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- Table of Contents
FBXW7 / F-box/WD repeat-containing protein 7 · Western blot design guide
Design a Western Blot for FBXW7
Real validated FBXW7 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-FBXW7 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
FBXW7 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 | ~79.7 kDa | |
| Observed band | ~75 kDa | |
| Gel | 8% (catalog A00406-2) | |
| Positive control | Adipose tissue (IHC candidate; verify WB) +4 more | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Ubl conjugation | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 3 isoform(s) |
Section 1
Real Curated FBXW7 Western Blot Protocols
The A00406-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human MCF-7, human 293T, human Jurkat (catalog A00406-2) |
| Gel % | 8% (catalog A00406-2) |
| Load | 30 ug; reducing conditions (catalog A00406-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A00406-2) |
| Membrane | nitrocellulose membrane (catalog A00406-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A00406-2) |
| Primary antibody | A00406-2 · 0.5 μg/mL (catalog A00406-2) |
| Primary incubation | overnight at 4°C (catalog A00406-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A00406-2) |
| Secondary incubation | 1.5 hour at RT (catalog A00406-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A00406-2) |
| Detection | ECL (catalog A00406-2) |
Section 2
What Is the Expected FBXW7 Western Blot Band Size?
FBXW7 is predicted at 79.7 kDa and observed near 75 kDa; the cause of the difference is not established.
| Band near 75 kDa | Empirical FBXW7 band; confirm identity with antibody and depletion controls |
| Band near 79.7 kDa | Near the UniProt predicted mass; migration at this position is not established by the supplied blot |
| Higher band near twice the monomer size | Could reflect an FBXW7 homodimer if it survives electrophoresis |
| Additional bands at different positions | Could reflect isoforms 1, 2, and 3, but distinct migration is unconfirmed |
| Subtle doublet | Could reflect phosphorylation, but a visible shift is unconfirmed |
💡Expected FBXW7 appearanceUniProt predicts 79.7 kDa, while the supplied Western blot reports approximately 75 kDa; confirm band identity with antibody and FBXW7 depletion controls before assigning the difference to a specific cause.
| UniProt predicted mass | 79.7 kDa predicted; the supplied blot reports approximately 75 kDa |
| Homodimer formation | could yield a higher band if the dimer survives electrophoresis |
| Isoforms 1, 2, and 3 | may differ in apparent size; their relative sizes and migration are not supplied |
| Phosphoserine 26 by ATM | could alter migration, but a visible size shift is unconfirmed |
| Phosphothreonine 205 | could alter migration, but a visible size shift is unconfirmed |
| Phosphoserine 227 by SGK1 | could alter migration, but a visible size shift is unconfirmed |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Nuclear FBXW7 may be insufficiently represented in the sample | Check nuclear enrichment and use a positive control lysate |
| Band higher than expected | FBXW7 homodimer may persist during electrophoresis | Compare denaturing conditions and verify identity by FBXW7 depletion |
| Band lower than expected | The approximately 75 kDa empirical band is below the 79.7 kDa prediction; the cause is unknown | Compare with the reported band and verify identity by FBXW7 depletion |
| Multiple bands | Isoforms or phosphorylation are possible, but distinct migration is unconfirmed | Check antibody specificity and compare bands after FBXW7 depletion |
| Weak or no signal | Nuclear FBXW7 may be underrepresented or antibody detection may be weak | Check sample loading, nuclear enrichment, and a positive control lysate |
Experiment evidence
Western blot images showing FBXW7 band
A00406-2 · FBXW7
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for FBXW7 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for FBXW7 in Western blot, you can use adipose tissue lysate, which HPA rates as high expression.
Positive control: Adipose tissue (IHC candidate; verify WB)
Negative control: Suggested KO / knockdown lysate
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: HPA lists no tissue with undetected expression, so use siRNA knockdown or a KO line as the negative control.
HPA tissue expression evidence for FBXW7
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 |
|---|---|---|---|---|
| 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 | adipocytes | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Cerebellum | cells in granular layer | Medium | Protein (IHC) | HPA → |
| Cervix | glandular cells | Medium | Protein (IHC) | HPA → |
| Endometrium | cells in endometrial stroma | Medium | Protein (IHC) | HPA → |
| Epididymis | glandular cells | Medium | Protein (IHC) | HPA → |
| Fallopian tube | glandular cells | Medium | Protein (IHC) | HPA → |
Section 3
Advanced FBXW7 Western Blot Tips
Deeper troubleshooting and optimisation questions for FBXW7, answered from its protein features.
How should FBXW7 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.
Could FBXW7 isoforms produce different bands?
Isoforms · Yes. UniProt lists three isoforms. Relative to isoform 1, isoform 2 lacks residues 1–80 and replaces 81–166; isoform 3 lacks 1–118 and replaces 119–167. Their distinct sequences may affect migration, but the features do not specify observed band positions.
Check the antibody epitope against each isoform’s sequence. An epitope within canonical residues 1–118 is absent from isoform 3; residues 1–80 are absent from isoform 2. Portions of the remaining N-terminal region are replaced in both variants, so an isoform 1 antibody may miss them.
Check the antibody epitope against each isoform’s sequence. An epitope within canonical residues 1–118 is absent from isoform 3; residues 1–80 are absent from isoform 2. Portions of the remaining N-terminal region are replaced in both variants, so an isoform 1 antibody may miss them.
Which FBXW7 phosphorylation sites matter when interpreting bands?
PTM · UniProt lists phosphoserine 26 by ATM, phosphothreonine 205, and phosphoserine 227 by SGK1, using canonical-sequence coordinates. Phosphorylation could affect migration, but these annotations alone do not demonstrate a visible shift. State the numbering convention when comparing antibody or paper site labels.
Does this guide establish induction of FBXW7?
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 FBXW7?
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 A00406-2 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 FBXW7 bands be quantified across samples?
Quantitation · Quantify the same identified band in comparable preparations. Record whether samples are whole-cell or nuclear fractions: UniProt places isoform 1 in the nucleoplasm and at chromosomes. Its three isoforms may not share an antibody epitope, so verify which species the assay detects before combining band intensities.
Why is FBXW7 observed near 75 kDa instead of 79.7 kDa?
Interpretation · The supplied apparent band is about 75 kDa, while the predicted mass is 79.7 kDa. These values use different measurements. The listed features do not establish the cause of the difference; identify the band with an appropriate specificity control.
UniProt reports that isoform 1 localizes to double-strand-break sites following ATM phosphorylation. If comparing samples with different DNA damage conditions, keep the preparation and fraction analyzed consistent. The feature supports a localization change, not a predicted change in total band intensity.
Consider the three annotated isoforms and antibody cross-reactivity when assessing extra bands. FBXW7 also forms homodimers, but that annotation does not establish a dimer band under Western-blot conditions. Confirm band identity before assigning a species or attributing migration to phosphorylation.
UniProt reports that isoform 1 localizes to double-strand-break sites following ATM phosphorylation. If comparing samples with different DNA damage conditions, keep the preparation and fraction analyzed consistent. The feature supports a localization change, not a predicted change in total band intensity.
Consider the three annotated isoforms and antibody cross-reactivity when assessing extra bands. FBXW7 also forms homodimers, but that annotation does not establish a dimer band under Western-blot conditions. Confirm band identity before assigning a species or attributing migration to phosphorylation.
Boster reagents
FBXW7 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-FBXW7 Antibody Picoband®
Cat # A00406-2
The catalog reports one anti-FBXW7 antibody, A00406-2, with reported human, mouse, and rat reactivity. Its WB image shows an approximately 75 kDa band in the tested lysates. No publication evidence is supplied.
Which to pick: A00406-2 is the only listed option. Its WB caption documents human MCF-7, 293T, Jurkat, and HepG2 cells; rat brain and PC-12; and mouse brain and Nih/3T3, using 30 µg reducing lysate and 0.5 µg/mL primary antibody.
Source: BosterBio FBXW7 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q969H0.
- Human Protein Atlas. FBXW7 tissue expression.
- PMC10809768 — target-verified WB comparison
- PMC6964669 — target-verified WB comparison
- PMC10054043 — target-verified WB comparison