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- Table of Contents
NAT10 / RNA cytidine acetyltransferase · Western blot design guide
Design a Western Blot for NAT10
Real validated NAT10 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-NAT10 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
NAT10 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 | ~115.7 kDa | |
| Observed band | ~120 kDa | |
| Gel | 5–20% (catalog A06226-2) | |
| Positive control | Cerebellum (IHC candidate; verify WB) +4 more | |
| Negative control | Appendix (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 | 2 isoform(s) |
Section 1
Real Curated NAT10 Western Blot Protocols
The A06226-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human A549, human Hela, human K562 (catalog A06226-2) |
| Gel % | 5–20% (catalog A06226-2) |
| Load | 30 ug; reducing conditions (catalog A06226-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A06226-2) |
| Membrane | nitrocellulose membrane (catalog A06226-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A06226-2) |
| Primary antibody | A06226-2 · 0.5 μg/mL (catalog A06226-2) |
| Primary incubation | overnight at 4°C (catalog A06226-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A06226-2) |
| Secondary incubation | 1.5 hour at RT (catalog A06226-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A06226-2) |
| Detection | ECL (catalog A06226-2) |
Section 2
What Is the Expected NAT10 Western Blot Band Size?
NAT10 is predicted at 115.7 kDa and observed near 120 kDa; the cause of the difference is not established.
| Band near 120 kDa | Empirically observed NAT10 band in whole-cell lysate |
| Band near 115.7 kDa | Consistent with the predicted NAT10 mass; confirm its identity |
| Two bands near the main band | Isoforms 1 and 2 are possible contributors, but distinct migration is unverified |
| One sharp band | Isoforms 1 and 2 need not appear as separate bands |
💡Expected NAT10 appearanceNAT10 has a predicted mass of 115.7 kDa and an empirical band near 120 kDa in whole-cell lysate; the difference is unexplained, so confirm band identity with appropriate controls.
| Predicted NAT10 mass | 115.7 kDa is the sequence-based reference; the observed band is near 120 kDa |
| Splice isoforms 1 and 2 | Their relative masses and whether they resolve as separate bands are unknown |
| N6-acetyllysine at residue 426 | No visible migration effect is established |
| Phosphothreonine at residue 716 and phosphoserines at residues 934, 984, and 987 | Modification-dependent migration is possible but unproven |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Nuclear NAT10 may be poorly recovered during lysate preparation | Check nuclear recovery and compare a nuclear-enriched fraction |
| Band higher than expected | The approximately 120 kDa empirical band lies above the 115.7 kDa prediction; the cause is unknown | Compare with the documented band and verify identity by NAT10 depletion |
| Band lower than expected | A lower band has no established NAT10 assignment from the supplied features | Check sample integrity and test whether NAT10 depletion removes the band |
| Multiple bands | Isoforms 1 and 2 exist, but their band positions are unknown | Use NAT10 depletion or an independent antibody to identify NAT10-specific bands |
| Weak or no signal | Recovery of nuclear NAT10 may be insufficient | Check loading and transfer, then compare nuclear-enriched material |
Experiment evidence
Western blot images showing NAT10 band
A06226-2 · NAT10
A06226-2 · NAT10
M06226 · NAT10
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for NAT10 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for NAT10 in Western blot, you can use cerebellum tissue, which HPA rates as highly positive.
Positive control: Cerebellum (IHC candidate; verify WB)
Negative control: Appendix (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: NAT10 is nucleolar, and HPA lists appendix as a feasible negative tissue.
HPA tissue expression evidence for NAT10
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 |
|---|---|---|---|---|
| Cerebellum | Purkinje cells | High | Protein (IHC) | HPA → |
| Cerebral cortex | neuronal cells | High | Protein (IHC) | HPA → |
| Hippocampus | neuronal cells | High | Protein (IHC) | HPA → |
| Caudate | neuronal cells | Medium | Protein (IHC) | HPA → |
| Cervix | squamous epithelial cells | Medium | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Appendix | glandular cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Colon | endothelial cells | Not detected | Protein (IHC) | HPA → |
| Endometrium | cells in endometrial stroma | Not detected | Protein (IHC) | HPA → |
| Heart muscle | cardiomyocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced NAT10 Western Blot Tips
Deeper troubleshooting and optimisation questions for NAT10, answered from its protein features.
How should NAT10 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 NAT10 isoforms produce separate bands?
Isoforms · UniProt lists isoforms 1 and 2; isoform 2 lacks residues 1–72 of isoform 1. Check whether your antibody recognizes a sequence retained in isoform 2 before assigning a second band to it. The feature does not establish where either isoform migrates.
Which NAT10 modifications are relevant to band interpretation?
PTM · UniProt lists N6-acetyllysine at 426, phosphothreonine at 716, and phosphoserines at 934, 984, and 987. These are UniProt coordinates; paper or antibody numbering may differ. Their presence alone does not demonstrate a visible band shift.
Does this guide establish induction of NAT10?
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.
What transfer method to use for NAT10 Western blot?
Transfer · NAT10 is predicted at 115.7 kDa, so choose transfer conditions suited to a protein around 120 kDa and check that this size range reached the membrane. The supplied features do not specify a particular transfer method or settings.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A06226-2 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 NAT10 bands be quantified across samples?
Quantitation · Quantify the same identified band consistently across samples, accounting for loading and transfer. If multiple bands appear, assess them separately before combining signals: the two listed isoforms and modification sites do not establish which bands represent NAT10.
Why does NAT10 appear near 120 kDa instead of 115.7 kDa?
Interpretation · The supplied apparent band is approximately 120 kDa, versus a predicted mass of 115.7 kDa. That difference alone does not identify a cause; the listed modifications do not establish a visible shift. Use the approximate 120 kDa region as a starting point for band identification.
Compare them with the approximately 120 kDa observed band and consider whether the antibody can detect isoform 2, which lacks residues 1–72. The listed modifications do not by themselves identify a shifted band. Verify band identity before assigning an unexpected signal to an isoform or modification.
UniProt locates NAT10 in the nucleus and nucleolus and also lists the midbody. If using fractionated samples, include a nuclear fraction when assessing NAT10 and compare like fractions across samples.
Compare them with the approximately 120 kDa observed band and consider whether the antibody can detect isoform 2, which lacks residues 1–72. The listed modifications do not by themselves identify a shifted band. Verify band identity before assigning an unexpected signal to an isoform or modification.
UniProt locates NAT10 in the nucleus and nucleolus and also lists the midbody. If using fractionated samples, include a nuclear fraction when assessing NAT10 and compare like fractions across samples.
Boster reagents
NAT10 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-NAT10 Antibody Picoband®
Cat # A06226-2
Real WB data
Anti-NAT10 Rabbit Monoclonal Antibody
Cat # M06226
The catalog reports two anti-NAT10 antibodies with Western blot images. A06226-2 shows an approximately 120 kDa band in three human cell lysates; M06226 shows an approximately 116 kDa band in the listed human, monkey, rat, and mouse cell lysates. Evidence is limited to these reported conditions.
Which to pick: For human lysates, either antibody has a WB image. Consider M06226 for monkey, rat, or mouse samples because its image includes examples from those species; A06226-2 lists human reactivity only. The pictured cell lines are tested examples, not universal validation.
Source: BosterBio NAT10 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q9H0A0.
- Human Protein Atlas. NAT10 tissue expression.
- PMC13478519 — target-verified WB comparison
- PMC9076013 — target-verified WB comparison
- PMC13464070 — target-verified WB comparison