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- Table of Contents
EBF1 / Transcription factor COE1 · Western blot design guide
Design a Western Blot for EBF1
Real validated EBF1 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-EBF1 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
EBF1 Western Blot Experimental Design Guide
Expected bands, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before running
| Expected band | ~64.5 kDa | |
| Gel | 12–15% (standard starting point) | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Acetylated + Ubl conjugation | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 2 isoform(s) |
Section 1
Real Curated EBF1 Western Blot Protocols
The M01341 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | Raji cell lysate (catalog M01341) |
| 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 | M01341; 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 EBF1 Western Blot Band Size?
EBF1 is predicted at 64.5 kDa; its isoforms and homodimer could affect migration, but no empirical band size or visible shift is established.
| Band near 64.5 kDa | consistent with the predicted EBF1 monomer size; confirm identity with controls |
| Band near 129 kDa under nonreducing conditions | could represent an EBF1 homodimer if it survives electrophoresis |
| Additional bands at unspecified positions | could reflect isoforms 1 and 2, whose migration is not established |
| Faint lysate band with stronger nuclear fraction signal | consistent with nuclear localization of EBF1 |
💡Expected EBF1 appearanceEBF1 has a predicted mass of 64.5 kDa, but no empirical band size is supplied; confirm any candidate band with antibody and depletion controls.
| Predicted EBF1 mass | places the monomer near 64.5 kDa before any unverified migration effects |
| Homodimer formation | could yield a band near twice the monomer size if the complex survives electrophoresis |
| Isoform 1 | may migrate differently from isoform 2; the direction is unknown |
| Isoform 2 | may migrate differently from isoform 1; the direction is unknown |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | nuclear EBF1 may be poorly recovered | check a nuclear fraction and validate recovery with a nuclear marker |
| Band higher than expected | an EBF1 homodimer may persist | compare reducing and nonreducing samples and confirm identity by depletion |
| Band lower than expected | an alternative isoform is possible, but its migration is unknown | compare isoform expression and test whether the band falls with EBF1 depletion |
| Multiple bands | isoforms 1 and 2 may contribute, though distinct bands are unproven | use EBF1 depletion and isoform-specific controls to assign bands |
| Weak or no signal | nuclear EBF1 may be diluted in whole-cell lysate | enrich nuclei and verify antibody performance with a positive control |
Experiment evidence
Western blot images showing EBF1 band
M01341 · EBF1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for EBF1 Western blot
🧪For positive controls for EBF1 in Western blot, you can use an HPA-IHC candidate positive sample once one is identified.
Positive control: No high/medium HPA tissue identified
Negative control: Suggested KO / knockdown lysate
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: HPA provides no positive or negative tissue candidates, so tissue controls cannot be selected from these data.
HPA tissue expression evidence for EBF1
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 |
|---|---|---|---|---|
| No high/medium HPA tissues identified in the supplied evidence. | ||||
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| No lower-expression tissue rows available in the supplied evidence. | ||||
Section 3
Advanced EBF1 Western Blot Tips
Deeper troubleshooting and optimisation questions for EBF1, answered from its protein features.
How should EBF1 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.
Can EBF1 isoforms produce different bands?
Isoforms · UniProt lists two isoforms. Relative to isoform 1, isoform 2 lacks residues 162–184 and 252–259 in UniProt numbering. These deletions could affect apparent size, but the features do not establish whether the isoforms resolve as separate bands. Check which isoform your antibody recognizes.
Which EBF1 modification matters when interpreting a band?
PTM · UniProt lists N-acetylmethionine at position 1. Keep that UniProt coordinate explicit when comparing other numbering schemes. Its presence does not establish a visible band shift. The supplied features list no glycosylation sites.
Does this guide establish induction of EBF1?
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 EBF1?
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 M01341 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 EBF1 Western blot signals be quantified?
Quantitation · Compare signals from consistently prepared samples and normalize for loading in the fraction analyzed. Because EBF1 is nuclear, changes in nuclear signal should be interpreted as changes in that fraction unless whole-cell abundance is measured separately.
Should EBF1 appear exactly at its predicted 64.5 kDa?
Interpretation · 64.5 kDa is the predicted mass; no observed Western blot band is supplied. Use it as a reference, not an exact migration requirement. The listed features alone cannot establish a visible shift or explain a difference between apparent and predicted mass.
EBF1 is listed in the nucleus. A nuclear fraction is therefore relevant when assessing its signal; compare the same fraction across samples and check fraction loading when interpreting weak or variable bands.
Consider the two listed isoforms, including isoform 2's missing segments, when evaluating band size. EBF1 is also reported to form homodimers, but that feature alone does not show that dimers survive Western blot preparation. Do not assign an unexpected band to an isoform, modification, or dimer from migration alone.
EBF1 is listed in the nucleus. A nuclear fraction is therefore relevant when assessing its signal; compare the same fraction across samples and check fraction loading when interpreting weak or variable bands.
Consider the two listed isoforms, including isoform 2's missing segments, when evaluating band size. EBF1 is also reported to form homodimers, but that feature alone does not show that dimers survive Western blot preparation. Do not assign an unexpected band to an isoform, modification, or dimer from migration alone.
Boster reagents
EBF1 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-EBF1 Rabbit Monoclonal Antibody
Cat # M01341
The catalog reports one anti-EBF1 antibody, M01341, a rabbit monoclonal with reported Human and Mouse reactivity. Its Western blot image shows EBF1 expression in Raji cell lysate; that image documents this sample context only.
Which to pick: M01341 is the only listed option. Consider its reported Human and Mouse reactivity for your sample, and use the Raji cell lysate Western blot image as the available example of WB performance.
Source: BosterBio EBF1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q9UH73.
- Human Protein Atlas. EBF1 tissue expression.
- PMC3870187 — target-verified WB comparison
- PMC2657874 — target-verified WB comparison
- PMC9753996 — target-verified WB comparison