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TSC22D3 / TSC22 domain family protein 3 · Western blot design guide
Design a Western Blot for TSC22D3
Source-linked TSC22D3 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-TSC22D3 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
TSC22D3 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 | ~14.8 kDa | |
| Gel | 15% (standard starting point) | |
| Positive control | Adrenal gland (IHC candidate; verify WB) +4 more | |
| Negative control | Adipose tissue (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Acetylated | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 3 isoform(s) |
Section 1
Source-Linked TSC22D3 Western Blot Protocol Options
The A03078 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human small intestine tissue lysate (catalog A03078) |
| Gel % | 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 | 0.2 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | A03078 · (A) 1 and (B) 2 μg/mL (catalog A03078) |
| 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 TSC22D3 Western Blot Band Size?
TSC22D3 is predicted at 14.8 kDa; isoforms and homodimerization could affect bands, but their migration effects are unproven and no empirical band is supplied.
| Band near 14.8 kDa | Consistent with the predicted size of TSC22D3; confirm its identity with controls |
| Band near 30 kDa | Could reflect a homodimer if the association survives sample preparation |
| Bands at different sizes | May reflect isoforms 1, 2, and 3; their migration differences are unknown |
| Weak cytoplasmic band during myoblast differentiation | Isoform 1 may relocate from cytoplasm to nucleus |
💡Expected TSC22D3 appearanceUniProt predicts 14.8 kDa for TSC22D3, but no empirical band size is supplied; confirm any candidate band with antibody specificity and appropriate sample controls.
| Predicted molecular mass | Sets a 14.8 kDa reference for the supplied sequence |
| Isoform 1 | Its migration relative to other isoforms is unknown |
| Isoforms 2 and 3 | Alternative splicing may change size, but their relative masses are unknown |
| Homodimer formation | Could yield a band near twice the monomer size if the dimer survives sample preparation |
| Situation | Likely cause | Next action |
|---|---|---|
| Band higher than expected | A homodimer may survive sample preparation | Compare denaturing conditions and verify band identity with an independent antibody |
| Band lower than expected | An alternative splice isoform may differ in size | Check isoform expression and verify the band with an independent antibody |
| Multiple bands | Isoforms 1, 2, and 3 may differ in migration | Compare isoform-specific expression or depletion controls |
| Weak or no signal | Isoform 1 localization may shift between cytoplasm and nucleus during myoblast differentiation | Compare nuclear and cytoplasmic fractions and include an expression-positive control |
| Fragments below expected size | Sample proteolysis may produce smaller immunoreactive fragments | Prepare fresh lysate with protease inhibitors and confirm identity with an independent antibody |
Experiment evidence
Western blot images showing TSC22D3 band
A03078 · TSC22D3
A03078-1 · TSC22D3
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for TSC22D3 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for TSC22D3 in Western blot, you can use adrenal gland tissue, which HPA scores High.
Positive control: Adrenal gland (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: HPA reports adipose tissue as not detected, though its negative signal should be verified in the tested lysate.
HPA tissue expression evidence for TSC22D3
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 → |
| Breast | glandular cells | High | Protein (IHC) | HPA → |
| Prostate | glandular cells | High | Protein (IHC) | HPA → |
| Skin | melanocytes | High | Protein (IHC) | HPA → |
| Tonsil | non-germinal center cells | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | adipocytes | Not detected | Protein (IHC) | HPA → |
| Cervix | glandular cells | Not detected | Protein (IHC) | HPA → |
| Duodenum | glandular cells | Not detected | Protein (IHC) | HPA → |
| Heart muscle | cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Lung | alveolar cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced TSC22D3 Western Blot Tips
Deeper troubleshooting and optimisation questions for TSC22D3, answered from its protein features.
What band size should I expect for TSC22D3?
Band shift · The canonical protein has a predicted mass of 14.8 kDa. No empirical apparent band size is supplied, so use 14.8 kDa as a reference, not a confirmed migration position. The listed features alone cannot explain any observed difference.
How might TSC22D3 isoforms affect band interpretation?
Isoforms · Three isoforms are listed. Isoform 3 lacks canonical residues 1–57, while isoform 2 replaces canonical residues 1–40 with a different sequence. These changes may affect migration, but the supplied features do not establish distinct visible bands.
Check the antibody epitope against each isoform’s sequence. An epitope within canonical residues 1–57 is absent from isoform 3; canonical residues 1–40 are replaced in isoform 2. Detection will depend on the actual epitope.
Check the antibody epitope against each isoform’s sequence. An epitope within canonical residues 1–57 is absent from isoform 3; canonical residues 1–40 are replaced in isoform 2. Detection will depend on the actual epitope.
Could TSC22D3 modifications cause a band shift?
PTM · The supplied UniProt features list N-acetylmethionine at position 1 and a position-102 entry labeled “Phosphoserine Phosphoserine Phosphoserine N-acetylmethionine.” These are UniProt coordinates; antibody or paper numbering may differ. The entries alone do not establish a visible shift or its direction.
Does this guide establish induction of TSC22D3?
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 TSC22D3?
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 A03078 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.
What should I check when quantifying TSC22D3 bands?
Quantitation · Define which band or bands the antibody detects before quantifying them. The three isoforms have different N-terminal sequences, and isoform 1 can occur in cytoplasm or nucleus. Keep the sampled fraction and differentiation state consistent across comparisons.
How should I compare cytoplasmic and nuclear TSC22D3 signals?
Interpretation · Isoform 1 is listed in both cytoplasm and nucleus, with localization depending on myoblast differentiation status by similarity. Compare matched differentiation states and account for the fraction analyzed when interpreting signal changes.
First check whether the antibody recognizes isoforms 1, 2, and 3, then compare the band with the canonical 14.8 kDa prediction. The listed modifications and potential homodimer formation are features to consider, but none establishes the identity or apparent mass of an unexpected band.
First check whether the antibody recognizes isoforms 1, 2, and 3, then compare the band with the canonical 14.8 kDa prediction. The listed modifications and potential homodimer formation are features to consider, but none establishes the identity or apparent mass of an unexpected band.
Boster reagents
TSC22D3 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-TSC22 domain family protein 3 TSC22D3 Antibody
Cat # A03078
Real WB data
Anti-TSC22 domain family protein 3 TSC22D3 Antibody
Cat # A03078-1
Two the supplier anti-TSC22D3 antibodies have WB images. A03078 was shown with human small intestine lysate at 1 and 2 μg/mL; A03078-1 was shown with 293T extracts at 1:1000. These captions document specific tested contexts, not broader validation.
Which to pick: For human small intestine lysate, consider A03078; it also lists mouse reactivity, though its supplied WB image uses human tissue. For 293T extracts, consider A03078-1, which lists human reactivity and has a WB image with reported conditions.
Source: BosterBio TSC22D3 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q99576.
- Human Protein Atlas. TSC22D3 tissue expression.
- PMC3598084 — target-verified WB comparison
- PMC7176855 — target-verified WB comparison
- PMC10415931 — target-verified WB comparison