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- Table of Contents
RAB23 / Ras-related protein Rab-23 · Western blot design guide
Design a Western Blot for RAB23
Source-linked RAB23 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-RAB23 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
RAB23 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 | ~26.7 kDa | |
| Observed band | ~27 kDa | |
| Gel | 5–20% (catalog A04593-2) | |
| 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 | Phosphorylated + Methylated | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked RAB23 Western Blot Protocol Options
The A04593-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human 293T, human MCF-7, human Jurkat, human HepG2, rat brain, mouse brain (catalog A04593-2) |
| Gel % | 5–20% (catalog A04593-2) |
| Load | 30 ug; reducing conditions (catalog A04593-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A04593-2) |
| Membrane | nitrocellulose membrane (catalog A04593-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A04593-2) |
| Primary antibody | A04593-2 · 0.5 μg/mL (catalog A04593-2) |
| Primary incubation | overnight at 4°C (catalog A04593-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A04593-2) |
| Secondary incubation | 1.5 hour at RT (catalog A04593-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A04593-2) |
| Detection | ECL (catalog A04593-2) |
Section 2
What Is the Expected RAB23 Western Blot Band Size?
RAB23 is predicted at 26.7 kDa and observed at ~27 kDa; the small difference has no established cause in the supplied evidence.
| Single band at ~27 kDa | Empirical RAB23 band, close to its 26.7 kDa predicted mass |
| Band near 26.7 kDa | Consistent with the predicted full-length mass; confirm identity with controls |
| Band slightly below the precursor | Could reflect removal of the 235–237 propeptide, if cleavage occurs and is detectable |
| Weak band in soluble fraction | May reflect RAB23 association with membranes through a lipid anchor |
💡Expected RAB23 appearanceRAB23 has a predicted mass of 26.7 kDa and an empirical band at ~27 kDa; confirm band identity with antibody and sample controls, especially for additional bands.
| UniProt predicted mass | Places the full-length protein near 26.7 kDa |
| Full-length 237-residue precursor | Provides the reference size before any propeptide cleavage |
| C-terminal propeptide at residues 235–237 | Its removal would slightly reduce mass, but visible separation is unestablished |
| Mature protein after propeptide removal | Could migrate slightly below the precursor if cleavage occurs |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Membrane-associated RAB23 may be poorly extracted | Check extraction and compare membrane-enriched material |
| Band higher than expected | The higher band has no established size-shifting explanation in the supplied features | Check antibody specificity with RAB23 depletion and a positive-control lysate |
| Band lower than expected | Propeptide removal could make a small difference; a larger shift is unexplained | Compare with a positive control and verify identity by RAB23 depletion |
| Multiple bands | The supplied features do not establish distinct resolvable RAB23 bands | Identify the RAB23-dependent band using depletion and a positive control |
| Weak or no signal | Low recovery of membrane-associated RAB23 may reduce signal | Check membrane extraction, loading, and a positive-control lysate |
Experiment evidence
Western blot images showing RAB23 band
A04593-2 · RAB23
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for RAB23 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for RAB23 in Western blot, you can use adrenal gland tissue, which shows high HPA expression.
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 such as Ponceau alongside the samples.
⚠️Feasibility: No tissue is listed as not detected, so use siRNA knockdown or a KO line for a clean negative control.
HPA tissue expression evidence for RAB23
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 → |
| Bronchus | respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Endometrium | glandular cells | High | Protein (IHC) | HPA → |
| Esophagus | squamous epithelial cells | High | Protein (IHC) | HPA → |
| Fallopian tube | glandular cells | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Lymph node | germinal center cells | Low | Protein (IHC) | HPA → |
| Spleen | cells in red pulp | Low | Protein (IHC) | HPA → |
| Adipose tissue | adipocytes | Medium | Protein (IHC) | HPA → |
| Appendix | glandular cells | Medium | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | Medium | Protein (IHC) | HPA → |
Section 3
Advanced RAB23 Western Blot Tips
Deeper troubleshooting and optimisation questions for RAB23, answered from its protein features.
How should RAB23 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.
Do annotated isoforms explain multiple RAB23 bands?
Isoforms · Only one isoform is supplied, with no alternative sequence. The supplied features therefore do not identify an isoform responsible for an additional band.
Which modifications should inform band interpretation?
PTM · UniProt lists phosphoserine at positions 186 and 187 and a cysteine methyl ester at position 234. These are UniProt coordinates. The annotations alone do not show that any modification causes a visible shift; establish the identity of an extra band experimentally.
Does this guide establish induction of RAB23?
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 RAB23?
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 A04593-2 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 RAB23 signal be quantified across samples?
Quantitation · Use a consistent sample preparation and compare equivalent fractions: RAB23 has both cytoplasmic and membrane-associated locations. A change in one fraction’s signal need not represent the same change in total RAB23.
Why is the RAB23 band near 27 kDa?
Interpretation · The observed band near 27 kDa agrees with RAB23’s predicted mass of 26.7 kDa. Its annotated modifications do not, by themselves, establish a detectable band shift.
UniProt annotates a propeptide at residues 235..237. Check whether the antibody recognizes this C-terminal region when assessing detection. These are UniProt coordinates; antibody or paper numbering may differ.
RAB23 is annotated in the cytoplasm and at membranes, including the cytoplasmic side of the cell membrane and the endosome membrane. Consider which fraction your preparation retains when comparing signal across samples.
Start with the expected band near 27 kDa. Check an extra band’s identity before assigning it to the annotated propeptide, phosphorylation, methylation, or lipid anchoring; feature presence alone does not establish a visible band shift.
UniProt annotates a propeptide at residues 235..237. Check whether the antibody recognizes this C-terminal region when assessing detection. These are UniProt coordinates; antibody or paper numbering may differ.
RAB23 is annotated in the cytoplasm and at membranes, including the cytoplasmic side of the cell membrane and the endosome membrane. Consider which fraction your preparation retains when comparing signal across samples.
Start with the expected band near 27 kDa. Check an extra band’s identity before assigning it to the annotated propeptide, phosphorylation, methylation, or lipid anchoring; feature presence alone does not establish a visible band shift.
Boster reagents
RAB23 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-RAB23 Antibody Picoband®
Cat # A04593-2
The catalog reports one anti-RAB23 antibody for Western blot. Its supplied blot shows an approximately 27 kDa band in human cell lines and rat and mouse brain lysates. No publication evidence or independent validation is supplied.
Which to pick: A04593-2 is the only listed option. It is catalogued for human, mouse and rat and has a WB image using human cell lines and rat and mouse brain lysates, with 30 µg loaded per lane and antibody at 0.5 µg/mL.
Source: BosterBio RAB23 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q9ULC3.
- Human Protein Atlas. RAB23 tissue expression.
- PMC10725407 — target-verified WB comparison
- PMC10809273 — target-verified WB comparison
- PMC6607405 — target-verified WB comparison