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- Table of Contents
RAB8A / Ras-related protein Rab-8A · Western blot design guide
Design a Western Blot for RAB8A
Real validated RAB8A Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-RAB8A 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
RAB8A 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 | ~23.7 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 | Phosphorylated + Methylated | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 2 isoform(s) |
Section 1
Real Curated RAB8A Western Blot Protocols
The M02180 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | HeLa cell lysate (catalog M02180) |
| Gel % | 12–15% (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 | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | M02180; 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 RAB8A Western Blot Band Size?
Full-length RAB8A is predicted at 23.7 kDa; propeptide cleavage and isoforms could affect bands, but their migration effects are unproven.
| Band near 23.7 kDa | Consistent with the predicted full-length RAB8A mass; identity requires confirmation |
| Band just below the precursor | Could reflect cleavage of the 205..207 propeptide; its migration is unverified |
| Multiple nearby bands | Could reflect isoforms 1 and 2; distinct migration is unverified |
| Stronger band in a membrane-enriched fraction | Consistent with lipid-anchored, membrane-associated RAB8A |
💡Expected RAB8A appearanceUniProt predicts 23.7 kDa for full-length RAB8A; no empirical band size is supplied, and any shifted band needs confirmation with antibody specificity and RAB8A depletion controls.
| Predicted full-length mass | 23.7 kDa is the sequence-based reference, not a measured migration |
| 205..207 propeptide cleavage | Could yield a slightly smaller mature protein; migration is unverified |
| Isoform 1 | May differ in size from isoform 2; relative size is unknown |
| Isoform 2 | May differ in size from isoform 1; relative size is unknown |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Lipid-anchored RAB8A may be poorly recovered from membranes | Check membrane extraction and a membrane-enriched fraction |
| Band higher than expected | The supplied features do not establish a higher migrating RAB8A species | Check the size marker and confirm band identity by RAB8A depletion |
| Band lower than expected | Propeptide cleavage could cause a small decrease; a large decrease remains unexplained | Confirm identity by RAB8A depletion and assess sample degradation |
| Multiple bands | Isoforms 1 and 2 exist, but distinct band sizes are unverified | Test which bands respond to RAB8A depletion |
| Weak or no signal | Membrane-associated RAB8A may be underrepresented after extraction | Check extraction efficiency and antibody performance with a positive lysate |
Experiment evidence
Western blot images showing RAB8A band
M02180 · RAB8A
PA2280 · RAB8A
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for RAB8A Western blot
🧪For positive controls for RAB8A in Western blot, you can use an independently validated RAB8A-positive lysate, since no HPA positive sample is supplied.
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 such as Ponceau alongside the samples.
⚠️Feasibility: HPA expression data are unavailable, so tissue controls cannot be selected from the supplied evidence.
HPA tissue expression evidence for RAB8A
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 RAB8A Western Blot Tips
Deeper troubleshooting and optimisation questions for RAB8A, answered from its protein features.
How should RAB8A 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 RAB8A isoforms produce different bands?
Isoforms · UniProt lists two isoforms. Isoform 2 replaces the canonical sequence at 161..205, so an antibody recognizing that region may detect the isoforms differently. The feature does not establish whether they resolve as separate bands.
Which RAB8A phosphorylation sites matter when interpreting bands?
PTM · UniProt lists phosphothreonine at 72 by LRRK2 and phosphoserines at 181 and 185. These are UniProt coordinates; check the numbering convention used by an antibody or paper before comparing sites. A phosphorylation annotation alone does not prove a detectable band shift.
Does this guide establish induction of RAB8A?
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 RAB8A Western blot?
Transfer · RAB8A is a 23.7 kDa protein annotated at membranes with a lipid anchor. Choose transfer conditions that retain proteins near this size, then check both the membrane and post-transfer gel to assess recovery. The supplied features do not identify one validated transfer method.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the M02180 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 RAB8A be quantified across samples?
Quantitation · RAB8A is annotated at the cell membrane, Golgi, and endosome membranes. Compare like fractions or use consistent whole-cell extraction across samples; changes in membrane recovery could affect the measured signal. Quantify a band only after establishing that the antibody detects RAB8A.
Why might RAB8A migrate away from its predicted 23.7 kDa?
Interpretation · Use 23.7 kDa as a starting point, not an exact band position. UniProt lists phosphorylation, a cysteine methyl ester at 204, and a propeptide at 205..207. These features alone do not establish a visible shift, and no observed band position is supplied.
Check whether the antibody recognizes the region replaced in isoform 2 (161..205). Also consider the annotated phosphorylation sites at 72, 181, and 185, cysteine methyl ester at 204, and propeptide at 205..207. None of these annotations identifies an unexpected band on its own.
The listed propeptide spans 205..207, and isoform 2 replaces residues 161..205. Check an antibody's epitope against both regions when comparing isoforms or processed protein. UniProt coordinates refer to the supplied canonical sequence; antibody or paper numbering may differ.
Check whether the antibody recognizes the region replaced in isoform 2 (161..205). Also consider the annotated phosphorylation sites at 72, 181, and 185, cysteine methyl ester at 204, and propeptide at 205..207. None of these annotations identifies an unexpected band on its own.
The listed propeptide spans 205..207, and isoform 2 replaces residues 161..205. Check an antibody's epitope against both regions when comparing isoforms or processed protein. UniProt coordinates refer to the supplied canonical sequence; antibody or paper numbering may differ.
Boster reagents
RAB8A 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-RAB8A Rabbit Monoclonal Antibody
Cat # M02180
Real WB data
Anti-Ras-related protein Rab-8A RAB8A Antibody Picoband®
Cat # PA2280
Both listed anti-RAB8A antibodies have Western blot images: M02180 in HeLa lysate and PA2280 in Caco-2, HepG2, and A549 lysates. PA2280’s caption reports a band near 24 kDa. The supplied evidence does not establish orthogonal specificity.
Which to pick: For human lysates, PA2280 has a detailed WB example using 50 μg of lysate per lane and 0.5 μg/mL antibody; M02180 has a HeLa image. For mouse or rat, M02180 lists reactivity, but its shown WB sample is human.
Source: BosterBio RAB8A gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P61006.
- Human Protein Atlas. RAB8A tissue expression.
- PMC11565912 — target-verified WB comparison
- PMC3547156 — target-verified WB comparison
- PMC5828482 — target-verified WB comparison