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- Table of Contents
BRF1 / Transcription factor IIIB 90 kDa subunit · Western blot design guide
Design a Western Blot for BRF1
Source-linked BRF1 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-BRF1 WB antibodies. Everything you need to plan the experiment before you commit precious samples.
Evidence assembled September 2026 · For research use; verify linked source records and product datasheet before use
BRF1 Western Blot Experimental Design Guide
Expected bands, source-linked protocol options, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before running
| Expected band | ~73.8 kDa | |
| Gel | 12–15% (standard starting point) | |
| Positive control | Adrenal gland (IHC candidate; verify WB) +4 more | |
| Negative control | Adipose tissue (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 9 isoform(s) |
Section 1
Source-Linked BRF1 Western Blot Protocol Options
The A03761-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | various cells (catalog A03761-1) |
| Gel % | 12–15% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Standard semi-dry transfer; verify efficiency (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | A03761-1; use the WB datasheet starting dilution (standard starting point) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | Species-matched HRP conjugate at validated dilution (standard starting point) |
| Secondary incubation | 1 h at room temperature (standard starting point) |
| Wash | 3 × 5 min in TBST (standard starting point) |
| Detection | ECL; bracket exposures to avoid saturation (standard starting point) |
Section 2
What Is the Expected BRF1 Western Blot Band Size?
BRF1 has a predicted mass of 73.8 kDa; isoforms could affect band patterns, but their migration has not been demonstrated here.
| Band near 73.8 kDa | Consistent with the predicted mass of BRF1; identity requires controls |
| Additional band above 73.8 kDa | Could reflect a BRF1 isoform; its mass is not supplied |
| Additional band below 73.8 kDa | Could reflect a BRF1 isoform; its mass is not supplied |
| Several bands at different positions | Could reflect isoforms 1–9; their separation is not established |
💡Expected BRF1 appearanceUniProt predicts 73.8 kDa for BRF1; no empirical band size is supplied, so confirm any candidate band with antibody specificity and nuclear fraction controls.
| UniProt predicted mass | Provides a 73.8 kDa reference, not a measured band position |
| Splice isoforms 1, 2 and 3 | May differ in size; individual masses and migration are unknown |
| Splice isoforms 4, 5 and 6 | May differ in size; individual masses and migration are unknown |
| Splice isoforms 7, 8 and 9 | May differ in size; individual masses and migration are unknown |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Nuclear BRF1 may be diluted in whole-cell lysate | Check a nuclear fraction and loading controls |
| Band higher than expected | A splice isoform is possible, but its migration is unknown | Verify identity with an independent BRF1 antibody or depletion control |
| Band lower than expected | A splice isoform is possible, but its migration is unknown | Verify identity with an independent BRF1 antibody or depletion control |
| Multiple bands | BRF1 has nine named splice isoforms, without supplied band positions | Test band identity with BRF1 depletion and an independent antibody |
| Weak or no signal | Nuclear BRF1 may be underrepresented in the sample | Enrich the nuclear fraction and check antibody and loading controls |
Experiment evidence
Western blot images showing BRF1 band
A03761-1 · BRF1
A03761-1 · BRF1
A30544 · BRF1
A30544 · BRF1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for BRF1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for BRF1 in Western blot, you can use adrenal gland tissue, which HPA reports as high expression.
Positive control: Adrenal gland (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: BRF1 is nuclear, so nuclear-enriched lysate may improve detection.
HPA tissue expression evidence for BRF1
Comprehensive Human Protein Atlas IHC scoring per tissue. Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Higher expression tissues · candidate positive controls from IHC
| 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 → |
Lower expression tissues · IHC evidence, not confirmed WB-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 Western Blot Tips
Deeper troubleshooting and optimisation questions for BRF1, answered from its protein features.
How should BRF1 band migration be interpreted?
Band shift · Use the separately labelled calculated mass and catalog-observed evidence above. A sequence annotation does not establish an observed migration shift. Verify target identity with orthogonal controls.
Which BRF1 isoforms could produce smaller bands?
Isoforms · UniProt lists nine isoforms. Isoforms 2, 4, and 6 lack large C-terminal regions; isoforms 7–9 have N-terminal deletions. These sequence differences could affect band size. Check which isoforms your antibody recognizes before assigning a band.
Match the antibody epitope to each isoform’s sequence. For example, isoforms 2 and 3 lack residues 1–204, while isoform 9 lacks residues 1–238. Isoforms 2, 4, and 6 also lack substantial C-terminal sequence. An epitope within a missing region cannot detect that isoform.
Match the antibody epitope to each isoform’s sequence. For example, isoforms 2 and 3 lack residues 1–204, while isoform 9 lacks residues 1–238. Isoforms 2, 4, and 6 also lack substantial C-terminal sequence. An epitope within a missing region cannot detect that isoform.
Which phosphorylation sites should I consider when interpreting BRF1 bands?
PTM · UniProt lists phosphothreonine 365 and phosphoserines 450 and 553, using the supplied canonical sequence coordinates. These sites do not establish that phosphorylation produces a resolvable band shift. Isoforms 2, 4, and 6 lack the C-terminal regions containing all three sites.
Does this guide establish induction of BRF1?
Induction · No general induction response is established by this guide. A pathway or gene-set association is not evidence of induction in a particular specimen. Verify the relevant treatment and control in a target-specific experiment.
How should transfer be checked for BRF1?
Transfer · Standard workflow guidance: verify transfer efficiency for the intended target size before interpreting a weak signal. Use total-protein assessment and optimize transfer for the membrane, gel and apparatus; the labelled catalog values take precedence.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A03761-1 datasheet where specified. Otherwise compare 5% milk or 5% BSA in TBST; for a phospho-specific assay start with BSA. Optimize background and specific signal with matched controls.
How should I quantify multiple BRF1 bands?
Quantitation · Define which isoforms the antibody can detect, then quantify the corresponding bands consistently across samples. UniProt lists nine isoforms with sequence differences, so combining bands without checking epitope coverage could mix distinct forms. The features alone do not establish which bands are present in a sample.
Why might BRF1 differ from its predicted mass?
Interpretation · The canonical sequence has a predicted mass of 73.8 kDa, despite the protein name ‘90 kDa subunit.’ No observed band mass is supplied. The listed phosphorylation sites and alternative isoforms are possibilities to consider, but their presence alone does not establish a visible shift or explain a measured difference.
Compare its apparent size with the canonical 73.8 kDa prediction and the listed isoform deletions or replacements. Check whether the antibody epitope is retained. The three phosphorylation sites are additional context, but the supplied features do not identify an observed band or prove its cause.
UniProt places BRF1 in the nucleus. A nuclear fraction is therefore relevant when assessing BRF1 detection; compare fractions consistently when measuring abundance. This location does not, by itself, identify a particular Western-blot band.
Compare its apparent size with the canonical 73.8 kDa prediction and the listed isoform deletions or replacements. Check whether the antibody epitope is retained. The three phosphorylation sites are additional context, but the supplied features do not identify an observed band or prove its cause.
UniProt places BRF1 in the nucleus. A nuclear fraction is therefore relevant when assessing BRF1 detection; compare fractions consistently when measuring abundance. This location does not, by itself, identify a particular Western-blot band.
Boster reagents
BRF1 Western Blot Antibodies
Catalog antibodies with Western blot application and product-specific WB images. Evaluate suitability with the reported sample, controls and experimental conditions.
Real WB data
Anti-TFIIIB90-1 Antibody
Cat # A03761-1
Real WB data
Anti-TF3B BRF1 Antibody
Cat # A30544
Two the supplier antibodies are listed for BRF1, both with stated Human and Mouse reactivity and WB images. The A03761-1 caption describes nuclear extracts from unspecified cells. The A30544 caption names TFIIIB90-1 Polyclonal Antibody, leaving its image attribution unclear.
Which to pick: For a nuclear-extract WB, start with A03761-1 because its caption reports that preparation. A30544 has the same stated species reactivity, but confirm that its WB image represents A30544 before relying on it.
Source: BosterBio BRF1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q92994.
- Human Protein Atlas. BRF1 tissue expression.
- PMC11414613 — target-verified WB comparison
- PMC8081602 — target-verified WB comparison
- PMC5709663 — target-verified WB comparison