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BUD31 / Protein BUD31 homolog · IHC design guide
Design Immunohistochemistry for BUD31
Plan chromogenic IHC for BUD31 in paraffin sections using the nuclear and cytoplasmic tissue pattern reported by HPA (HPA tissue IHC). This guide covers fixation, retrieval, controls and scoring, with the catalog antibody’s 1:50–1:200 IHC dilution range (datasheet: 1:50–1:200).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
BUD31 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 | Nuclear (UniProt); nuclear and cytoplasmic in tissue (HPA tissue IHC) | |
| Staining pattern | Glandular and other cells show nuclear and cytoplasmic staining (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen) | |
| Positive control | Adipose tissue+4 more · see all | |
| Negative control | Adrenal gland+3 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 | Staining and RNA show medium consistency (HPA tissue IHC) | |
| Regulation | Low tissue RNA specificity (HPA tissue RNA) | |
| Isoform / epitope | 2 isoforms; epitope coverage is unestablished (UniProt) |
Section 1
Recommended BUD31 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol (datasheet) with published BUD31 protocols for FFPE sections (PMC10094631; PMC9587234).
| Sample | Paraffin-embedded human tonsil carcinoma tissue; fixative not specified (datasheet A11662) |
| Fixation | Image fixative and duration unreported (datasheet A11662); 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 Tris-EDTA buffer, pH 9.0, 20 min at 95–98 °C (standard rule: nuclear 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-BUD31, 1:50-1:200 (datasheet A11662) |
| 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 | BUD31-positive staining in adipocytes of adipose tissue (HPA tissue IHC: Medium). HPA tissue profile: Cytoplasmic and nuclear expression in several tissues. No signal in the no-primary control. |
💡Decision noteStart with Tris-EDTA pH 9.0 HIER at 95–98 °C for 20 min (page retrieval rule); the published protocols use citrate or EDTA (PMC10094631; PMC9587234).
Section 2
What Is the Expected BUD31 Staining Pattern?
BUD31 is a nuclear protein with no transmembrane segment (UniProt P41223: localization and topology). In paraffin sections, expect nuclear staining in some epithelial and stromal cells, with possible cytoplasmic staining: HPA reports cytoplasmic and nuclear expression across several tissues, while UniProt reports protein in benign prostate hyperplasia epithelial and stromal cells (HPA tissue IHC; UniProt P41223: tissue specificity). HPA rates its tissue IHC profile Approved, with medium consistency against RNA expression (HPA tissue IHC).
| Nuclear staining in appendix or colon glandular cells, with possible cytoplasmic staining (HPA tissue IHC). | This fits the reported tissue pattern: both cell groups have Medium staining, and HPA describes cytoplasmic and nuclear expression (HPA tissue IHC). Score the compartments separately; cytoplasmic color alone is less persuasive for a protein UniProt places in the nucleus (UniProt P41223: localization). |
| Strong membrane outlining or staining restricted to a compartment other than the nucleus. | A membrane pattern conflicts with the reported nuclear localization and absence of a transmembrane segment (UniProt P41223: localization and topology). Cytoplasmic staining can occur (HPA tissue IHC), so assess whether convincing nuclei accompany it before calling the pattern an artefact. |
| Prominent color in adrenal gland glandular cells or ovarian stroma cells (HPA tissue IHC: Not detected). | Those sampled cell groups were Not detected by HPA; unexpected staining warrants review for cross-reactivity or endogenous chromogenic activity (HPA tissue IHC; general IHC practice). These observations do not prove every specimen from either tissue is BUD31-negative. |
| Diffuse color across tissue, including areas without clear cell boundaries or nuclear detail. | That distribution is difficult to assign to BUD31-positive cells; nonspecific reagent binding or detection background are possible causes (general IHC practice). Compare a matched control slide and judge cell-localized signal after background is controlled (general IHC practice). |
| No staining in appendix or colon glandular cells (HPA tissue IHC: Medium). | These are reasonable positive-control cell groups, though Medium is an observed level, not a guaranteed result in every section (HPA tissue IHC). Check tissue preservation, retrieval, antibody dilution and detection controls before concluding the specimen lacks BUD31 (general IHC practice). |
💡Expected BUD31 appearanceCall a result consistent when identifiable nuclei stain in an HPA-positive cell group, such as appendix or colon glandular cells with reported Medium staining; cytoplasmic color may accompany nuclei, whereas isolated membrane outlining or diffuse, cell-independent color is suspect (HPA tissue IHC; UniProt P41223: localization and topology; general IHC practice).
| Compartment and topology | UniProt places BUD31 in the nucleus and reports no transmembrane segment; HPA tissue IHC also reports cytoplasmic expression (UniProt P41223: localization and topology; HPA tissue IHC). Interpret mixed nuclear and cytoplasmic staining in that context, while investigating a membrane-only pattern. |
| Cell type and reference intensity | HPA reports Medium staining in appendix and colon glandular cells, Low staining in prostate glandular cells, and Not detected in adrenal gland glandular cells (HPA tissue IHC). Compare the same cell type when using these references; Low and Not detected are observed categories, not universal biological absolutes. |
| Strength of IHC evidence | HPA calls tissue IHC Approved but describes only medium consistency between staining and RNA expression; its listed antibody HPA028943 is IHC Approved (HPA tissue IHC; HPA antibodies). This supports a reference pattern without establishing that every positive structure in a new assay is specific. |
| Isoforms and antigen retrieval | UniProt lists 2 isoforms, but the supplied record gives no antibody epitope or isoform-specific IHC behavior (UniProt P41223: isoforms). Retrieval conditions should therefore be checked against the chosen antibody's validated procedure; no BUD31-specific retrieval effect is established here. |
| IF/ICC Q: What pattern should a separate IF/ICC assay show? | A: HPA supports mainly nucleoplasmic localization; additional microtubule and centrosome localizations are uncertain (HPA subcellular ICC-IF). Use that evidence to interpret IF images, while keeping this section's primary readout chromogenic IHC in paraffin sections. |
| Situation | Likely cause | Next action |
|---|---|---|
| Nuclei are unstained in appendix or colon glandular cells (HPA tissue IHC: Medium). | A missed assay step or inadequate signal is possible; the HPA level alone cannot identify the cause (HPA tissue IHC; general IHC practice). | Check section quality, retrieval, antibody dilution and detection reagents using the chosen antibody's validated IHC procedure (general IHC practice). |
| Only cytoplasm stains, with no convincing nuclear signal. | HPA reports cytoplasmic and nuclear tissue staining, but UniProt places BUD31 in the nucleus (HPA tissue IHC; UniProt P41223: localization). | Review nuclear counterstain and cellular boundaries, then compare with a known HPA-positive cell group and a matched control (HPA tissue IHC; general IHC practice). |
| Cell membranes form the dominant staining pattern. | That pattern is unsupported by the nuclear localization and lack of a transmembrane segment (UniProt P41223: localization and topology). | Reassess specificity with appropriate controls and inspect whether precipitate or edge staining is producing apparent membrane outlines (general IHC practice). |
| Adrenal gland glandular cells show prominent color (HPA tissue IHC: Not detected). | Cross-reactivity or endogenous detection activity is possible; the HPA observation cannot distinguish these causes (HPA tissue IHC; general IHC practice). | Compare a control without primary antibody and the chosen detection system's endogenous-activity control, then judge cell-specific staining (general IHC practice). |
| Color spreads over nuclei, cytoplasm and empty spaces alike. | Diffuse reagent background or chromogen deposit can obscure cellular localization (general IHC practice). | Inspect matched controls, washing and chromogen development; score only signal whose cellular compartment remains clear (general IHC practice). |
| A weak prostate glandular-cell result disagrees with stronger staining elsewhere. | HPA reports Low staining in prostate glandular cells but Medium in appendix and colon glandular cells (HPA tissue IHC). | Compare equivalent cell types and assay conditions before treating different tissue intensities as an assay failure (HPA tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing BUD31 staining
A11662 · IHCImmunohistochemistry (IHC) analyzes of BUD31 (E24) pAb in paraffin-embedded human tonsil carcinoma tissue at 1:50.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000106245-BUD31/tissue/cerebral+cortex#imid_7593536
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000106245-BUD31/tissue/colon#imid_7593462
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000106245-BUD31/tissue/liver#imid_7593455
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000106245-BUD31/tissue/kidney#imid_7593464
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA028943; green = BUD31, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA028943; green = BUD31, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for BUD31 IHC & IF
🧪Run appendix first and require staining in its glandular cells (Medium; HPA: appendix glandular cells). Use adrenal gland glandular cells as the negative tissue (Not detected; HPA: adrenal gland glandular cells); any adjacent cells used as internal negatives on the appendix slide should show only background chromogen, although their BUD31-negative status is unverified (HPA: appendix glandular cells).
Positive control tissue: Adipose tissue (Adipocytes, HPA Medium)
Negative control tissue: Adrenal gland (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show BUD31 in A-431, U-251MG, U2OS, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) control, a host-species- and clonality-matched isotype control, and BUD31 knockout material or antigen-peptide competition if available (standard IHC practice). For appendix sections, check and quench endogenous peroxidase before chromogenic detection (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported; the A11662 paraffin-section caption gives 1:50 for human tonsil carcinoma but does not state a fixative (A11662 caption). BUD31-specific antigen-retrieval dependence is unreported, so optimize retrieval and assess nuclear staining (UniProt P41223: nucleus; standard IHC practice). Frozen-section feasibility and whether IF is easier are unreported; ICC-IF images support nucleoplasmic localization, while endogenous peroxidase may complicate chromogenic assessment in appendix (HPA: nucleoplasm supported; standard IHC practice).
HPA tissue IHC evidence for BUD31
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Medium | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Medium | Protein (IHC) | HPA → |
| Caudate | Glial cells | Medium | Protein (IHC) | HPA → |
| Cerebellum | Cells in molecular layer | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Ovary | Ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Soft tissue | Peripheral nerve | Not detected | Protein (IHC) | HPA → |
| Spleen | Cells in red pulp | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced BUD31 IHC Tips
Troubleshooting BUD31 staining in paraffin sections, with one entry for IF/ICC assay design.
How should I retrieve BUD31 in paraffin sections when nuclear staining is weak?
Use Tris-EDTA at pH 9.0 for heat-induced retrieval at 95–98 °C for 20 min (page retrieval setting: nuclear antigen). Keep section thickness, heating vessel and cooling time consistent while comparing runs, since these affect staining intensity (standard IHC practice). If nuclear signal remains weak, compare a shorter and a longer heating interval on adjacent sections before changing antibody concentration (standard IHC practice). Judge improvement by crisp staining within cells rather than by overall tissue darkness: BUD31 is nuclear by UniProt, while HPA tissue IHC reports both cytoplasmic and nuclear expression (UniProt P41223; HPA tissue IHC).
Could fixation be responsible for weak or uneven BUD31 staining?
The selected product image specifies paraffin-embedded human tonsil carcinoma tissue at 1:50 but does not report a fixative (caption: A11662). Target-specific fixation sensitivity is therefore unknown; neither the HPA staining pattern nor BUD31 topology or acetylation establishes one (HPA tissue IHC; UniProt P41223). Record fixative, fixation duration and tissue thickness for each specimen, then compare matched sections using the same retrieval and detection conditions (standard IHC practice). If staining varies across a block, inspect preservation and processing differences before attributing the pattern to BUD31 abundance (standard IHC practice).
Should I accept cytoplasmic BUD31 staining without a nuclear signal?
Treat a nucleus-free cytoplasmic pattern as a finding that needs confirmation: UniProt places BUD31 in the nucleus, including chromatin at promoters of AR target genes (UniProt P41223). HPA nevertheless reports cytoplasmic and nuclear expression across several tissues, with medium consistency between antibody staining and RNA expression (HPA tissue IHC: Approved). Compare nuclear and cytoplasmic staining within the same cell population and check whether the pattern survives an antibody dilution series (standard IHC practice). Document the compartment separately for each cell type; a cytoplasmic signal alone does not establish where BUD31 is functioning (UniProt P41223; standard IHC interpretation).
Can this IHC stain distinguish BUD31 isoforms or a modified epitope?
Do not assign an isoform from chromogenic staining alone: the record lists 2 BUD31 isoforms, but the supplied IHC caption does not map the antibody epitope (UniProt P41223; caption: A11662). The annotated protein spans residues 1–144 and includes an acetyllysine at position 125; those facts do not establish antibody sensitivity to that modification (UniProt P41223). Obtain the antibody's epitope information before claiming isoform selectivity, and check whether the proposed epitope is shared by the relevant sequences (standard assay validation practice). Until then, report staining as BUD31 immunoreactivity and keep compartment and cell-type observations separate (standard IHC reporting practice).
How can I assess BUD31 alongside a cell-type marker by IF/ICC?
For the separate IF/ICC assay, pair BUD31 with a marker validated for the cell type being examined and include single-stain controls to assess channel bleed-through (standard IF practice). Choose a fluorophore channel with low tissue autofluorescence in the specimen, verifying the choice on an unstained section (standard IF practice). BUD31 is a nuclear protein with no annotated transmembrane segment, so permeabilisation must permit antibody access to its intracellular epitope; optimise it against preservation of nuclear structure (UniProt P41223; standard IF practice). Compare nuclear signal with a DNA counterstain, while treating HPA's microtubule and centrosome locations as uncertain (HPA subcellular: supported nucleoplasm; uncertain additional locations).
What should I check when DAB staining obscures BUD31-positive nuclei?
First inspect a section processed without primary antibody to assess detection-system background, and examine an unstained section for pigment that could resemble DAB (standard chromogenic IHC practice). Block endogenous peroxidase before chromogen development, then compare a shorter DAB development with the original run (standard chromogenic IHC practice). Titrate the catalog antibody around the documented 1:50 image condition on matched paraffin sections, because that caption shows a tested image condition rather than a universal dilution (caption: A11662; standard IHC practice). Score signal only where cell boundaries and nuclei remain interpretable; diffuse brown deposit should not be counted as BUD31-positive cells (standard IHC scoring practice).
How should I quantify BUD31 when nuclear and cytoplasmic staining differ? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, because UniProt places BUD31 in the nucleus while HPA tissue IHC describes both nuclear and cytoplasmic expression (UniProt P41223; HPA tissue IHC). For nuclear chromogenic staining, record the percentage of positive nuclei and intensity categories, then calculate an H-score as the sum of each intensity score multiplied by its percentage (standard IHC scoring practice). Normalise positive-cell counts to evaluable nuclei in the same defined region, or report positive-cell density per mm² of evaluable tissue (standard IHC quantification practice). Exclude folds, necrosis and staining edges using the same rules across sections (standard IHC scoring practice).
How do I distinguish convincing BUD31 staining from section artefact?
A convincing result shows reproducible staining in intact cells, with nuclear localisation assessed explicitly because UniProt assigns BUD31 to the nucleus (UniProt P41223; standard IHC interpretation). HPA reports medium staining in several cell populations, including appendix glandular cells, and no detected staining in adrenal gland glandular cells; use those observations as context rather than absolute controls (HPA tissue IHC: Approved). Question signal restricted to cut edges, necrotic regions or damaged cells, and check a no-primary control for endogenous enzyme or detection background (standard chromogenic IHC practice). Record unexpected cell types and cytoplasmic-only staining separately for follow-up, since HPA also describes cytoplasmic expression (HPA tissue IHC).
Boster reagents
Best BUD31 / Protein BUD31 homolog IHC Antibodies
A11662 has real BUD31 IHC data from a paraffin-embedded human tissue sample (catalog image caption); the catalog lists Human, Mouse, and Rat reactivity, with no IF data (catalog applications and images).
Real IHC data
Anti-BUD31 (E24) Antibody
Cat # A11662
A11662 will render with an IHC image of paraffin-embedded human tonsil carcinoma tissue at 1:50 (catalog image caption). Its listed applications are IHC and WB, and its listed reactivity is Human, Mouse, and Rat (catalog applications and reactivity).
Which to pick: Choose A11662 for paraffin-section IHC: its own image caption documents staining in paraffin-embedded human tonsil carcinoma tissue at 1:50; the fixative is unreported (catalog image caption). For Mouse or Rat tissue, A11662 has listed reactivity, but its IHC image documents only a human sample (catalog reactivity and image caption). No catalog antibody here is listed for IF/ICC, so there is no supported IF/ICC pick (catalog applications and images).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P41223 (BUD31_HUMAN, Protein BUD31 homolog).
- Human Protein Atlas. BUD31 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. BUD31 subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the centrosome and microtubules..
- Human Protein Atlas. BUD31 antibody validation summary (1 antibodies).
- Downregulation of BUD31 Promotes Prostate Cancer Cell Proliferation and Migration via Activation of p-AKT and Vimentin In Vitro. International journal of molecular sciences 2023 — PMC10094631.
- The role of BUD31 in clear cell renal cell carcinoma: prognostic significance, alternative splicing, and tumor immune environment. Clinical and experimental medicine 2024 — PMC11322202.
- Splicing factor BUD31 promotes ovarian cancer progression through sustaining the expression of anti-apoptotic BCL2L12. Nature communications 2022 — PMC9587234.
- Single-cell proteo-genomic reveals a comprehensive map of centrosome-associated spliceosome components. iScience 2023 — PMC10214398.
- PubMed PMID:7841202 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:12690205 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for BUD31 (Protein BUD31 homolog). Expected localisation Nuclear (UniProt); nuclear and cytoplasmic in tissue (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen). Validated IHC antibody: A11662 at 1:50-1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Adipose tissue (Adipocytes); negative Adrenal gland. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human tonsil carcinoma tissue; fixative not specified (datasheet A11662); 2. Fixation: Image fixative and duration unreported (datasheet A11662); 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 Tris-EDTA buffer, pH 9.0 , 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-BUD31, 1:50-1:200 (datasheet A11662); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Staining and RNA show medium consistency (HPA tissue IHC); Isoform / epitope: 2 isoforms; epitope coverage is unestablished (UniProt); Regulation: Low tissue RNA specificity (HPA tissue RNA). Evidence: UniProt P41223, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet A11662, PMC10094631, PMC9587234. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/BUD31/immunohistochemistry-ihc-guide.