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NR1D2 / Nuclear receptor subfamily 1 group D member 2 · Western blot design guide
Design a Western Blot for NR1D2
Source-linked NR1D2 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-NR1D2 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
NR1D2 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 | ~64.6 kDa | |
| Observed band | ~68 kDa | |
| Gel | 5–20% (catalog A04958-2) | |
| Positive control | Fallopian tube (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 | Modification-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked NR1D2 Western Blot Protocol Options
The A04958-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human Hela, human Jurkat, human MCF-7, human HepG2, rat liver, rat brain, mouse liver, mouse brain (catalog A04958-2) |
| Gel % | 5–20% (catalog A04958-2) |
| Load | 30 ug; reducing conditions (catalog A04958-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A04958-2) |
| Membrane | nitrocellulose membrane (catalog A04958-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A04958-2) |
| Primary antibody | A04958-2 · 0.5 μg/mL (catalog A04958-2) |
| Primary incubation | overnight at 4°C (catalog A04958-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A04958-2) |
| Secondary incubation | 1.5 hour at RT (catalog A04958-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A04958-2) |
| Detection | ECL (catalog A04958-2) |
Section 2
What Is the Expected NR1D2 Western Blot Band Size?
NR1D2 is predicted at 64.6 kDa and observed near 68 kDa on reducing Western blots; the cause of the difference is not established.
| Band near 68 kDa | Empirical NR1D2 band in reducing whole-cell lysates |
| Band near 65 kDa | Close to the 64.6 kDa predicted monomer mass; confirm identity with controls |
| Higher band that decreases with stronger reduction | Could reflect retained NR1D2 homodimers; their persistence on SDS-PAGE is unestablished |
| Weak whole-cell band with stronger nuclear signal | Consistent with NR1D2 nuclear localization |
💡Expected NR1D2 appearanceNR1D2 has a predicted mass of 64.6 kDa and an empirical band near 68 kDa in reducing lysates; the difference is unexplained, so confirm identity with antibody and depletion controls.
| Predicted monomer mass | 64.6 kDa calculated; the empirical band is near 68 kDa |
| Monomeric DNA binding | Supports a monomer-sized species without establishing its migration |
| Homodimeric DNA binding | Could yield a higher band if dimers survive sample preparation; this is unproven |
| Intrachain disulfides at residues 337–343 and 374–384 | May affect conformation, but no apparent size change is established |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | NR1D2 may be enriched in the nucleus | Check a nuclear fraction and include a validated positive lysate |
| Band higher than expected | A retained homodimer is possible, but unconfirmed | Compare reducing conditions and confirm identity by NR1D2 depletion |
| Band lower than expected | Protein degradation or an unrelated signal is possible | Use fresh lysate with protease inhibitors and check NR1D2 depletion |
| Multiple bands | Additional signals have no supported isoform explanation | Check which band disappears after NR1D2 depletion |
| Weak or no signal | Whole-cell lysate may dilute nuclear NR1D2 | Enrich nuclei and verify sample loading |
| Fragments below expected size | Sample degradation is possible | Prepare fresh lysate with protease inhibitors and confirm fragment identity |
Experiment evidence
Western blot images showing NR1D2 band
A04958-2 · NR1D2
A04958-1 · NR1D2
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for NR1D2 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for NR1D2 in Western blot, you can use fallopian tube tissue, which has high HPA expression.
Positive control: Fallopian tube (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: NR1D2 is nuclear and can also be cytoplasmic, so nuclear enrichment may improve detection.
HPA tissue expression evidence for NR1D2
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 |
|---|---|---|---|---|
| Fallopian tube | ciliated cells (ciliary rootlets) | High | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Medium | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | Medium | Protein (IHC) | HPA → |
| Bronchus | ciliated cells (cell body) | Medium | Protein (IHC) | HPA → |
| Cerebellum | Purkinje cells | Medium | 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 → |
| Hippocampus | glial cells | Not detected | Protein (IHC) | HPA → |
| Liver | cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Ovary | follicle cells | Not detected | Protein (IHC) | HPA → |
| Skeletal muscle | myocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced NR1D2 Western Blot Tips
Deeper troubleshooting and optimisation questions for NR1D2, answered from its protein features.
How should NR1D2 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.
Are multiple NR1D2 bands explained by listed isoforms?
Isoforms · UniProt supplies one isoform and no alternative sequence. The listed features therefore do not establish an isoform explanation for multiple bands. Verify the identity of each band before assigning it to NR1D2.
Which NR1D2 modifications are relevant when interpreting bands?
PTM · UniProt lists phosphoserine at position 46 by GSK3-beta and N6-acetyllysine at positions 162 and 163 by KAT5. These are UniProt sequence coordinates; antibody or paper numbering may differ. Their presence does not establish that a visible band shift will occur.
Does this guide establish induction of NR1D2?
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 NR1D2?
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 A04958-2 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 NR1D2 be quantified across sample preparations?
Quantitation · NR1D2 is listed in both nucleus and cytoplasm. Keep the sampled compartment consistent across comparisons, or measure matched fractions separately, so changes in their proportions do not confound band intensity. Confirm the quantified band’s identity, particularly if more than one band appears.
Why might NR1D2 appear near 68 kDa rather than 64.6 kDa?
Interpretation · The reported apparent band is about 68 kDa, while the predicted mass is 64.6 kDa. UniProt lists phosphorylation and acetylation sites, but their presence alone does not establish the cause of this difference. Use the 68 kDa observation as a reference and verify band identity experimentally.
UniProt lists NR1D2 in the nucleus and cytoplasm. Compare matched whole-cell and fractionated samples if signal varies with preparation. Its note that CSNK1E phosphorylation enhances cytoplasmic localization is transferred from another species, so do not assume the same effect has been demonstrated for this protein.
Compare them with the reported approximately 68 kDa band and the 64.6 kDa predicted mass. UniProt lists one isoform, no signal peptide or propeptide, and no glycosylation sites. Its modification sites alone cannot identify an unexpected band or prove a mobility shift; verify band identity experimentally.
UniProt lists NR1D2 in the nucleus and cytoplasm. Compare matched whole-cell and fractionated samples if signal varies with preparation. Its note that CSNK1E phosphorylation enhances cytoplasmic localization is transferred from another species, so do not assume the same effect has been demonstrated for this protein.
Compare them with the reported approximately 68 kDa band and the 64.6 kDa predicted mass. UniProt lists one isoform, no signal peptide or propeptide, and no glycosylation sites. Its modification sites alone cannot identify an unexpected band or prove a mobility shift; verify band identity experimentally.
Boster reagents
NR1D2 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-NR1D2 Antibody Picoband®
Cat # A04958-2
Real WB data
Anti-NR1D2 Antibody (Center)
Cat # A04958-1
Both listed anti-NR1D2 antibodies have Western blot images. A04958-2 shows a band near 68 kDa in the stated human cell and rodent tissue lysates; A04958-1 shows detection in HeLa lysate. These images document the reported conditions, without independent target validation.
Which to pick: Choose A04958-2 for the reported human, mouse, or rat reactivity and its eight-lane blot covering human cell lines and rodent tissues. For human HeLa lysate, A04958-1 also has a Western blot image. Match your sample and conditions to the reported evidence.
Source: BosterBio NR1D2 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q14995.
- Human Protein Atlas. NR1D2 tissue expression.
- PMC11896154 — target-verified WB comparison
- PMC7217318 — target-verified WB comparison
- PMC7810220 — target-verified WB comparison