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- Table of Contents
DIO2 / Type II iodothyronine deiodinase · Western blot design guide
Design a Western Blot for DIO2
Real validated DIO2 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-DIO2 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
DIO2 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 | ~30.6 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 | Ubl conjugation | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 3 isoform(s) |
Section 1
Real Curated DIO2 Western Blot Protocols
The A02249-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | lysates from human heart and human skeletal muscle tissue lysate (from left to right), (catalog A02249-1) |
| Gel % | 12–15% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Standard semi-dry transfer; verify efficiency (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | A02249-1; use the WB datasheet starting dilution (standard starting point) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | goat anti-rabbit IgG, 1:10000 (catalog A02249-1) |
| 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 DIO2 Western Blot Band Size?
DIO2 has a predicted 30.6 kDa monomer; possible isoforms and homodimers could affect bands, but no empirical migration is supplied.
| Band near 30.6 kDa | consistent with the predicted DIO2 monomer; confirm identity with controls |
| Band near twice the monomer size | could reflect a retained DIO2 homodimer; its migration on this blot is unestablished |
| Several bands at different positions | isoforms 1, 2, and 3 are possible contributors, but distinct migration is unestablished |
| Little signal in a soluble fraction | DIO2 is an endoplasmic reticulum membrane protein |
💡Expected DIO2 appearanceUniProt predicts a 30.6 kDa DIO2 monomer; no empirical band size is supplied, so confirm any candidate band with membrane recovery and band-identity controls.
| Predicted DIO2 monomer mass | sets a 30.6 kDa reference, not a measured band position |
| Possible DIO2 homodimer | could yield a higher band if the complex persists during electrophoresis |
| Splice isoform 1 | has no supplied isoform-specific mass or migration position |
| Splice isoform 2 | may differ in size from other isoforms; its relative position is unknown |
| Splice isoform 3 | may differ in size from other isoforms; its relative position is unknown |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | incomplete recovery of membrane-bound DIO2 | check detergent extraction and test a membrane-enriched fraction |
| Band higher than expected | possible retained DIO2 homodimer | compare fully denatured samples and verify the band with DIO2 depletion |
| Band lower than expected | an alternative isoform is possible, but its migration is unknown | verify identity with DIO2 depletion and an independent antibody |
| Multiple bands | isoforms 1, 2, and 3 are possible contributors | compare isoform-specific controls and DIO2 depletion |
| Weak or no signal | limited recovery of endoplasmic reticulum membrane protein | check membrane extraction and sample loading |
Experiment evidence
Western blot images showing DIO2 band
A02249-1 · DIO2
A02249-1 · DIO2
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for DIO2 Western blot
🧪For positive controls for DIO2 in Western blot, you can use an HPA-confirmed positive tissue once one is identified; none is supplied here.
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.
⚠️Feasibility: HPA supplies no tissue controls, and DIO2's ER membrane location may require effective solubilization.
HPA tissue expression evidence for DIO2
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 DIO2 Western Blot Tips
Deeper troubleshooting and optimisation questions for DIO2, answered from its protein features.
How should DIO2 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.
How could DIO2 isoforms affect band interpretation?
Isoforms · UniProt lists three isoforms. Isoform 3 lacks canonical residues 1–53 and 147–273 and has an alternative sequence at 75–146. Isoform 2 has an alternative sequence beginning at canonical residue 74. These are UniProt canonical coordinates; antibody or paper numbering may differ. Check whether the antibody’s epitope is retained in each isoform.
Does UniProt identify a DIO2 modification site causing a shift?
PTM · The supplied record lists ubiquitin-like conjugation as a keyword but provides no modified-residue coordinates. Conjugation could be relevant to additional bands, but the feature does not establish that any band is modified or how far it would migrate.
Does this guide establish induction of DIO2?
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 DIO2?
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 A02249-1 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 be quantified when several DIO2 bands appear?
Quantitation · Define the band or bands used for quantitation and apply that choice consistently. The record lists three isoforms and possible homodimers, so a single band may not represent every DIO2 form. The supplied features cannot identify an unexpected band without additional evidence.
Why might DIO2 migrate differently from 30.6 kDa?
Interpretation · 30.6 kDa is the predicted mass; no observed band is supplied. UniProt lists alternative isoforms and ubiquitin-like conjugation, but these features alone do not establish a visible shift or explain a measured mass difference. Compare the band with a molecular-weight marker and identify which isoform the antibody recognizes.
DIO2 is an endoplasmic-reticulum, single-pass membrane protein. Include a membrane-containing fraction when preparing samples, and check whether DIO2 remains in any material discarded during extraction. A weak band could reflect sample recovery.
DIO2 is predominantly monomeric but can form homodimers; dimerization is not essential for enzyme activity. A higher band is therefore compatible with, but does not establish, a dimer. Compare it with the monomer band before assigning its identity.
DIO2 is an endoplasmic-reticulum, single-pass membrane protein. Include a membrane-containing fraction when preparing samples, and check whether DIO2 remains in any material discarded during extraction. A weak band could reflect sample recovery.
DIO2 is predominantly monomeric but can form homodimers; dimerization is not essential for enzyme activity. A higher band is therefore compatible with, but does not establish, a dimer. Compare it with the monomer band before assigning its identity.
Boster reagents
DIO2 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-DIO2 Antibody (Center)
Cat # A02249-1
A02249-1 is listed for human and mouse DIO2. Its Western blot image shows human heart and skeletal muscle lysates at 35 µg per lane, with the primary antibody diluted 1:1000. The supplied evidence does not show a mouse blot.
Which to pick: A02249-1 is the only listed option. It has a Western blot image from human heart and skeletal muscle; mouse reactivity is listed, but no mouse blot is supplied.
Source: BosterBio DIO2 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q92813.
- Human Protein Atlas. DIO2 tissue expression.
- PMC8913869 — target-verified WB comparison
- PMC6031622 — target-verified WB comparison
- PMC8584701 — target-verified WB comparison