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SPTBN1 / Spectrin beta chain, non-erythrocytic 1 · Western blot design guide
Design a Western Blot for SPTBN1
Source-linked SPTBN1 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-SPTBN1 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
SPTBN1 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 | ~274.6 kDa | |
| Gel | 4–12% gradient (standard starting point) | |
| 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 | Glycosylated + Phosphorylated | |
| Caveat | Peptide-blocking control | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 3 isoform(s) |
Section 1
Source-Linked SPTBN1 Western Blot Protocol Options
The A03164 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | the lysates from Jurkat cells (catalog A03164) |
| Gel % | 4–12% gradient (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Wet/tank transfer; optimize duration (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | A03164; 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 SPTBN1 Western Blot Band Size?
SPTBN1 is predicted at 274.6 kDa; annotated isoforms and Ser2324 O-GlcNAc may affect migration, but no empirical band or visible shift is established.
| Band near 275 kDa | consistent with the 274.6 kDa sequence prediction; confirm identity with antibody controls |
| Additional high-mass bands | could reflect Long, Short, or 2 isoforms, but their migration is unknown |
| Lower band alongside the main band | could reflect the Short isoform; identity requires confirmation |
| Weak band in a soluble lysate fraction | SPTBN1 may remain associated with the cytoskeleton or membrane |
💡Expected SPTBN1 appearanceSPTBN1 has a predicted mass of 274.6 kDa, but no empirical band size is supplied; use antibody specificity controls to identify any band near that size.
| UniProt predicted mass | sets a 274.6 kDa sequence-based reference, not a measured migration |
| O-linked GlcNAc at Ser2324 | adds one monosaccharide; a visible shift is not established |
| Long isoform | may differ in size from Short; its migration is not supplied |
| Short isoform | may be smaller than Long; its migration is not supplied |
| Isoform 2 | may differ in size; its migration is not supplied |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | cytoskeletal or membrane association may limit recovery | check extraction and analyze the insoluble fraction |
| Band higher than expected | an isoform could differ from the sequence-based reference | confirm identity with peptide competition or a second antibody |
| Band lower than expected | the Short isoform is possible, but its migration is unknown | confirm identity with an antibody recognizing another region |
| Multiple bands | Long, Short, and 2 isoforms are annotated | check which bands respond to peptide competition or SPTBN1 depletion |
| Weak or no signal | cytoskeletal association may reduce recovery in the sampled fraction | compare soluble and insoluble fractions and check sample loading |
| Fragments below expected size | breakdown of this large protein is possible but unestablished | compare promptly prepared samples and verify fragment identity |
Experiment evidence
Western blot images showing SPTBN1 band
A03164 · SPTBN1
A03164 · SPTBN1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for SPTBN1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for SPTBN1 in Western blot, you can use adrenal gland lysate, 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 alongside the samples.
⚠️Feasibility: No tissue is listed as not detected, so use siRNA knockdown or a KO line for a negative control.
HPA tissue expression evidence for SPTBN1
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 → |
| Appendix | glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | ciliated cells (cell body) | High | Protein (IHC) | HPA → |
| Cerebral cortex | endothelial cells | High | Protein (IHC) | HPA → |
| Colon | endothelial cells | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Bone marrow | hematopoietic cells | Low | Protein (IHC) | HPA → |
| Caudate | glial cells | Low | Protein (IHC) | HPA → |
| Cervix | glandular cells | Low | Protein (IHC) | HPA → |
| Esophagus | squamous epithelial cells | Low | Protein (IHC) | HPA → |
| Hippocampus | glial cells | Low | Protein (IHC) | HPA → |
Section 3
Advanced SPTBN1 Western Blot Tips
Deeper troubleshooting and optimisation questions for SPTBN1, answered from its protein features.
Where should the full-length SPTBN1 band appear?
Band shift · The canonical sequence is 2,364 residues with a predicted mass of 274.6 kDa. Use that as a reference, but no observed band position is supplied. The listed modifications alone do not establish a visible shift or explain any difference from the predicted mass.
Could SPTBN1 isoforms produce different bands?
Isoforms · Yes. UniProt lists Long, Short, and isoform 2. Relative to the canonical sequence, Short replaces residues 2141–2168 and lacks 2169–2364. Isoform 2 replaces residues 1–49 and 2141–2225, and lacks 2226–2364. Check whether the antibody recognizes a region retained in the isoform being measured.
Which phosphorylation sites matter when interpreting SPTBN1 bands?
PTM · UniProt lists numerous sites, including Ser36, Tyr1805, and a cluster at residues 2102–2195. These are canonical UniProt coordinates; antibody or paper numbering may differ. Some cluster residues are replaced or absent in shorter isoforms. Phosphorylation alone does not establish a resolvable band shift.
Yes. UniProt lists O-linked GlcNAc on Ser2324 in canonical numbering. That residue is absent from Short and isoform 2 because their C-terminal sequences are deleted. A listed site does not establish a visible mobility shift; account for isoform identity when interpreting a band.
Yes. UniProt lists O-linked GlcNAc on Ser2324 in canonical numbering. That residue is absent from Short and isoform 2 because their C-terminal sequences are deleted. A listed site does not establish a visible mobility shift; account for isoform identity when interpreting a band.
Does this guide establish induction of SPTBN1?
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 SPTBN1 Western blot?
Transfer · SPTBN1 has a predicted canonical mass of 274.6 kDa. Choose and check transfer conditions suitable for a protein of that size, including whether protein remains in the gel after transfer. The supplied features do not establish one specific transfer method.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A03164 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 SPTBN1 bands be quantified across samples?
Quantitation · Compare the same assigned band and antibody-recognized region across samples. The three listed isoforms differ at the N terminus or C terminus, so an antibody may measure different isoforms differently. If measuring a modification, distinguish its signal from total SPTBN1; a listed site does not establish its occupancy in your samples.
What could an unexpected SPTBN1 band represent?
Interpretation · First check whether its size and antibody epitope are consistent with Short or isoform 2, which have substantial sequence replacements and deletions. Compare it with the 274.6 kDa canonical prediction. The supplied features do not provide an observed band position or establish that any modification causes the band.
Boster reagents
SPTBN1 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-SPTBN1 Antibody
Cat # A03164
The catalog reports one anti-SPTBN1 antibody for Western blotting: A03164. It is catalogued as reactive with human, mouse, and rat, and its Western blot image uses Jurkat cell lysate. The supplied evidence does not establish performance across all specimens.
Which to pick: A03164 is the only listed option. Its Western blot image uses Jurkat cell lysate; check that this tested context and the catalogued human, mouse, and rat reactivity fit your experiment.
Source: BosterBio SPTBN1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q01082.
- Human Protein Atlas. SPTBN1 tissue expression.
- PMC9667201 — target-verified WB comparison
- PMC12369245 — target-verified WB comparison
- PMC8864474 — target-verified WB comparison