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- Table of Contents
Plan FUBP1 paraffin-section IHC around the widespread nuclear staining reported in tissue (HPA tissue IHC). This guide covers fixation consistency, antibody dilution, chromogenic detection and interpretation of nuclear staining.
Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
| Expected localisation | Nuclear staining in tissue (HPA tissue IHC) | |
| Staining pattern | Nuclear staining across many cell types (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet M03126) | |
| Positive control | Adipose tissue+4 more · see all | |
| Negative control | None in HPA (detected in all 45 tissues); use no-primary + isotype controls |
| Fixation | Keep fixation consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Ubiquitous staining limits negative tissue controls (HPA tissue IHC) | |
| Regulation | Low tissue specificity (HPA tissue RNA) | |
| Isoform / epitope | 2 isoforms; epitope coverage is unresolved (UniProt) |
The catalog antibody’s IHC-P protocol (datasheet M03126) is followed by published FUBP1 staining procedures for murine cell preparations (PMC5447289) and human paraffin sections (PMC10725354; PMC8902172).
| Sample | Paraffin-embedded human breast cancer tissue; fixative not specified (datasheet M03126) |
| Fixation | Image fixative and duration unreported (datasheet M03126); 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 M03126); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet M03126) |
| Primary antibody | Rabbit monoclonal (clone AHC-6) anti-FUBP1, 1:50 (datasheet M03126) |
| Primary incubation | Overnight at 4 °C (datasheet M03126) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet M03126) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | FUBP1-positive staining in adipocytes of adipose tissue (HPA tissue IHC: High). HPA tissue profile: Ubiquitous nuclear expression. No signal in the no-primary control. |
FUBP1 should appear predominantly in nuclei (UniProt Q96AE4: nucleus), with nucleoplasmic localisation also observed by ICC-IF (HPA subcellular: nucleoplasm). Tissue IHC describes ubiquitous nuclear expression and high staining in several listed cell populations (HPA tissue IHC: Supported; medium consistency with RNA). FUBP1 has no transmembrane segment, so a membrane pattern is unexpected (UniProt Q96AE4: topology).
| Many cell nuclei stain across the section, including the expected cells in the tissue examined. | This fits the reported ubiquitous nuclear pattern (HPA tissue IHC). High staining is reported in adipocytes, glandular cells, hematopoietic cells, respiratory epithelial cells and specified brain cell populations (HPA tissue IHC: High in listed cells). Compare nuclei within the same section; do not require identical intensity in every cell. |
| Cytoplasm or cell borders stain more strongly than nuclei. | A dominant extranuclear pattern conflicts with the nuclear tissue profile and nucleoplasmic ICC-IF localisation (HPA tissue IHC; HPA subcellular). Treat it as a possible artefact or off-target signal, then inspect morphology and detection controls before interpreting it as FUBP1. |
| Only an unexpected cell population stains while expected nuclei remain blank. | Consider cross-reactivity or endogenous detection activity (general IHC practice). Cell identity alone cannot prove either cause: HPA reports ubiquitous nuclear expression and low RNA tissue specificity (HPA tissue IHC). Recheck compartment, controls and the expected positive cells before assigning specificity. |
| Colour covers tissue broadly, including spaces between cells, with poorly defined nuclei. | This does not provide convincing nuclear localisation (HPA tissue IHC: ubiquitous nuclear expression). Diffuse background can arise from nonspecific reagent binding or detection activity (general IHC practice). Judge the result against a no-primary control and look for discrete nuclear signal above background. |
| No nuclei stain in a tissue and cell population listed as high. | For example, HPA reports high staining in bone marrow hematopoietic cells and breast glandular cells (HPA tissue IHC). Absence of signal warrants a technical check, but does not alone establish loss of FUBP1: HPA rates tissue IHC Supported, with medium antibody–RNA consistency (HPA tissue IHC). |
| Expected compartment | Nuclear tissue staining is the main reference pattern (UniProt Q96AE4: nucleus; HPA tissue IHC). Nucleoplasmic ICC-IF localisation supports the compartment, but IF/ICC has its own guide and does not supply an IHC protocol here (HPA subcellular). |
| Distribution across tissues | HPA describes ubiquitous nuclear expression and low RNA tissue specificity (HPA tissue IHC). The supplied high examples are positive comparators, while the absence of negative examples gives no evidence for a deliberately negative tissue control (HPA tissue IHC: positive list; negative list empty). |
| Antibody evidence | The listed antibody HPA006149 has Supported IHC and Enhanced ICC validation (HPA antibodies). Those ratings apply to that antibody and assay; they do not establish identical staining with another antibody or turn an ICC-IF image into paraffin-section validation. |
| Protein forms | UniProt records two isoforms and a chain spanning residues 2–644 (UniProt Q96AE4: isoforms; processing). Without an epitope map for the antibody used, the record cannot establish whether both isoforms are detected or whether a particular modified residue affects staining. |
| Membrane and secretion context | No transmembrane segment, signal peptide or propeptide is recorded (UniProt Q96AE4: topology; processing). These annotations support prioritising intracellular nuclear signal; they do not identify an epitope or predict the outcome of antigen retrieval in a particular specimen. |
| Situation | Likely cause | Next action |
|---|---|---|
| Known positive tissue shows no nuclear colour. | The staining run may have failed, or the antibody conditions may be unsuitable (general IHC practice); HPA's high cell-level examples provide comparators (HPA tissue IHC). | Check that the expected cells are present, then review the IHC-validated antibody instructions, retrieval and detection steps; compare with a concurrently stained positive section (general IHC practice). |
| Nuclei are weak while the entire section is faint. | Low overall signal may reflect insufficient detection or an unsuitable antibody concentration (general IHC practice). A weak result alone cannot establish low FUBP1 expression in a listed high population (HPA tissue IHC). | Review reagent performance, dilution and chromogen development using the antibody's IHC instructions and a positive control (general IHC practice). Score cells only when nuclear colour is distinguishable from background. |
| Strong diffuse colour obscures nuclei. | Nonspecific binding or endogenous detection activity may contribute (general IHC practice); this appearance conflicts with the nuclear reference pattern (HPA tissue IHC). | Inspect a no-primary control, then review blocking, washes and detection chemistry (general IHC practice). Reassess the slide only after discrete nuclear staining can be separated from diffuse colour. |
| Cell borders or cytoplasm dominate the stain. | The compartment is inconsistent with nuclear FUBP1 and its lack of a transmembrane segment (UniProt Q96AE4: nucleus; topology). Off-target signal or a staining artefact is possible. | Compare with the no-primary control and a positive section; check whether any nuclei retain a distinct signal before calling the pattern FUBP1 (general IHC practice; HPA tissue IHC: nuclear expression). |
| Only one cell population stains in an otherwise blank section. | Selective off-target staining or endogenous activity is possible (general IHC practice), but HPA's ubiquitous nuclear profile does not define any supplied cell population as negative (HPA tissue IHC). | Confirm cell identity and nuclear localisation, inspect controls, and compare expected positive cells in the same section (general IHC practice; HPA tissue IHC). Avoid declaring other cells truly negative from this slide alone. |
| ICC-IF images and paraffin IHC appear different. | They are different assays: HPA reports nucleoplasmic ICC-IF localisation and tissue IHC nuclear expression, with separate validation ratings for HPA006149 (HPA subcellular; HPA antibodies). | Compare compartment and assay-specific controls before judging agreement (general IHC practice). Use the separate IF/ICC guide for fluorescence workflow; do not use ICC-IF validation as a paraffin IHC protocol. |
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Supported — Medium 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 |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | High | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: FUBP1 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Use the documented M03126 tissue staining conditions as the IHC starting point, then judge signal against FUBP1’s expected nuclear localisation.
Anti-FUBP1 IHC images cover human breast and colon cancer, mouse spleen and thymus, and rat thymus (catalog IHC captions). IF/ICC data show U2OS cells (A03126-2 IF caption).
M03126 has IHC images from paraffin sections of human breast and colon cancer, mouse thymus, and rat thymus (M03126 IHC captions). A03126-2 has IHC images from paraffin sections of mouse spleen and human breast and colon cancer, plus IF/ICC data from U2OS cells (A03126-2 IHC and IF captions).
Which to pick: For tissue IHC across the pictured species, choose monoclonal M03126: its paraffin-section captions show human, mouse, and rat samples at 1:50 (M03126 catalog; M03126 IHC captions). For IF/ICC, choose A03126-2, which has a U2OS cell image at 5 μg/ml; M03126 lists IF/ICC as applications but has no IF image in this payload (A03126-2 IF caption; M03126 catalog). Both IHC examples use EDTA retrieval at pH 8.0, and neither caption reports the fixative (M03126 and A03126-2 IHC captions).