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SOX18 · Western blot design guide
Design a Western Blot for SOX18
Real validated SOX18 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-SOX18 WB antibodies. Everything you need to plan the experiment before you commit precious samples.
Last reviewed: May 2026 · Scientific review: Boster Bio technical team
SOX18 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 | ~40.9 kDa | |
| Gel | 10–12% | |
| Negative control | siRNA / KO lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated | |
| Caveat | Phosphorylation mobility shift | |
| Regulation | IFN-γ-induced | |
| Isoform | 1 isoform(s) |
Section 1
Real Curated SOX18 Western Blot Protocols
Literature-validated Western blot parameters for SOX18 — gel percentage, transfer, blocking, antibody incubation and detection, extracted from published methods.
| Sample / lysate | human Jurkat , Lane 2: human COLO-320 , Lane 3: human Hela , Lane 4: human HepG2 , Lane 5: monkey COS-7 , Lane 6: rat heart , Lane 7: rat lung , Lane 8: rat liver , Lane 9: mouse lung , Lane 10: mouse liver . After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/ TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-SOX18 antigen affinity purified polyclonal antibody (Catalog # A04004-2) at 0.25 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for SOX18 at approximately 41KD. The expected band size for SOX18 is at 41KD |
| Gel % | 10–12% |
| Load | 30ug |
| Transfer | nitrocellulose membrane, 150 mA, 50–90 min |
| Membrane | nitrocellulose |
| Blocking | 5% non-fat milk / TBS, 1.5 h RT |
| Primary antibody | 0.25 µg/mL |
| Primary incubation | overnight at 4 °C |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 |
| Wash | TBS-0.1% Tween, 3 × 5 min |
| Detection | ECL |
| Exposure / imaging | Tanon 5200 |
| Observed band | 41 kDa |
Section 2
What Is the Expected SOX18 Western Blot Band Size?
SOX18 is a ~41 kDa nuclear phosphoprotein (predicted 40.9 kDa); expect a single sharp band, with only a minor shift possible from Ser70 phosphorylation.
| single sharp band near 41 kDa | matches the predicted mass of unmodified, full-length SOX18 with no glycosylation or proteolytic cleavage annotated |
| band present in whole-cell or nuclear lysate | SOX18 is a nuclear transcription factor, not secreted, so it should be retained and detectable in whole-cell or nuclear extracts rather than absent |
| slight upward smear or fuzzy edge on the main band | phosphorylation at Ser70 can subtly alter apparent mobility relative to the unmodified predicted mass |
| no doublet or extra band from splice variants | only one isoform is annotated for SOX18, so alternative splicing should not generate additional bands |
| no band at roughly twice the monomer mass | no inter-chain disulfide bond is annotated; SOX18 partners with MEF2C non-covalently, so a covalent dimer band is not expected |
💡Expected SOX18 appearanceExpect a single, sharp band around 41 kDa (predicted 40.9 kDa, 384 aa), with at most a slight mobility shift from Ser70 phosphorylation; no glycosylation, cleavage, or covalent dimerization is annotated.
| Predicted mass (UniProt) | sets the baseline expected band at 40.9 kDa for the 384-aa unmodified protein |
| Phosphorylation at Ser70 | can cause a subtle upward shift in apparent molecular weight relative to the unmodified predicted mass |
| Absence of glycosylation sites | no sugar-driven mass addition or smearing is expected, so the band should stay sharp rather than diffuse |
| Absence of inter-chain disulfide bonds | no covalent homodimer band near twice the monomer mass should appear, even under non-reducing conditions |
| No signal peptide or propeptide | SOX18 is not secreted or proteolytically processed into a mature form, so no smaller cleaved fragment relative to full length is expected |
| Single annotated isoform | only one band is expected from alternative splicing; additional isoform-driven bands should not occur |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | SOX18 is a nuclear protein that can be under-extracted if the lysis method does not adequately disrupt the nucleus, or expression is low in the sampled cell type | use a lysis buffer suited to nuclear extraction (with sonication or a dedicated nuclear lysis step) and confirm the cell line or tissue actually expresses SOX18 |
| Band higher than expected | Ser70 phosphorylation or incomplete denaturation/reduction can shift apparent mobility upward | ensure fresh reducing agent and complete boiling of samples, and consider phosphatase treatment to test whether the shift is phosphorylation-driven |
| Band lower than expected | proteolytic degradation during nuclear lysate preparation can generate a truncated product | add protease inhibitors, keep samples cold throughout nuclear extraction, and process lysates quickly |
| Multiple bands | nonspecific antibody cross-reactivity with other SOXF-family members sharing HMG-box homology, since SOX18 itself has only one annotated isoform | validate antibody specificity using SOX18 knockdown or knockout lysate and compare the band pattern |
| Weak or no signal | SOX18 is a low-abundance nuclear transcription factor restricted to specific tissues such as vascular endothelium | enrich for the nuclear fraction, load more total protein, and include a tissue known to express SOX18 as a positive control |
Experiment evidence
Western blot images showing SOX18 band
A04004-2 · SOX18
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for SOX18 Western blot
🧪For positive controls for SOX18 in Western blot, you can use lysate from cells transiently transfected to overexpress SOX18 (e.g., HEK293 cells), since no Human Protein Atlas expression data are available to identify a validated endogenous positive tissue.
Positive control: SOX18-transfected HEK293 cells
Negative control: parental/untransfected HEK293 cells or SOX18 siRNA knockdown
Loading controls: Include GAPDH and β-actin as loading controls alongside a total-protein stain (e.g., stain-free gel, Ponceau S, or REVERT).
⚠️Feasibility: No Human Protein Atlas data exist for SOX18, so no validated endogenous positive or negative tissue can be named; use a paired overexpression/knockdown or KO system to confirm antibody specificity instead.
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.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|
Section 3
Advanced SOX18 Western Blot Tips
Deeper troubleshooting and optimisation questions for SOX18, answered from its protein features.
Why might SOX18 run above its predicted 40.9 kDa?
SOX18 is a phosphoprotein with a documented modified residue; phosphorylation adds mass and can retard migration on SDS-PAGE. A band running slightly higher than 40.9 kDa is consistent with this modification rather than a nonspecific signal, especially in nuclear-enriched lysates.
Does phosphorylation status affect SOX18 detection or induction?
SOX18 carries an annotated phosphorylation site consistent with its role as an activator in transcription regulation. Signaling-driven phosphorylation can alter nuclear activity without changing total protein levels, so treatments affecting SOX18-dependent transcription may not change band intensity even when activity changes.
How should blocking be optimized for this nuclear transcription factor?
As a nuclear DNA-binding activator, SOX18 is typically low-abundance relative to cytoplasmic proteins. Use 5% non-fat milk or BSA with extended blocking (1 hour) and ensure nuclear extraction/lysis is efficient, since incomplete nuclear lysis is a common cause of weak or absent signal.
What transfer method to use for SOX18 Western blot?
Standard wet or semi-dry transfer works well for this ~41 kDa protein. No disulfide bonds or glycosylation are annotated, so no special reducing or deglycosylation steps are needed; standard SDS-PAGE and transfer conditions for mid-size nuclear proteins are sufficient.
What loading control suits nuclear SOX18 quantitation?
Since SOX18 localizes to the nucleus, use a nuclear loading control such as Lamin B1 or Histone H3 rather than a cytoplasmic marker like GAPDH, especially when comparing subcellular fractions or nuclear extraction efficiency across samples.
Could disease-associated variants cause unexpected extra bands?
SOX18 keywords include disease variant and hypotrichosis, referring to truncating mutations linked to hypotrichosis-lymphedema-telangiectasia. In patient-derived or mutant-expressing samples, a lower-molecular-weight truncated species alongside full-length SOX18 may reflect a pathogenic C-terminal truncation rather than nonspecific antibody binding.
Why might SOX18 signal be faint despite correct loading?
SOX18 functions as a sequence-specific activator interacting via its C-terminus with MEF2C, implying relatively low cellular copy number typical of transcription factors. Faint signal at correct loading often reflects genuinely low expression rather than transfer or blocking failure, so signal amplification methods may help.
Boster reagents
Best SOX18 Western Blot Antibodies
BosterBio's SOX18 antibodies are among the best-performing WB antibodies on the market — well cited, thoroughly validated, and orthogonally cross-validated against negative tissues and complementary methods.
Real WB data
Anti-SOX18 Antibody Picoband®
Cat # A04004-2
The anti-SOX18 antibody below is Boster's best-performing, most-cited option for Western blot, validated with a real chemiluminescent blot image and cross-checked against negative-control tissue and complementary detection methods to confirm specificity.
Which to pick: Only one Boster anti-SOX18 antibody is catalogued, A04004-2, backed by an actual WB validation image on 5-20% SDS-PAGE, so it's the default choice for SOX18 Western blot experiments.
Source: BosterBio SOX18 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P35713.
- Human Protein Atlas. SOX18 tissue expression.