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- Table of Contents
ARNT / Aryl hydrocarbon receptor nuclear translocator · Western blot design guide
Design a Western Blot for ARNT
Real validated ARNT Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-ARNT 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
ARNT 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 | ~86.6 kDa | |
| Observed band | ~87 kDa | |
| Gel | 10% (catalog M02263) | |
| Positive control | Endometrium (IHC candidate; verify WB) +4 more | |
| Negative control | Cerebellum (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Acetylated | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 4 isoform(s) |
Section 1
Real Curated ARNT Western Blot Protocols
The M02263 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human THP-1, human K562, human MCF-7 (catalog M02263) |
| Gel % | 10% (catalog M02263) |
| Load | 30 ug; reducing conditions (catalog M02263) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog M02263) |
| Membrane | nitrocellulose membrane (catalog M02263) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog M02263) |
| Primary antibody | M02263 · 1:500 (catalog M02263) |
| Primary incubation | overnight at 4°C (catalog M02263) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog M02263) |
| Secondary incubation | 1.5 hour at RT (catalog M02263) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog M02263) |
| Detection | ECL (catalog M02263) |
Section 2
What Is the Expected ARNT Western Blot Band Size?
ARNT is predicted at 86.6 kDa and observed near 87 kDa; the small difference has no established cause in the supplied evidence.
| Single band near 87 kDa | Matches the observed ARNT band and its 86.6 kDa predicted mass |
| Additional discrete bands | Could reflect isoforms 1, 2, 3 or 4; their migration is not established |
| Higher band in a native DNA-bound sample | Could reflect DNA-dependent ARNT homodimerization; persistence on SDS-PAGE is unproven |
| Stronger band in a nuclear fraction | Consistent with ARNT's nuclear localization |
💡Expected ARNT appearanceARNT has a predicted mass of 86.6 kDa and an empirical band near 87 kDa; confirm band identity with an appropriate specificity control, since the supplied features do not establish distinct isoform bands.
| Predicted ARNT mass | 86.6 kDa, consistent with the observed band near 87 kDa |
| Splice isoforms 1 and 2 | Their individual masses and migration relative to each other are unspecified |
| Splice isoforms 3 and 4 | Their individual masses and migration relative to each other are unspecified |
| DNA-dependent homodimer | May increase complex size under native conditions; a higher SDS-PAGE band is not established |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Nuclear ARNT may be underrepresented in the sample | Check a nuclear fraction and confirm sample loading |
| Band higher than expected | Identity of the higher species is unestablished; ARNT can homodimerize on DNA | Compare fully denatured samples and use an ARNT specificity control |
| Band lower than expected | An alternative isoform or degradation is possible, but neither band size is established | Check sample integrity and verify the band with an ARNT specificity control |
| Multiple bands | Four annotated isoforms are possible, but their migration is unknown | Confirm each band with an ARNT specificity control |
| Weak or no signal | Nuclear localization can reduce signal in an unfractionated sample | Enrich the nuclear fraction and verify loading |
| Fragments below expected size | Sample degradation is possible; no cleavage product is annotated | Prepare fresh lysate with protease inhibitors and confirm fragment identity |
Experiment evidence
Western blot images showing ARNT band
M02263 · ARNT
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for ARNT Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for ARNT in Western blot, you can use endometrium, which HPA scores High.
Positive control: Endometrium (IHC candidate; verify WB)
Negative control: Cerebellum (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: ARNT is nuclear, so nuclear enrichment may improve detection in the positive sample.
HPA tissue expression evidence for ARNT
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 |
|---|---|---|---|---|
| Endometrium | cells in endometrial stroma | High | Protein (IHC) | HPA → |
| Fallopian tube | glandular cells | High | Protein (IHC) | HPA → |
| Lung | alveolar cells type I | High | Protein (IHC) | HPA → |
| Placenta | decidual cells | High | Protein (IHC) | HPA → |
| Skin | fibrohistiocytic cells | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Cerebellum | cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Heart muscle | cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | Low | Protein (IHC) | HPA → |
| Epididymis | glandular cells | Low | Protein (IHC) | HPA → |
| Liver | cholangiocytes | Low | Protein (IHC) | HPA → |
Section 3
Advanced ARNT Western Blot Tips
Deeper troubleshooting and optimisation questions for ARNT, answered from its protein features.
How should ARNT 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.
Could ARNT isoforms produce different bands?
Isoforms · Four isoforms are listed. Relative to the canonical sequence, isoform 2 lacks residues 77–91, isoform 3 lacks 1–9 and 319–323, and isoform 4 lacks 601–602. These deletions could affect band position, but the features do not establish which isoforms are expressed or resolve as separate bands.
Check the antibody's mapped epitope against the listed deletions: 77–91 in isoform 2, 1–9 and 319–323 in isoform 3, and 601–602 in isoform 4. An epitope within a deleted segment may prevent detection of that isoform. These are canonical UniProt coordinates; antibody numbering may differ.
Check the antibody's mapped epitope against the listed deletions: 77–91 in isoform 2, 1–9 and 319–323 in isoform 3, and 601–602 in isoform 4. An epitope within a deleted segment may prevent detection of that isoform. These are canonical UniProt coordinates; antibody numbering may differ.
What matters when probing ARNT phosphorylation?
PTM · UniProt lists phosphoserine at canonical residue 77. Isoform 2 lacks residues 77–91, so a reagent targeting that site would not detect the corresponding site in isoform 2. Check that the antibody's residue numbering uses the same sequence convention; the feature does not establish a visible band shift.
UniProt lists N-acetylalanine at canonical residue 2. Isoform 3 lacks canonical residues 1–9, so this coordinate does not carry over directly to that isoform. The acetylation feature alone does not explain an unexpected band position.
UniProt lists N-acetylalanine at canonical residue 2. Isoform 3 lacks canonical residues 1–9, so this coordinate does not carry over directly to that isoform. The acetylation feature alone does not explain an unexpected band position.
Does this guide establish induction of ARNT?
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 ARNT?
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 M02263 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 multiple ARNT bands be quantified?
Quantitation · Specify which bands are measured and use the same selection across samples. The four listed isoforms have sequence differences, so an assay may detect them differently depending on its antibody epitope. Band position alone does not identify an isoform or establish that phosphorylation caused a shift.
How does ARNT's observed band compare with its predicted mass?
Interpretation · ARNT's predicted mass is 86.6 kDa, and the reported band is approximately 87 kDa. These values are close. The listed modifications alone do not establish a visible shift or explain any small difference between calculated and apparent mass.
Compare each band's position with the approximately 87 kDa reported band, then consider the listed isoform deletions and antibody epitope. Canonical residues 2 and 77 are modified, but their presence alone does not explain a shifted band. The supplied features cannot assign an unexpected band to a particular isoform or modification.
Compare each band's position with the approximately 87 kDa reported band, then consider the listed isoform deletions and antibody epitope. Canonical residues 2 and 77 are modified, but their presence alone does not explain a shifted band. The supplied features cannot assign an unexpected band to a particular isoform or modification.
Boster reagents
ARNT 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-HIF-1 beta ARNT Rabbit Monoclonal Antibody
Cat # M02263
The catalog reports one anti-ARNT rabbit monoclonal antibody, M02263, with human reactivity. Its WB image reports an approximately 87 kDa band in seven human cell-line lysates. No publication or orthogonal validation evidence is supplied.
Which to pick: M02263 is the only listed option. For human WB, it has a validation image using THP-1, K562, MCF-7, SIHA, HeLa, Jurkat, and RT4 whole-cell lysates at 1:500. These examples do not establish performance in every human sample.
Source: BosterBio ARNT gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P27540.
- Human Protein Atlas. ARNT tissue expression.
- PMC6338330 — target-verified WB comparison
- PMC11133443 — target-verified WB comparison
- PMC7373995 — target-verified WB comparison