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- Table of Contents
Plan chromogenic DBN1 IHC in paraffin sections using cerebellar granular-layer processes as a strong-staining reference (HPA tissue IHC). The guide covers fixation consistency, staining expectations and how to interpret weaker hippocampal neuronal staining (HPA tissue IHC).
Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
| Expected localisation | Cytoplasmic and membranous tissue staining (HPA tissue IHC) | |
| Staining pattern | Strong staining in cerebellar granular-layer processes (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A05530-1) | |
| Positive control | Cerebellum+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
| Fixation | Keep fixation consistent across sections (standard IHC practice; not target-specific). Selected-image fixative and duration unreported (datasheet A05530-1); verify before use. | |
| Caveat | Hippocampal neurons stain low despite high cerebellar processes (HPA tissue IHC) | |
| Regulation | Intensity regulation is unspecified (UniProt) | |
| Isoform / epitope | 3 isoforms; isoform-specific epitope coverage is unknown (UniProt) |
The catalog antibody’s IHC protocol uses EDTA pH 8.0 heat retrieval (datasheet: A05530-1). The published options below report only the IHC steps stated in each article (PMC12262305; PMC7352383).
| Sample | Paraffin-embedded human laryngeal carcinoma tissue; fixative not specified (datasheet A05530-1) |
| Fixation | Image fixative and duration unreported (datasheet A05530-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: EDTA pH 8.0 (datasheet A05530-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A05530-1) |
| Primary antibody | Rabbit anti-DBN1, 2-5 μg/ml (datasheet A05530-1) |
| Primary incubation | Overnight at 4 °C (datasheet A05530-1) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A05530-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | DBN1-positive staining in processes in granular layer of cerebellum (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic and membranous expression in several tissues, most abundant in CNS. No signal in the no-primary control. |
DBN1 is a cytoplasmic, cortical and cell projection protein with no transmembrane segment (UniProt Q16643). In paraffin-section IHC, expect cytoplasmic or membranous staining, most abundant in CNS tissue (HPA tissue IHC: Enhanced). Examine neuronal processes and neuropil as well as cell bodies: HPA reports high staining in cerebellar granular-layer processes, medium staining in cerebral cortical neuropil and low staining in hippocampal neuronal cells (HPA tissue IHC).
| Cerebellar granular-layer processes stain strongly; cerebral cortical neuropil stains less strongly (HPA tissue IHC). | This distribution fits HPA's high cerebellar process and medium cortical neuropil observations (HPA tissue IHC). Assess the named structures separately from neighboring cells; a section-wide intensity judgment can obscure the expected pattern. UniProt also places DBN1 in dendrites, the cell cortex and growth cones (UniProt Q16643). |
| The dominant signal is confined to nuclei or appears as a sharply extracellular deposit. | That compartment does not fit the reported cytoplasmic, projection and junction locations (UniProt Q16643), or HPA's cytoplasmic and membranous tissue profile (HPA tissue IHC). Treat it as suspect until controls and tissue morphology support it; a compartment mismatch alone does not identify the source of artefact (standard IHC interpretation). |
| Signal appears in adipocytes or cardiomyocytes, while expected positive structures stain poorly (HPA tissue IHC). | HPA reports DBN1 as not detected in those particular cell populations (HPA tissue IHC). Investigate cross-reactivity or endogenous detection activity before assigning the signal to DBN1 (standard IHC practice). A negative HPA entry concerns the specified cell type; it does not establish that every cell in that tissue is negative. |
| Uniform chromogen covers cells, stroma and otherwise pale areas, obscuring structure. | A broad haze is difficult to reconcile with HPA's cell-specific staining levels and cytoplasmic or membranous profile (HPA tissue IHC). Consider nonspecific binding, endogenous detection activity or excess chromogen development (standard IHC practice). Judge specificity from recognizable cells and compartments rather than overall slide darkness. |
| No signal appears in cerebellar granular-layer processes (HPA tissue IHC). | This misses an HPA high reference pattern, so interpret other negative fields cautiously (HPA tissue IHC). Check section integrity and the staining run before concluding DBN1 is absent (standard IHC practice). HPA's Enhanced reliability describes agreement between staining and RNA data; it does not guarantee that every specimen or run will stain. |
| Cell and tissue choice | Use cerebellar granular-layer processes as a strong pattern reference; HPA scores them high, whereas hippocampal neuronal cells are low (HPA tissue IHC). A weak hippocampal result therefore has less power to diagnose a failed run. UniProt reports expression in hippocampal regions (UniProt Q16643); that statement does not override HPA's cell-specific IHC score. |
| Antibody evidence | HPA rates tissue IHC reliability Enhanced and lists 2 rabbit polyclonal antibodies, HPA051452 and HPA056940, with Enhanced IHC validation (HPA tissue IHC; HPA antibodies). These observations support the reference pattern but do not validate an unspecified antibody, dilution or detection system. |
| Epitope and molecular form | UniProt lists 3 isoforms, a chain spanning residues 2–649, an ADF-H domain at 3–134 and multiple modified residues (UniProt Q16643). Epitope location for the antibody used here is unreported; these features cannot predict which isoforms it detects or whether retrieval changes its signal. |
| IF/ICC Q&A: where should cellular signal appear? | HPA's ICC-IF summary places DBN1 mainly at the plasma membrane, with additional actin-filament localization (HPA subcellular ICC-IF). UniProt also reports cytoplasmic and cell-cortex locations (UniProt Q16643). This answers cellular localization only; ICC-IF observations do not set a paraffin-section IHC staining score or an IF protocol. |
| Situation | Likely cause | Next action |
|---|---|---|
| No staining in the chosen test tissue. | The selected cell population may have a weak reference pattern: hippocampal neuronal cells are low by HPA IHC (HPA tissue IHC). | Review a cerebellar section containing granular-layer processes, scored high by HPA (HPA tissue IHC). If that reference also fails, check reagent integrity, detection steps and section quality (standard IHC practice); do not infer DBN1 absence from an unverified run. |
| Cerebellar processes are present but unstained. | A failed staining step or unsuitable assay conditions are possibilities (standard IHC practice); the payload gives no DBN1-specific fixation sensitivity or retrieval requirement. | Confirm tissue morphology and the run controls, then review the antibody's documented IHC conditions and the retrieval and detection steps used (standard IHC practice). Record any adjustment as assay optimization, without claiming that fixation masked a DBN1 epitope. |
| Strong nuclear-only staining dominates. | Nuclear restriction conflicts with UniProt's cytoplasmic, cortical and projection locations and HPA's tissue profile (UniProt Q16643; HPA tissue IHC). | Inspect morphology, compare expected positive structures and repeat with appropriate negative controls (standard IHC practice). If available, compare an independently validated antibody; HPA lists 2 antibodies with Enhanced IHC validation (HPA antibodies). |
| Chromogen appears in HPA-negative cell populations. | Cross-reactivity or endogenous detection activity may explain staining in adipocytes or cardiomyocytes, both not detected by HPA IHC (HPA tissue IHC; standard IHC practice). | Check a no-primary control and any detection-system controls appropriate to the run, then reassess the exact cell type (standard IHC practice). Do not extend a cell-specific HPA negative call to the entire tissue. |
| Diffuse background hides cell boundaries. | Nonspecific binding, incomplete blocking or detection background can reduce contrast (standard IHC practice); HPA does not establish a DBN1-specific cause. | Review blocking, washing, primary-antibody concentration and chromogen development against the run controls (standard IHC practice). Score only clearly localized signal after background is controlled, using the cytoplasmic and membranous HPA pattern as a reference (HPA tissue IHC). |
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.
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Cerebellum | Processes in granular layer | High | Protein (IHC) | HPA → |
| Testis | Spermatogonia cells | High | Protein (IHC) | HPA → |
| Cerebral cortex | Neuropil | Medium | Protein (IHC) | HPA → |
| Endometrium | Cells in endometrial stroma | Medium | Protein (IHC) | HPA → |
| Epididymis | Glandular cells | Medium | Protein (IHC) | HPA → |
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Appendix | Endocrine cells | Not detected | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Not detected | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Not detected | Protein (IHC) | HPA → |
Troubleshoot DBN1 staining in paraffin sections by checking retrieval, detection, and the expected cellular pattern before comparing samples.
The catalog shows paraffin-section IHC images from human tissue and rat brain (IHC image captions), plus an IF image from CACO-2 cells (A05530-1 IF image caption).
A05530-1 has IHC images from human laryngeal carcinoma and cerebellum (A05530-1 IHC image captions) and an IF image from CACO-2 cells (A05530-1 IF image caption). M05530-4 has IHC images from human ovarian serous carcinoma and rat brain (M05530-4 IHC image captions).
Which to pick: For paraffin-section IHC, choose A05530-1 for a rabbit primary or M05530-4 for a mouse monoclonal primary (catalog hosts and clone; each SKU’s IHC image captions); both captions report EDTA retrieval at pH 8.0, while fixation is unreported (IHC image captions). For IF/ICC, choose A05530-1: IF and ICC are listed applications, with a CACO-2 IF image at 5 μg/ml (A05530-1 catalog applications and IF image caption). For cross-species tissue work, M05530-4 lists human, mouse, rat and monkey reactivity, with IHC images for human and rat; A05530-1 lists human, rat and monkey reactivity, with IHC images for human (catalog reactivity; each SKU’s IHC image captions).