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- Table of Contents
UBR5 / E3 ubiquitin-protein ligase UBR5 · Western blot design guide
Design a Western Blot for UBR5
Real validated UBR5 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-UBR5 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
UBR5 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 | ~309.4 kDa | |
| Observed band | ~280 kDa | |
| Gel | 8% (catalog A03160-1) | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Acetylated | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 2 isoform(s) |
Section 1
Real Curated UBR5 Western Blot Protocols
The A03160-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human T-47D, human MCF-7, rat brain (catalog A03160-1) |
| Gel % | 8% (catalog A03160-1) |
| Load | 30 ug; reducing conditions (catalog A03160-1) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A03160-1) |
| Membrane | nitrocellulose membrane (catalog A03160-1) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A03160-1) |
| Primary antibody | A03160-1 · 0.5 μg/mL (catalog A03160-1) |
| Primary incubation | overnight at 4°C (catalog A03160-1) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A03160-1) |
| Secondary incubation | 1.5 hour at RT (catalog A03160-1) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A03160-1) |
| Detection | ECL (catalog A03160-1) |
Section 2
What Is the Expected UBR5 Western Blot Band Size?
UBR5 is predicted at 309.4 kDa, while antibody QC detects ~280 kDa; the cause of the difference is not established.
| Band at ~280 kDa | Empirical UBR5 band in reducing whole-cell lysates; confirm identity with antibody controls. |
| Band near 309 kDa | Possible migration near the 309.4 kDa predicted monomer mass; identity requires confirmation. |
| Doublet or multiple high-mass bands | Isoforms 1 and 2 are possible contributors, but distinct migration has not been established. |
| Very high-mass band or material near the well | Possible retained UBR5 oligomers; UBR5 forms a homotetramer, but persistence during SDS-PAGE is unproven. |
💡Expected UBR5 appearanceUBR5 has a predicted monomer mass of 309.4 kDa, while antibody QC shows a ~280 kDa band in reducing lysates; the difference is unexplained, so confirm identity with appropriate controls.
| Predicted UBR5 monomer mass | Sequence predicts 309.4 kDa; the empirical band is ~280 kDa, with no established cause for the difference. |
| Isoform 1 | May migrate differently from isoform 2; its individual mass and band position are unknown. |
| Isoform 2 | May migrate differently from isoform 1; its individual mass and band position are unknown. |
| Homotetrameric assembly | Could produce higher-mass signal if oligomers persist, though persistence during SDS-PAGE is unproven. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | UBR5 may be insufficiently represented in the loaded sample or poorly transferred at its high mass. | Check sample loading and high-mass transfer with a positive lysate. |
| Band higher than expected | UBR5 oligomers may persist if denaturation is incomplete. | Compare fully denatured samples and confirm the band with an independent antibody. |
| Band lower than expected | The ~280 kDa empirical band is below the 309.4 kDa sequence prediction; its cause is unknown. | Compare with a positive lysate and confirm identity with an independent antibody or UBR5 depletion. |
| Multiple bands | UBR5 has isoforms 1 and 2, but their distinct migration has not been established. | Check which bands decrease after UBR5 depletion. |
| Weak or no signal | Transfer of this large protein may be inefficient. | Inspect high-mass transfer and use a documented positive lysate. |
| Fragments below expected size | Sample degradation or nonspecific antibody binding may produce lower bands; no UBR5 cleavage product is specified. | Process samples promptly with protease inhibitors and check band identity by UBR5 depletion. |
Experiment evidence
Western blot images showing UBR5 band
A03160-1 · UBR5
M03160 · UBR5
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for UBR5 Western blot
🧪For positive controls for UBR5 in Western blot, you can use an HPA-IHC candidate positive sample once one is identified; none is supplied here.
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.
⚠️Feasibility: HPA tissue data are unavailable, so a positive tissue and a clean negative tissue cannot be selected.
HPA tissue expression evidence for UBR5
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 UBR5 Western Blot Tips
Deeper troubleshooting and optimisation questions for UBR5, answered from its protein features.
How should UBR5 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 UBR5 isoforms produce different bands?
Isoforms · UniProt lists isoforms 1 and 2; isoform 2 lacks residue 2474 in canonical UniProt numbering. That single-residue difference alone does not establish a visibly separate band. Check which isoform and epitope your antibody recognizes.
Could phosphorylation change UBR5 band appearance?
PTM · UniProt lists numerous phosphorylation sites, including Ser2026, Ser2028, Thr2030, Ser2484 and Ser2486 in canonical numbering. Their presence does not demonstrate a visible shift. If comparing phosphatase-treated and untreated samples, interpret any change alongside a total UBR5 blot; verify numbering before comparing sites with antibody documentation.
Does this guide establish induction of UBR5?
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.
What transfer method to use for UBR5 Western blot?
Transfer · UBR5 is large, with a predicted mass of 309.4 kDa. Use a transfer setup suitable for high-molecular-weight proteins, then check both the membrane and the post-transfer gel to see whether UBR5 transferred. The supplied features do not specify a membrane type or transfer setting.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A03160-1 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 UBR5 bands be quantified?
Quantitation · Quantify the same consistently identified UBR5 band across samples; the reported apparent band is about 280 kDa. Normalize to an appropriate loading measure and keep exposure within its linear range. Do not combine other bands with the main band unless their identity has been verified.
Why is the observed UBR5 band below its predicted mass?
Interpretation · UBR5 has a predicted mass of 309.4 kDa, while the reported band is about 280 kDa. Its listed modifications and isoforms do not establish the cause of that difference. Check the band with an antibody to a second region of UBR5 before assigning it as full length.
UBR5 is reported in both nucleus and cytoplasm and forms a homotetramer. Its two listed isoforms differ by one missing residue, which alone does not identify a distant band. Compare fractions and use an antibody to another UBR5 region to assess band identity; the supplied features do not establish the cause of an unexpected band.
UBR5 is reported in both nucleus and cytoplasm and forms a homotetramer. Its two listed isoforms differ by one missing residue, which alone does not identify a distant band. Compare fractions and use an antibody to another UBR5 region to assess band identity; the supplied features do not establish the cause of an unexpected band.
Boster reagents
UBR5 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-UBR5 Antibody Picoband®
Cat # A03160-1
Real WB data
Anti-EDD Rabbit Monoclonal Antibody
Cat # M03160
Two the supplier antibodies are listed for UBR5 Western blotting. A03160-1 has a WB image using human cell and rat and mouse brain lysates; the reported band is approximately 280 kDa versus 309 kDa expected. M03160 has an SH-SY5Y lysate image with limited caption detail.
Which to pick: For rat samples, A03160-1 is the listed option with stated rat reactivity. For human or mouse samples, both list reactivity; A03160-1 provides more detailed WB conditions, while M03160 shows an SH-SY5Y lysate blot.
Source: BosterBio UBR5 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry O95071.
- Human Protein Atlas. UBR5 tissue expression.
- PMC9665233 — target-verified WB comparison
- PMC8319180 — target-verified WB comparison
- PMC7994773 — target-verified WB comparison