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- Table of Contents
NOP56 / Nucleolar protein 56 · Western blot design guide
Design a Western Blot for NOP56
Source-linked NOP56 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-NOP56 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
NOP56 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 | ~66.1 kDa | |
| Gel | 12–15% (standard starting point) | |
| Positive control | Testis (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 | Phosphorylated + Acetylated | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked NOP56 Western Blot Protocol Options
The A04335 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | HEPG2 Hela cells (catalog A04335) |
| 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 | A04335 · 1:1000 (catalog A04335) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | 1:20000 (catalog A04335) |
| 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 NOP56 Western Blot Band Size?
NOP56 is predicted at 66.1 kDa; listed modifications could affect migration, but no empirical band or visible shift is established.
| Band near 66.1 kDa | Consistent with predicted NOP56 size; identity needs confirmation |
| Closely spaced bands near 66.1 kDa | Could reflect modified NOP56, but distinct migration is unverified |
| Band above 66.1 kDa | Could reflect modified NOP56; confirm its identity |
| Weak band in whole-cell lysate | Nucleolar NOP56 may be poorly recovered during extraction |
💡Expected NOP56 appearanceNOP56 has a predicted mass of 66.1 kDa and no supplied empirical band size; confirm a candidate band near that size with a NOP56 depletion control.
| UniProt predicted mass | Places unmodified NOP56 near 66.1 kDa |
| Phosphoserine at residue 314 | May affect migration; a visible shift is unestablished |
| Phosphorylation at residues 466 to 468 | May affect migration; distinct bands are unestablished |
| Omega-N-methylarginine at residue 359 | Its effect on apparent band size is unestablished |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Nucleolar NOP56 may be poorly recovered | Check nuclear extraction and loading controls |
| Band higher than expected | Modification or unrelated antibody binding is possible | Test whether the band decreases after NOP56 depletion |
| Band lower than expected | Degradation or unrelated antibody binding is possible | Use protease inhibitors and a NOP56 depletion control |
| Multiple bands | Modified forms are possible, but distinct NOP56 bands are unverified | Identify bands that decrease after NOP56 depletion |
| Weak or no signal | Nucleolar protein recovery may be limited | Check extraction efficiency and antibody performance |
Experiment evidence
Western blot images showing NOP56 band
A04335 · NOP56
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for NOP56 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for NOP56 in Western blot, you can use testis lysate, which has the highest HPA signal among the supplied tissues.
Positive control: Testis (IHC candidate; verify WB)
Negative control: Suggested KO / knockdown lysate
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: No Not-detected tissue is listed, so a knockdown or KO line is the clearest negative control.
HPA tissue expression evidence for NOP56
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 |
|---|---|---|---|---|
| Testis | Leydig cells | High | Protein (IHC) | HPA → |
| Appendix | glandular cells | Medium | Protein (IHC) | HPA → |
| Breast | glandular cells | Medium | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | Medium | Protein (IHC) | HPA → |
| Caudate | glial cells | Medium | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | adipocytes | Low | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Low | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | Low | Protein (IHC) | HPA → |
| Cervix | glandular cells | Low | Protein (IHC) | HPA → |
| Colon | endothelial cells | Low | Protein (IHC) | HPA → |
Section 3
Advanced NOP56 Western Blot Tips
Deeper troubleshooting and optimisation questions for NOP56, answered from its protein features.
Where should the main NOP56 band appear?
Band shift · NOP56 has a predicted mass of 66.1 kDa. No observed Western blot band is supplied, so use 66.1 kDa as a reference rather than an established apparent mass.
Could different NOP56 bands represent isoforms?
Isoforms · The supplied record lists one isoform and no alternative sequence. Do not assign additional bands to NOP56 isoforms on this evidence alone.
Which phosphorylation sites matter when interpreting NOP56 bands?
PTM · UniProt lists phosphoserine at 314, 466, 467, 511, 519, 520, 537, 563, 569, 570, 579 and 581, and phosphothreonine at 468. These are UniProt coordinates; antibody or paper numbering may differ. The sites alone do not establish a visible shift.
UniProt lists omega-N-methylarginine at 359 and N6-acetyllysine at 561. These are UniProt coordinates. Their presence does not establish a visible band shift or explain an apparent mass difference.
UniProt lists omega-N-methylarginine at 359 and N6-acetyllysine at 561. These are UniProt coordinates. Their presence does not establish a visible band shift or explain an apparent mass difference.
Does this guide establish induction of NOP56?
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 NOP56?
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 A04335 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 NOP56 signal be quantified across samples?
Quantitation · Compare the same band and sample fraction across lanes. NOP56 occurs in a large pre-ribosomal ribonucleoprotein complex and in multiple listed compartments, so changes in fraction or extraction can complicate interpretation of signal differences.
Which sample fractions could contain NOP56?
Interpretation · NOP56 is listed in the nucleolus and nucleoplasm, with cytoplasmic localization also reported. Keep the fraction consistent across samples when comparing bands or signal levels.
Compare them with the 66.1 kDa predicted mass and the expected band pattern for the assay. The supplied features list modifications and one isoform, but provide no observed band mass; they cannot identify an unexpected band or establish its cause.
Compare them with the 66.1 kDa predicted mass and the expected band pattern for the assay. The supplied features list modifications and one isoform, but provide no observed band mass; they cannot identify an unexpected band or establish its cause.
Boster reagents
NOP56 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-NOP56 Antibody
Cat # A04335
A04335 is an anti-NOP56 polyclonal antibody listed for human, mouse, and rat. Its WB image shows HepG2 and HeLa cell samples with primary antibody at 1:1000 and secondary antibody at 1:20000; no mouse or rat WB example is supplied.
Which to pick: A04335 is the only listed option. It has a WB image using human cell samples; mouse and rat reactivity is listed, but those species are not shown in the supplied WB evidence.
Source: BosterBio NOP56 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry O00567.
- Human Protein Atlas. NOP56 tissue expression.
- PMC12984107 — target-verified WB comparison
- PMC8162121 — target-verified WB comparison