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- Table of Contents
RRAS / Ras-related protein R-Ras · Western blot design guide
Design a Western Blot for RRAS
Source-linked RRAS Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-RRAS 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
RRAS 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 | ~23.5 kDa | |
| Observed band | ~23 kDa | |
| Gel | 5–20% (catalog A04310-1) | |
| Positive control | Adrenal gland (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 | Methylated + Cleaved | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked RRAS Western Blot Protocol Options
The A04310-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human A549, human Hela, human PC-3 (catalog A04310-1) |
| Gel % | 5–20% (catalog A04310-1) |
| Load | 30 ug; reducing conditions (catalog A04310-1) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A04310-1) |
| Membrane | nitrocellulose membrane (catalog A04310-1) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A04310-1) |
| Primary antibody | A04310-1 · 0.5 μg/mL (catalog A04310-1) |
| Primary incubation | overnight at 4°C (catalog A04310-1) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A04310-1) |
| Secondary incubation | 1.5 hour at RT (catalog A04310-1) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A04310-1) |
| Detection | ECL (catalog A04310-1) |
Section 2
What Is the Expected RRAS Western Blot Band Size?
RRAS is predicted at 23.5 kDa and observed near 23 kDa; the cause of the difference is not established.
| Single band at ~23 kDa | Matches the empirical RRAS band in whole-cell lysates. |
| Band near 23.5 kDa | Consistent with the UniProt predicted mass; band identity still needs confirmation. |
| Band slightly below the precursor | Could reflect removal of the 216–218 propeptide, but a visible shift is not established. |
| Weak band in a soluble fraction | Could reflect RRAS localization at the cell membrane. |
💡Expected RRAS appearanceUniProt predicts 23.5 kDa, while antibody QC shows a ~23 kDa band in whole-cell lysates; confirm identity with an appropriate RRAS control.
| UniProt predicted mass | Places the expected protein near 23.5 kDa. |
| 218-residue sequence | Underlies the predicted 23.5 kDa mass. |
| Propeptide at residues 216–218 | Is present in the precursor represented by the predicted mass. |
| Propeptide cleavage at residues 216–218 | Could slightly lower mature protein mass; a resolvable shift is not established. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Membrane-associated RRAS may be poorly recovered during extraction. | Check a membrane-enriched preparation and a validated RRAS-positive lysate. |
| Band higher than expected | The supplied features do not establish a larger RRAS species. | Compare with the ~23 kDa positive control and test band identity by RRAS depletion. |
| Band lower than expected | Removal of the 216–218 propeptide could make only a small mass difference. | Compare with the ~23 kDa control and verify the smaller band's identity. |
| Multiple bands | Only one isoform is listed, so additional bands have no established isoform assignment. | Check which band decreases after RRAS depletion. |
| Weak or no signal | Recovery of membrane-associated RRAS may be low. | Check extraction and loading against a validated RRAS-positive lysate. |
Experiment evidence
Western blot images showing RRAS band
A04310-1 · RRAS
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for RRAS Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for RRAS in Western blot, you can use adrenal gland tissue, which HPA rates as highly expressed.
Positive control: Adrenal gland (IHC candidate; verify WB)
Negative control: Suggested KO / knockdown lysate
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: No HPA tissue is reported as not detected, so use siRNA knockdown or a KO line for a negative control.
HPA tissue expression evidence for RRAS
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 → |
| Bone marrow | hematopoietic cells | High | Protein (IHC) | HPA → |
| Colon | glandular cells | High | Protein (IHC) | HPA → |
| Duodenum | glandular cells | High | Protein (IHC) | HPA → |
| Gallbladder | glandular cells | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Liver | cholangiocytes | Low | Protein (IHC) | HPA → |
| Adipose tissue | adipocytes | 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 → |
Section 3
Advanced RRAS Western Blot Tips
Deeper troubleshooting and optimisation questions for RRAS, answered from its protein features.
How should RRAS 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 RRAS isoforms explain multiple bands?
Isoforms · UniProt lists one isoform and no alternative sequence here. Do not assign additional bands to RRAS isoforms on this evidence alone.
Do annotated modifications prove a band shift?
PTM · The linked UniProt record describes protein features. A modification annotation alone does not demonstrate a visible shift; retain any condition or experimental qualifier attached to it.
Does this guide establish induction of RRAS?
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 RRAS Western blot?
Transfer · RRAS is a ~23.5 kDa protein associated with the cytoplasmic side of the cell membrane through a lipid anchor. Choose transfer conditions that retain proteins near 23 kDa, and verify transfer before interpreting a weak band. The supplied features do not specify a particular transfer method.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A04310-1 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 an RRAS Western blot band?
Quantitation · Quantify the band near the reported ~23 kDa position within the assay’s linear range, with consistent loading and transfer across samples. The supplied features do not establish that band intensity measures GTP-bound RRAS or its interactions with PLCE1, RGS14, or OSBPL3.
Why is the RRAS band near 23 kDa rather than 23.5 kDa?
Interpretation · The reported ~23 kDa band is close to the predicted 23.5 kDa mass. The listed features alone cannot explain the small difference in apparent mass or establish a visible shift.
UniProt marks a cysteine methyl ester at position 215 and a propeptide at 216..218. These are UniProt coordinates; paper or antibody numbering may differ. The annotations do not establish whether either feature produces a visible band shift.
First check whether they reproduce and whether they track with RRAS using an independent specificity check. UniProt lists one isoform, no glycosylation sites, a methyl ester at Cys215, and a propeptide at 216..218. None of those annotations alone identifies an unexpected band or proves a shift.
UniProt marks a cysteine methyl ester at position 215 and a propeptide at 216..218. These are UniProt coordinates; paper or antibody numbering may differ. The annotations do not establish whether either feature produces a visible band shift.
First check whether they reproduce and whether they track with RRAS using an independent specificity check. UniProt lists one isoform, no glycosylation sites, a methyl ester at Cys215, and a propeptide at 216..218. None of those annotations alone identifies an unexpected band or proves a shift.
Boster reagents
RRAS 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-RRAS Antibody Picoband®
Cat # A04310-1
A04310-1 is a rabbit polyclonal anti-RRAS antibody listed for human samples. Its Western blot image shows an approximately 23 kDa band in A549, HeLa, and PC-3 whole-cell lysates at 0.5 μg/mL. Evidence supplied is limited to these tested contexts.
Which to pick: A04310-1 is the only listed option. Its Western blot image uses human A549, HeLa, and PC-3 lysates, making it the documented choice for those sample contexts; performance in other samples is not established here.
Source: BosterBio RRAS gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P10301.
- Human Protein Atlas. RRAS tissue expression.
- PMC10086563 — target-verified WB comparison
- PMC12722598 — target-verified WB comparison
- PMC7319537 — target-verified WB comparison