This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
RNF181 / E3 ubiquitin-protein ligase RNF181 · Western blot design guide
Design a Western Blot for RNF181
Source-linked RNF181 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-RNF181 WB antibodies. Everything you need to plan the experiment before you commit precious samples.
Evidence assembled October 2026 · For research use; verify linked source records and product datasheet before use
RNF181 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 | ~17.9 kDa | |
| Gel | 15% (standard starting point) | |
| Positive control | Testis (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 + Ubl conjugation | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked RNF181 Western Blot Protocol Options
The A10627 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | extracts of various cell lines, (catalog A10627) |
| Gel % | 15% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Short semi-dry transfer; verify retention (standard starting point) |
| Membrane | 0.2 µm PVDF (standard starting point) |
| Blocking | Blocking buffer: 3% nonfat dry milk in TBST (catalog A10627) |
| Primary antibody | A10627 · 1:3000 (catalog A10627) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | Goat Anti-Rabbit IgG, 1:10000 (catalog A10627) |
| Secondary incubation | 1 h at room temperature (standard starting point) |
| Wash | 3 × 5 min in TBST (standard starting point) |
| Detection | ECL (catalog A10627) |
Section 2
What Is the Expected RNF181 Western Blot Band Size?
RNF181 is predicted at 17.9 kDa; Thr153 phosphorylation is annotated, but its effect on migration is unproven and no empirical band size is supplied.
| Band near 17.9 kDa | compatible with RNF181's predicted mass; confirm its identity |
| Faint band near 17.9 kDa | compatible with RNF181, but identity remains unconfirmed |
| No band near 17.9 kDa | RNF181 is undetected under these conditions |
| Doublet near 17.9 kDa | RNF181 identity and any contribution from Thr153 phosphorylation remain unestablished |
💡Expected RNF181 appearanceRNF181 has a predicted mass of 17.9 kDa, but no empirical band size is supplied; a band near that size needs confirmation with band-identity controls.
| UniProt predicted mass | sets a 17.9 kDa reference, not a validated migration position |
| 153 amino acid sequence | underlies the full-length predicted mass; no processing site is annotated |
| Single annotated isoform | provides no basis for predicting distinct isoform bands |
| Thr153 phosphorylation | is annotated, but no visible migration shift is established |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | RNF181 is undetected; its location is not supplied | check loading and use a positive control |
| Band higher than expected | the higher band's identity or migration is unverified | compare with a validated positive control and test RNF181 depletion |
| Band lower than expected | the lower band's identity is unverified; no cleavage site is annotated | test RNF181 depletion and check sample integrity |
| Multiple bands | additional bands are not explained by the single annotated isoform | identify which bands respond to RNF181 depletion |
| Weak or no signal | RNF181 abundance or assay sensitivity may be insufficient | check loading, antibody performance and a positive control |
Experiment evidence
Western blot images showing RNF181 band
A10627 · RNF181
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for RNF181 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for RNF181 in Western blot, you can use testis tissue, which HPA rates as highly expressed.
Positive control: Testis (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: HPA identifies a high-expression tissue and a not-detected tissue, so tissue controls are feasible.
HPA tissue expression evidence for RNF181
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 |
|---|---|---|---|---|
| Testis | pachytene spermatocytes | High | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | glandular cells | Medium | Protein (IHC) | HPA → |
| Breast | glandular cells | Medium | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | Medium | 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 → |
| Caudate | glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebral cortex | endothelial cells | Not detected | Protein (IHC) | HPA → |
| Hippocampus | glial cells | Not detected | Protein (IHC) | HPA → |
| Skeletal muscle | myocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced RNF181 Western Blot Tips
Deeper troubleshooting and optimisation questions for RNF181, answered from its protein features.
How should RNF181 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 RNF181 isoforms explain multiple bands?
Isoforms · UniProt lists one isoform and no alternative sequence. Do not assign additional bands to RNF181 isoforms based on these features alone.
Does RNF181 phosphorylation affect band interpretation?
PTM · UniProt lists phosphothreonine at position 153, the last residue of the 153-residue sequence. This is UniProt sequence numbering; antibody or paper numbering may differ. The site alone does not establish a detectable band shift.
Does this guide establish induction of RNF181?
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 RNF181 Western blot?
Transfer · RNF181 is predicted to be 17.9 kDa. Choose and verify transfer conditions that retain a protein of this size, including checking for protein passing through the membrane. The supplied features do not specify a particular transfer method.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A10627 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 RNF181 bands be quantified?
Quantitation · Quantify a consistently identified band near the predicted 17.9 kDa across comparable samples. If bands differ between conditions, avoid combining them as RNF181 without confirming their identity; the listed phosphothreonine site does not establish that separate bands represent different phosphorylation states.
Should RNF181 run exactly at its predicted 17.9 kDa?
Interpretation · Use 17.9 kDa as the predicted mass, not an exact band position. No observed band size is supplied, and the listed features do not establish a visible shift or explain any difference between apparent and predicted mass.
The supplied features list no glycosylation sites or alternative sequence, so neither provides an explanation for extra bands. RNF181 directly interacts with ITGA2B and the ITGA2B/ITGB3 integrin, but that interaction alone does not identify another band as RNF181 or establish a covalent complex.
The supplied features list no glycosylation sites or alternative sequence, so neither provides an explanation for extra bands. RNF181 directly interacts with ITGA2B and the ITGA2B/ITGB3 integrin, but that interaction alone does not identify another band as RNF181 or establish a covalent complex.
Boster reagents
RNF181 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-RNF181 Antibody
Cat # A10627
The catalog reports one anti-RNF181 antibody, A10627, with stated human, mouse, and rat reactivity. Its WB image uses extracts from various cell lines at 1:3000 and 25 µg per lane. The caption does not identify the cell lines or their species.
Which to pick: A10627 is the only listed option and has a WB image. Consider it for the stated human, mouse, or rat reactivity, but the image does not establish which species were tested.
Source: BosterBio RNF181 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q9P0P0.
- Human Protein Atlas. RNF181 tissue expression.
- PMC5840266 — target-verified WB comparison