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BRF1 / Transcription factor IIIB 90 kDa subunit · IHC design guide
Design Immunohistochemistry for BRF1
Plan BRF1 paraffin-section IHC around variable nuclear staining in most tissues (HPA tissue IHC). Compare kidney tubular cells with high staining against adipocytes with no detected staining (HPA tissue IHC), and titrate the catalog antibody within 1:100–1:300 (datasheet).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
BRF1 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 staining (HPA tissue IHC) | |
| Staining pattern | Variable nuclear staining in most tissues (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen) | |
| Positive control | Adrenal gland+4 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 sections (standard IHC practice; not target-specific) | |
| Caveat | Presumed off-target staining was observed (HPA tissue IHC) | |
| Regulation | Expression regulation unreported (UniProt) | |
| Isoform / epitope | 9 isoforms; check epitope coverage (UniProt) |
Section 1
Recommended BRF1 IHC & IF Protocols
The catalog antibody protocol is paired with four published BRF1 IHC examples (PMC8081602; PMC4868753; PMC7336232; PMC11369479).
| Sample | Paraffin-embedded human brain tissue; fixative not specified (datasheet A03761-1) |
| Fixation | Image fixative and duration unreported (datasheet A03761-1); 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 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 monoclonal (clone 2F1) anti-BRF1, 1:100-1:300 (datasheet A03761-1) |
| 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 | BRF1-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Nuclear expression of varying levels in most tissues. No signal in the no-primary control. |
💡Decision noteStart with the page’s Tris-EDTA pH 9.0 retrieval for this nuclear target (UniProt Q92994; page retrieval rule); compare citrate retrieval where reported (PMC4868753; PMC7336232; PMC11369479).
Section 2
What Is the Expected BRF1 Staining Pattern?
BRF1 should show predominantly nuclear staining in paraffin tissue sections (UniProt Q92994: nucleus; HPA: nuclear expression at varying levels in most tissues). High staining is reported in kidney tubular cells, cerebral cortex neurons, and testis spermatogonia (HPA: High). A membrane pattern is unexpected (UniProt Q92994: no transmembrane segment). HPA rates its tissue IHC evidence Approved, while noting presumed off-target binding was disregarded and external verification is pending.
| Distinct nuclear staining in kidney tubular cells, with nuclei in the same cell population staining more strongly than surrounding cytoplasm. | This fits BRF1 localisation and a reported high-staining cell population (UniProt Q92994: nucleus; HPA: High in kidney tubular cells). Judge the nuclear pattern within identified tubules; staining elsewhere in the section needs its own cell-type assessment. |
| Strong cytoplasmic or membrane staining dominates while nuclei remain pale. | This conflicts with the expected compartment (UniProt Q92994: nucleus; HPA: nuclear expression). Treat the pattern as suspect and check controls and detection conditions before scoring it as BRF1; HPA notes that presumed off-target binding was disregarded in its tissue assessment (HPA: Approved reliability description). |
| Prominent staining appears in adipocytes or prostate glandular cells, especially without convincing nuclear staining in an expected positive population. | Those named cell populations were reported as not detected (HPA: adipocytes, Not detected; prostate glandular cells, Not detected). Investigate nonspecific antibody binding or endogenous detection activity (standard IHC practice). The HPA observations do not establish that every cell in either whole tissue must be negative. |
| A uniform haze covers nuclei, cytoplasm, and extracellular spaces, obscuring cell boundaries. | Diffuse background prevents a reliable compartment call (standard IHC practice). Assess blocking, washing, antibody concentration, and detection controls (standard IHC practice); the supplied UniProt and HPA records do not identify a BRF1-specific cause of background. |
| No nuclear signal is visible in an included high-staining cell population. | A blank kidney tubular, neuronal, or spermatogonial population is discordant with the corresponding HPA High observations (HPA: kidney tubules, cerebral cortex neurons, testis spermatogonia). Check that the expected cells are present, then review assay controls and workflow (standard IHC practice) before interpreting the specimen as negative. |
💡Expected BRF1 appearanceCall a result positive when identifiable expected cells show nuclear chromogenic signal, potentially strong in kidney tubules, cerebral cortex neurons, or spermatogonia (UniProt Q92994: nucleus; HPA: High in those cells); dominant membrane staining or signal confined to reported not-detected cell populations is suspect (UniProt Q92994: no transmembrane segment; HPA: Not detected).
| Cell type and baseline abundance | Compare like cells with like cells: HPA reports high staining in thyroid glandular cells and medium staining in appendix glandular cells, alongside low staining in stomach glandular cells (HPA: tissue IHC). A weak result in a low-staining population carries less weight than a blank high-staining population. |
| IHC antibody evidence | The listed tissue-IHC antibody has Approved status (HPA: HPA074990 IHC Approved). HPA also reports presumed off-target binding was disregarded and external verification is pending (HPA: reliability description). Interpret unexpected staining against cell type and compartment rather than treating Approved as proof of specificity in every specimen. |
| Antigen retrieval | Retrieval conditions can be evaluated with an expected positive tissue and detection controls when nuclear staining is absent (standard IHC practice). No BRF1-specific retrieval condition or fixation sensitivity is established by the supplied UniProt and HPA evidence; a failed signal alone cannot identify epitope masking as its cause. |
| Isoforms and epitope coverage | UniProt lists nine BRF1 isoforms and distinct transcriptional roles for isoforms 1 and 2 (UniProt Q92994). The payload gives no antibody epitope or isoform-coverage data, so staining cannot be assigned to one isoform, and isoform differences cannot explain a particular negative slide from these sources alone. |
| IF/ICC Q&A: Where should BRF1 fluorescence appear? | HPA places BRF1 in the nucleoplasm and nuclear bodies, both Approved (HPA: subcellular ICC-IF). That supports a nuclear IF/ICC interpretation; the separately listed ICC antibody is HPA051918, ICC Approved (HPA: antibodies). These observations do not supply an IF/ICC protocol for this IHC section. |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected high-staining nuclei are blank. | The assay may have failed, the expected cell population may be absent, or detection may be inadequate (standard IHC practice); HPA alone cannot distinguish these causes. | Confirm the named cells on the counterstained section, inspect a known positive control, and review retrieval and detection steps (standard IHC practice; HPA: High in kidney tubular cells). |
| Signal is mainly cytoplasmic or membranous. | The pattern disagrees with the reported nuclear location (UniProt Q92994: nucleus; HPA: nuclear tissue expression). Its precise cause is unproven. | Check a positive tissue and a detection control, then reassess whether any reproducible nuclear signal remains (standard IHC practice). Do not score dominant membrane staining as the expected BRF1 pattern (UniProt Q92994: no transmembrane segment). |
| Reported not-detected cells stain strongly. | Nonspecific binding or endogenous detection activity is possible (standard IHC practice); HPA has also noted presumed off-target binding in its tissue assessment (HPA: reliability description). | Identify the stained cell type, compare it with HPA's cell-level result, and inspect antibody-omission and detection controls (standard IHC practice; HPA: adipocytes and prostate glandular cells, Not detected). |
| Brown haze is widespread and nuclei cannot be distinguished. | High background can obscure a real nuclear pattern (standard IHC practice); the supplied sources do not establish a BRF1-specific mechanism. | Review blocking, washes, antibody concentration, and chromogenic detection controls (standard IHC practice). Reassess only when nuclear staining can be separated from the haze (UniProt Q92994: nucleus). |
| Staining is weak in a low-staining population. | Low signal may match the reported baseline; HPA lists stomach and small-intestine glandular cells as Low (HPA: tissue IHC). | Compare the same cell type and use a high-staining control population to judge assay performance (standard IHC practice; HPA: kidney tubular cells, High). Avoid calling the run failed solely from a weak low-staining population. |
| A positive tissue stains, but the study tissue is negative. | BRF1 staining varies among tissues and cell types (HPA: nuclear expression at varying levels in most tissues); the supplied evidence gives no diagnosis-specific expectation for the study specimen. | Score the identified cell population and nuclear signal with the positive control in view (standard IHC practice; UniProt Q92994: nucleus). Report the study result as observed, without inferring an isoform or fixation effect from absence alone. |
Experiment evidence
IHC & IF images showing BRF1 staining
A03761-1 · IHCImmunohistochemistry analysis of paraffin-embedded human brain tissue, using TF3B Antibody. The picture on the right is blocked with the syn
A03761-1 · IFImmunofluorescence analysis of MCF7 cells, using TF3B Antibody. The picture on the right is blocked with the synthesized peptide.
A30544 · IHCImmunohistochemistry analysis of paraffin-embedded human brain tissue, using TF3B Antibody. The picture on the right is blocked with the syn
A30544 · IFImmunofluorescence analysis of MCF7 cells, using TF3B Antibody. The picture on the right is blocked with the synthesized peptide.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000185024-BRF1/tissue/cerebral+cortex#imid_20575955
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000185024-BRF1/tissue/colon#imid_20575881
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000185024-BRF1/tissue/liver#imid_20575882
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000185024-BRF1/tissue/kidney#imid_20575875
HPA · ICC-IF MCF-7Human Protein Atlas immunofluorescence, MCF-7, antibody HPA051918; green = BRF1, blue = nucleus, red = microtubules.
HPA · ICC-IF MCF-7Human Protein Atlas immunofluorescence, MCF-7, antibody HPA051918; green = BRF1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for BRF1 IHC & IF
🧪Run cerebral cortex first and expect staining in neuronal cells (HPA: High in cerebral cortex neuronal cells; A03761-1 caption: staining in paraffin-embedded human brain). Use parathyroid gland as the negative tissue and expect its glandular cells to lack staining (HPA: Not detected in parathyroid gland glandular cells); on the cortex slide, cells without nuclear signal provide an internal background check, but the supplied HPA rows do not establish a specific cortical cell type as BRF1-negative (UniProt Q92994: nucleus; HPA: cerebral cortex neuronal cells High).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show BRF1 in MCF-7, PC-3, U2OS, NIH 3T3, with annotated localisation: Nucleoplasm (approved), Nuclear bodies (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and an isotype control matched to the primary antibody's host species and clonality (standard IHC control practice); use peptide blocking as a biological specificity control, as shown for the selected antibody (A03761-1 caption: signal blocked with synthesized peptide). For chromogenic detection in brain, quench endogenous peroxidase and check the no-primary slide for residual signal (standard IHC practice).
⚠️Feasibility: The selected antibody has a paraffin-section brain IHC image, but its caption does not report the fixative (A03761-1 caption: paraffin-embedded human brain; fixative unreported). A BRF1-specific fixation window and antigen-retrieval dependency are unreported in the supplied evidence, so optimize retrieval on control sections (supplied target/application evidence). The evidence does not establish whether frozen sections or IF are easier; if adapting the assay to IF in brain, assess tissue autofluorescence with a no-primary control (standard IF practice).
HPA tissue IHC evidence for BRF1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Presumed off target binding observed and disregarded. 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 → |
| Cerebral cortex | Neuronal cells | High | Protein (IHC) | HPA → |
| Endometrium | Cells in endometrial stroma | High | Protein (IHC) | HPA → |
| Kidney | Cells in tubules | High | Protein (IHC) | HPA → |
| Testis | Spermatogonia cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Liver | Cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Ovary | Ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Parathyroid gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Prostate | Glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced BRF1 IHC Tips
Troubleshoot BRF1 staining in paraffin sections by checking nuclear localisation, tissue controls and the limits of antibody specific evidence (UniProt Q92994; HPA tissue IHC).
What should I change when BRF1 nuclear staining is weak after retrieval?
Start with Tris-EDTA at pH 9.0 for heat-induced retrieval at 95–98 °C for 20 min (page retrieval setting: nuclear antigen). If nuclei remain weak, confirm the bath reaches the stated temperature and that sections stay covered by buffer throughout heating (standard IHC practice). Compare a short retrieval time series on adjacent sections while keeping antibody concentration and chromogen development constant (standard IHC practice). Judge improvement by nuclear signal in an expected positive cell population, such as cerebral cortex neurons, alongside background in the same section (HPA: High in cerebral cortex neuronal cells; UniProt Q92994: nucleus).
Could fixation explain weak BRF1 staining in paraffin sections?
BRF1-specific sensitivity to fixation is unknown from the supplied evidence; the catalog image identifies paraffin-embedded human brain tissue but does not state its fixative (caption: A03761-1). Record the actual fixative, fixation duration and processing history before comparing staining across blocks (standard IHC practice). If a well-characterised positive block stains and the test block does not, compare adjacent sections using the same pH 9.0, 95–98 °C, 20 min retrieval and detection run (page retrieval setting; standard IHC practice). Treat any difference as a processing-associated observation until controls distinguish fixation from antigen abundance or tissue condition (standard IHC practice).
How should I assess BRF1 staining outside the nucleus?
Score convincing BRF1 staining in nuclei, with nucleoplasm and nuclear bodies as more specific expected patterns (UniProt Q92994: nucleus; HPA subcellular: approved nucleoplasm and nuclear bodies). Diffuse cytoplasmic or membrane staining should prompt review of background and antibody specificity before it is called BRF1 (UniProt Q92994: no transmembrane segment; standard IHC practice). Inspect a matching negative reagent control and compare staining in intact cells away from tissue edges (standard IHC practice). Use the peptide-blocked companion image as product-specific supporting evidence, while recognising that peptide competition alone cannot establish every stained structure as specific (caption: A03761-1; standard IHC practice).
Can this IHC stain distinguish BRF1 isoforms or phosphorylation states?
The record lists 9 BRF1 isoforms, but the supplied IHC caption does not define the antibody epitope or its isoform coverage (UniProt Q92994: isoforms 1–9; caption: A03761-1). Do not assign an isoform from nuclear chromogenic staining alone (standard IHC interpretation). Isoform 1 participates in transcription of tRNA, VA1, 7SL and 5S RNA, whereas isoform 2 is required for U6 promoter transcription; those roles do not identify which isoform a stained nucleus contains (UniProt Q92994: function). Phosphorylation is annotated at residues 365, 450 and 553, but phosphorylation-specific recognition requires direct epitope evidence (UniProt Q92994: modified residues; standard IHC interpretation).
How can I check a BRF1 IF signal against the IHC pattern?
Use this as a cross-check of localisation: BRF1 should appear in nuclei, including nucleoplasm and nuclear bodies, when the IF antibody is suitable (UniProt Q92994: nucleus; HPA subcellular: approved locations). In a multiplex experiment, pair BRF1 with a validated marker for the expected cell population, such as cerebral cortex neurons, and assign fluorophores after checking tissue autofluorescence in unstained sections (HPA: High in cerebral cortex neuronal cells; standard IF practice). Because BRF1 is nuclear and has no transmembrane segment, permeabilise fixed cells sufficiently to expose a nuclear epitope (UniProt Q92994: localisation and topology; standard IF practice). Confirm bleed-through with single-colour controls before comparing IF and chromogenic nuclear patterns (standard IF practice).
What controls help separate BRF1 signal from chromogenic background?
First compare stained sections with a negative reagent control and a positive tissue control processed in the same run (standard IHC practice). Quench endogenous peroxidase before an HRP and DAB detection sequence, and check whether pigment or tissue edges retain colour independently of primary antibody (standard chromogenic IHC practice). If background is widespread, review blocking, washes, primary-antibody concentration and DAB development time one variable at a time (standard IHC practice). The A03761-1 brain image includes a peptide-blocked companion, which can support assessment of antibody-dependent staining but does not replace localisation and tissue controls (caption: A03761-1; UniProt Q92994: nucleus).
How should I quantify BRF1 staining across tissue samples? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and count intact nuclei within a consistent tissue area before scoring chromogenic intensity (standard IHC quantification; UniProt Q92994: nucleus). Report the percentage of positive nuclei and, if intensity matters, an H-score calculated as the summed percentage at each intensity multiplied by its intensity grade (standard IHC quantification). Alternatively, report positive nuclear density per mm² of evaluable tissue, with the area and exclusion rules stated (standard IHC quantification). Normalise comparisons to the same cell type, section quality, detection run and scoring threshold; avoid pooling populations with different baseline staining (standard IHC quantification; HPA: varying nuclear levels across tissues).
When is a BRF1-positive section biologically convincing?
A convincing result shows reproducible nuclear staining in intact cells and agrees with the expected nucleoplasm or nuclear-body pattern (UniProt Q92994: nucleus; HPA subcellular: approved locations). Cerebral cortex neuronal cells provide an example of a population reported High, whereas adipocytes are reported Not detected; use these as context, not absolute proof for an individual section (HPA tissue IHC). Question cytoplasmic-only signal, staining concentrated at cut edges or necrotic regions, and colour persisting in a negative reagent control (UniProt Q92994: nucleus; standard IHC practice). Check endogenous enzyme activity when using peroxidase detection, and remember HPA reports presumed off-target binding with external verification pending (standard chromogenic IHC practice; HPA: Approved reliability description).
Boster reagents
Best BRF1 / Transcription factor IIIB 90 kDa subunit IHC Antibodies
Two anti-BRF1 antibodies have IHC images of paraffin-embedded human brain and IF images of MCF7 cells (catalog image captions); both list human and mouse reactivity (catalog).
Real IHC data
Anti-TFIIIB90-1 Antibody
Cat # A03761-1
Real IHC data
Anti-TF3B BRF1 Antibody
Cat # A30544
A03761-1 lists IHC and IF, with its own images showing paraffin-embedded human brain IHC and MCF7 IF (A03761-1 catalog/image captions). A30544 lists IHC, IF and ICC, with its own images showing paraffin-embedded human brain IHC and MCF7 IF (A30544 catalog/image captions).
Which to pick: For tissue IHC, choose A03761-1 if a defined rabbit clone is useful: clone 2F1 has a listed IHC dilution of 1:100–1:300, and its own image shows a paraffin-embedded human brain section; the fixative is unreported (A03761-1 catalog/IHC caption). For IF/ICC, choose polyclonal A30544 because both applications are listed and its own IF image shows MCF7 cells; its own IHC image also shows paraffin-embedded human brain, with fixative unreported (A30544 catalog/IF and IHC captions). For cross-species planning, either SKU lists human and mouse reactivity, although both supplied IHC images show human tissue (A03761-1 and A30544 catalogs/IHC captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q92994 (TF3B_HUMAN, Transcription factor IIIB 90 kDa subunit).
- Human Protein Atlas. BRF1 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. BRF1 subcellular location (ICC-IF): Localized to the nucleoplasm and nuclear bodies..
- Human Protein Atlas. BRF1 antibody validation summary (2 antibodies).
- Exploring the Role and Mechanism of pAMPKα-Mediated Dysregulation of Brf1 and RNA Pol III Genes. Oxidative medicine and cellular longevity 2021 — PMC8081602.
- The significance of Brf1 overexpression in human hepatocellular carcinoma. Oncotarget 2016 — PMC4868753.
- Mitogen- and Stress-Activated Protein Kinase 1 Mediates Alcohol-Upregulated Transcription of Brf1 and tRNA Genes to Cause Phenotypic Alteration. Oxidative medicine and cellular longevity 2020 — PMC7336232.
- BRF1 promotes the odontogenic differentiation of dental pulp stem cells in pulpitis by inducing autophagy. Heliyon 2024 — PMC11369479.
- PubMed PMID:7624363 — UniProt-cited evidence.
- PubMed PMID:8943358 — UniProt-cited evidence.
- PubMed PMID:10921893 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for BRF1 (Transcription factor IIIB 90 kDa subunit). Expected localisation Nuclear staining (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen). Validated IHC antibody: A03761-1 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human brain tissue; fixative not specified (datasheet A03761-1); 2. Fixation: Image fixative and duration unreported (datasheet A03761-1); 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 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 monoclonal (clone 2F1) anti-BRF1, 1:100-1:300 (datasheet A03761-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Caveat: Presumed off-target staining was observed (HPA tissue IHC); Isoform / epitope: 9 isoforms; check epitope coverage (UniProt); Regulation: Expression regulation unreported (UniProt). Evidence: UniProt Q92994, HPA tissue IHC · Approved, HPA ICC-IF · nucleoplasm and nuclear bodies., Datasheet A03761-1, PMC8081602, PMC4868753. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/BRF1/immunohistochemistry-ihc-guide.