antibodies). Use the tissue pattern as a comparison, then verify unexpected results with controls (general IHC practice). |
| IHC versus ICC-IF localization | HPA reports general cytoplasmic tissue staining (HPA tissue IHC), while ICC-IF additionally reports a supported basal body location and an uncertain nucleoplasm location (HPA subcellular ICC-IF). Preserve the application distinction when interpreting a paraffin section (general IHC interpretation). |
| Situation | Likely cause | Next action |
|---|---|---|
| A documented high population stains weakly or not at all (HPA tissue IHC). | The staining run may lack sensitivity, or tissue identification may be wrong (general IHC troubleshooting). | Confirm the cell population and run controls; check retrieval, primary-antibody conditions and chromogenic detection against the applicable IHC-P instructions (general IHC practice). |
| Brown signal appears broadly in cells listed as Not detected (HPA tissue IHC). | Cross-reactivity or endogenous detection activity can mimic target staining (general IHC troubleshooting). | Inspect a no-primary control and the detection-system controls; compare localization with documented high cell populations before scoring the signal (general IHC practice; HPA tissue IHC). |
| The section shows uniform haze or heavy staining outside identifiable cells (general IHC interpretation). | Nonspecific binding, insufficient blocking or excessive detection signal are possible causes (general IHC troubleshooting). | Review blocking, washes and detection exposure using the same controls and tissue type; score cell-associated staining only after background separates from it (general IHC practice). |
| A nuclear-only pattern appears in tissue IHC (HPA tissue IHC comparison). | The tissue reference describes general cytoplasmic expression; ICC-IF nucleoplasm localization is uncertain (HPA tissue IHC; HPA subcellular ICC-IF). | Check counterstain and no-primary control, then seek independent specificity evidence before calling nuclear B9D1 positive in the paraffin section (general IHC practice). |
| Ciliated cell bodies stain, but no discrete transition-zone dot is visible (HPA tissue IHC; UniProt Q9UPM9). | A chromogenic paraffin section may not resolve the small ciliary structure described by UniProt (general IHC interpretation; UniProt Q9UPM9 subcellular location). | Record the supported cell-body and cytoplasmic pattern; reserve a claim of precise subcellular localization for appropriately resolved imaging (HPA tissue IHC; general microscopy practice). |
| Can an ICC-IF image establish the expected paraffin-section pattern? | The applications have different reference observations: tissue IHC is generally cytoplasmic, while ICC-IF includes supported basal body and uncertain nucleoplasm assignments (HPA tissue IHC; HPA subcellular ICC-IF). | Use ICC-IF as localization context only; judge the IHC section against HPA tissue cell-type patterns and IHC controls (HPA tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing B9D1 staining
A10722-1 · IHCImmunohistochemistry of paraffin-embedded human stomach using B9D1 antibody at dilution of 1:100 .Perform microwave antigen retrieval with 1
A10722-1 · IHCImmunohistochemistry of paraffin-embedded rat kidney using B9D1 antibody at dilution of 1:100 .Perform microwave antigen retrieval with 10 m
A10722 · IFImmunocytochemistry of B9D1 in 293 cells with B9D1 antibody at 5 μg/mL.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000108641-B9D1/tissue/cerebral+cortex#imid_5948384
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000108641-B9D1/tissue/colon#imid_5948310
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000108641-B9D1/tissue/liver#imid_5948319
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000108641-B9D1/tissue/kidney#imid_5948320
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA022957; green = B9D1, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA022957; green = B9D1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for B9D1 IHC & IF
🧪Run bronchus first and look for staining in ciliated cells (cell body), reported as High (HPA: bronchus). Use smooth muscle cells as the negative tissue comparator (HPA: smooth muscle, Not detected). On the bronchus slide, assess neighboring nonciliated cells for lower staining or background, without assuming they are B9D1-negative (HPA: bronchus identifies ciliated cells only).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Caudate (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show B9D1 in A-431, U-251MG, U2OS, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), Sperm, with annotated localisation: Nucleoplasm (uncertain), Acrosome (approved), Equatorial segment (approved), Mid piece (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only section and an isotype control matched to the primary antibody’s host species and clonality; confirm specificity with knockout tissue or a validated peptide block (standard IHC practice). Block endogenous peroxidase and inspect inflammatory cells in bronchus for residual chromogen signal (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A10722-1 paraffin-section stomach caption does not state a fixative (selected tissue-IHC caption). The caption reports microwave retrieval in 10 mM PBS at pH 7.2 and a 1:100 dilution for stomach, but does not establish retrieval dependence for bronchus (selected tissue-IHC caption). The supplied evidence does not show that frozen sections or IF are easier; for bronchus, distinguish ciliated-cell staining from inflammatory-cell peroxidase background (HPA: bronchus, High in ciliated cells; standard IHC practice).
HPA tissue IHC evidence for B9D1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Medium consistency between antibody staining and RNA expression data. 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 |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Appendix | Lymphoid tissue | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Ciliated cells (cell body) | High | Protein (IHC) | HPA → |
Section 3
Advanced B9D1 IHC Tips
Use compartment-aware controls to optimize B9D1 staining in paraffin sections and assess whether chromogenic signal fits its ciliary localization.
Which retrieval conditions should I try first for weak B9D1 staining?
Start with citrate buffer at pH 6.0, heat-induced retrieval at 95–98 °C for 20 min (page retrieval setting). If staining remains weak, test microwave retrieval in 10 mM PBS at pH 7.2 on a parallel section; that condition was used for the catalog antibody in paraffin-embedded human stomach at 1:100 (A10722-1 tissue-IHC caption). Keep detection and exposure to chromogen consistent across conditions so that signal and background can be compared (standard IHC practice). Assess cytoplasmic staining separately from small basal-body or transition-zone signals, since B9D1 is associated with both compartments (HPA tissue IHC; UniProt Q9UPM9 subcellular location).
Could fixation explain inconsistent B9D1 staining between paraffin blocks?
The supplied B9D1 tissue-IHC caption does not report its fixative, so target-specific fixation sensitivity is unknown (A10722-1 tissue-IHC caption). Record each block’s fixative, fixation duration, processing history, and section age before comparing staining; differences in these variables can alter epitope accessibility in IHC (standard IHC practice). Test blocks side by side with the same pH 6.0 citrate retrieval for 20 min and the same detection run (page retrieval setting; standard IHC practice). Include a reference section from one block in every run, and interpret weaker staining cautiously until processing differences and tissue preservation have been assessed (standard IHC practice).
Should B9D1 appear throughout the cytoplasm or at the ciliary base?
Expect that routine chromogenic IHC may show general cytoplasmic staining, which is the reported tissue pattern (HPA tissue IHC). B9D1 is also assigned to the basal body, axoneme, and transition zone between them, where a small localized signal may be difficult to resolve in a tissue section (UniProt Q9UPM9 subcellular location). Compare stained cells with adjacent morphology and a matched negative control before calling a tiny deposit a ciliary focus (standard IHC practice). Nuclear-only staining warrants caution: HPA lists nucleoplasm as uncertain, whereas its basal-body assignment is supported (HPA subcellular); inspect both the compartment and staining controls before interpreting it as B9D1.
How should I assess an antibody epitope when B9D1 has two isoforms?
B9D1 has 2 annotated isoforms and a C2 B9-type domain spanning residues 9–127 (UniProt Q9UPM9 isoforms and domains). Check the catalog antibody’s stated immunogen and epitope against both isoform sequences before assuming that one IHC result represents both (standard antibody-validation practice). The supplied record identifies a 204-aa chain, no transmembrane segment, and no annotated glycosylation or modified residues; those annotations do not establish which epitope survives tissue processing (UniProt Q9UPM9 processing, topology, and PTMs). If isoform distinction matters, pair IHC with an independently validated isoform-specific assay, and report the antibody’s documented recognition scope rather than assigning an isoform from staining intensity alone (standard antibody-validation practice).
How can I check B9D1 localization with multiplex immunofluorescence?
For a separate IF experiment, pair B9D1 with a validated marker of the cell type being examined and, when testing a ciliary focus, a basal-body or ciliary marker (UniProt Q9UPM9 subcellular location; standard IF practice). Choose a fluorophore in a channel with low measured tissue autofluorescence, and check single-label controls for bleed-through before interpreting overlap (standard IF practice). B9D1 has no annotated transmembrane segment and is assigned to intracellular ciliary and cytoplasmic sites, so permeabilize after fixation to expose an intracellular epitope, then optimize its strength against preserved structure (UniProt Q9UPM9 topology and subcellular location; standard IF practice). The listed ICC/IF images establish imaging contexts, but supply no fixation or permeabilization conditions to transfer into this protocol (HPA subcellular).
What should I change when B9D1 chromogenic staining looks diffuse or granular?
Run a no-primary control and inspect unstained tissue for pigment or other deposits before attributing diffuse brown signal to B9D1 (standard IHC practice). Include a peroxidase block when using HRP and DAB, control chromogen development time, and compare background in stromal and glandular regions within the same section (standard chromogenic IHC practice). Titrate the catalog antibody around its reported 1:100 stomach-image dilution only as an optimization exercise, since that caption does not establish an optimum for other specimens (A10722-1 tissue-IHC caption; standard IHC practice). A broad cytoplasmic pattern can be compatible with reported tissue staining, but granular deposits alone do not establish a transition-zone signal (HPA tissue IHC; UniProt Q9UPM9 subcellular location).
How should I score B9D1 staining across tissue samples? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring; an H-score or percentage of positive cells can summarize cytoplasmic staining, while discrete candidate ciliary foci can be counted as density per mm² (HPA tissue IHC; UniProt Q9UPM9 subcellular location; standard IHC quantification practice). Apply the same positivity threshold and chromogen-development conditions to every sample (standard IHC quantification practice). Normalize cell-based scores to the number of evaluable cells in the specified population, and focus density to evaluable tissue area or the relevant cell count (standard IHC quantification practice). Report cytoplasmic intensity separately from candidate basal-body signal, because those measures answer different localization questions for B9D1 (HPA tissue IHC; UniProt Q9UPM9 subcellular location).
Which staining features would make a B9D1 positive call unconvincing?
A plausible result may include cytoplasmic staining or a localized basal-body or transition-zone signal, interpreted in the context of tissue morphology (HPA tissue IHC; UniProt Q9UPM9 subcellular location). Be cautious with nuclear-only staining, because HPA marks nucleoplasm uncertain, and with signal assigned to a cell population without checking its morphology (HPA subcellular; standard IHC practice). Edge-restricted staining, necrotic deposits, and signal persisting in a no-primary control favor processing or detection artefact (standard IHC practice). Exclude endogenous peroxidase or pigment when reading DAB, and seek agreement between staining pattern, reference tissue, and controls before making a B9D1-specific claim (standard chromogenic IHC practice; HPA tissue IHC).
Boster reagents
Best B9D1 / B9 domain-containing protein 1 IHC Antibodies
The catalog shows B9D1 IHC in paraffin-embedded human stomach, rat kidney and mouse lung, plus IF/ICC in 293 cells (catalog image captions).
Real IHC data
Anti-B9 domain-containing protein 1 B9D1 Antibody
Cat # A10722-1
Real IF data
Anti-B9 domain-containing protein 1 B9D1 Antibody
Cat # A10722
A10722-1 has IHC images from paraffin-embedded human stomach, rat kidney and mouse lung (A10722-1 IHC captions). A10722 has IF/ICC images from 293 cells (A10722 IF/ICC captions).
Which to pick: Choose A10722-1 for tissue IHC: IHC is listed for human, mouse and rat samples (A10722-1 catalog), and its own captions show paraffin-embedded sections stained at 1:100 after microwave retrieval in 10 mM PBS, pH 7.2 (A10722-1 IHC captions); the fixative is unreported (A10722-1 IHC captions). Choose A10722 for IF/ICC in human samples: those applications and human reactivity are listed (A10722 catalog), with pictured 293-cell staining at 5 μg/mL for ICC and 20 μg/mL for IF (A10722 IF/ICC captions). For mouse or rat IF/ICC, A10722-1 lists those species and applications (A10722-1 catalog); clonality is unreported for both SKUs (catalog clone fields).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9UPM9 (B9D1_HUMAN, B9 domain-containing protein 1).
- Human Protein Atlas. B9D1 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. B9D1 subcellular location (ICC-IF): Mainly localized to the nucleoplasm, acrosome, equatorial segment and mid piece. In addition localized to vesicles, cytosol, basal body, principal piece and end piece..
- Human Protein Atlas. B9D1 antibody validation summary (1 antibodies).
- TMEM231, mutated in orofaciodigital and Meckel syndromes, organizes the ciliary transition zone. The Journal of cell biology 2015 — PMC4395494.
- PubMed PMID:15489334 — UniProt-cited evidence.
- PubMed PMID:19208769 — UniProt-cited evidence.
- PubMed PMID:21493627 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for B9D1 (B9 domain-containing protein 1). 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: A10722-1 at 1:50-1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Caudate. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human stomach tissue; fixative not specified (datasheet A10722-1); 2. Fixation: Image fixative and duration unreported (datasheet A10722-1); 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-B9D1, 1:50-1:200 (datasheet A10722-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep tissue fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Staining has medium agreement with RNA expression (HPA tissue IHC); Isoform / epitope: 2 isoforms; epitope coverage is unspecified (UniProt); Regulation: Expression regulator not annotated (UniProt). Evidence: UniProt Q9UPM9, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the nucleoplasm, acrosome, equatorial segment…, Datasheet A10722-1, PMC4395494. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/B9D1/immunohistochemistry-ihc-guide.