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- Table of Contents
NUSAP1 / Nucleolar and spindle-associated protein 1 · Western blot design guide
Design a Western Blot for NUSAP1
Source-linked NUSAP1 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-NUSAP1 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
NUSAP1 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 | ~49.5 kDa | |
| Observed band | Approximately 52 and 60 kDa | |
| Gel | 10% (catalog A06066-2) | |
| Positive control | Appendix (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 | 7 isoform(s) |
Section 1
Source-Linked NUSAP1 Western Blot Protocol Options
The A06066-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human 293T, human Hela, human Jurkat, human U251 (catalog A06066-2) |
| Gel % | 10% (catalog A06066-2) |
| Load | 30 ug; reducing conditions (catalog A06066-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A06066-2) |
| Membrane | nitrocellulose membrane (catalog A06066-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A06066-2) |
| Primary antibody | A06066-2 · 1:1000 (catalog A06066-2) |
| Primary incubation | overnight at 4°C (catalog A06066-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A06066-2) |
| Secondary incubation | 1.5 hour at RT (catalog A06066-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A06066-2) |
| Detection | ECL (catalog A06066-2) |
Section 2
What Is the Expected NUSAP1 Western Blot Band Size?
NUSAP1 is predicted at 49.5 kDa; empirical bands occur near 52 and 60 kDa, and the cause of the difference is not established.
| Band near 52 kDa | An empirical NUSAP1 antibody band; confirm identity with controls. |
| Band near 60 kDa | Another empirical NUSAP1 antibody band; its difference from the predicted mass is unexplained. |
| Band near 49.5 kDa | Near the sequence-predicted mass, though band identity still needs confirmation. |
| Several bands at different masses | Splice isoforms are possible contributors, but their migration is not established. |
💡Expected NUSAP1 appearanceNUSAP1 has a predicted mass of 49.5 kDa and empirical antibody bands near 52 and 60 kDa; their difference is unexplained, so confirm band identity with appropriate controls.
| UniProt predicted mass | 49.5 kDa is the sequence-based reference, while empirical bands occur near 52 and 60 kDa. |
| Splice isoforms 1 and 2 | Alternative sequences may affect apparent size; individual masses are unavailable. |
| Splice isoforms 3 and 4 | Alternative sequences may affect apparent size; individual masses are unavailable. |
| Splice isoforms 5, 6 and 7 | Alternative sequences may affect apparent size; individual masses are unavailable. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Nuclear or spindle-associated NUSAP1 may be poorly recovered in the sampled lysate fraction. | Check extraction and compare nuclear or cytoskeletal fractions. |
| Band higher than expected | A band near 60 kDa is reported, but its size difference has no established cause. | Compare with a second antibody and an appropriate positive control. |
| Band lower than expected | An unassigned splice isoform could differ in size. | Check isoform expression and whether the antibody recognizes its sequence. |
| Multiple bands | Seven splice isoforms are listed, but their band positions are unknown. | Use an independent antibody or NUSAP1 depletion to identify specific bands. |
| Weak or no signal | The sampled fraction may miss NUSAP1 in the nucleus or spindle. | Check extraction and test the relevant cellular fraction. |
Experiment evidence
Western blot images showing NUSAP1 band
A06066-2 · NUSAP1
A06066-2 · NUSAP1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for NUSAP1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for NUSAP1 in Western blot, you can use appendix tissue, which has high HPA expression.
Positive control: Appendix (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: High expression in appendix and no detection in adipose tissue make tissue controls feasible.
HPA tissue expression evidence for NUSAP1
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 |
|---|---|---|---|---|
| Appendix | endocrine cells | High | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | High | Protein (IHC) | HPA → |
| Colon | endocrine cells | High | Protein (IHC) | HPA → |
| Duodenum | endocrine cells | High | Protein (IHC) | HPA → |
| Rectum | endocrine cells | High | 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 → |
| Caudate | glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Cerebral cortex | endothelial cells | Not detected | Protein (IHC) | HPA → |
| Heart muscle | cardiomyocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced NUSAP1 Western Blot Tips
Deeper troubleshooting and optimisation questions for NUSAP1, answered from its protein features.
How should NUSAP1 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.
Which NUSAP1 isoforms could produce different bands?
Isoforms · UniProt lists seven isoforms. Relative to the canonical sequence, isoform 7 lacks residues 32–54 and 336–374; isoforms 4 and 6 lack 102–116; isoforms 3 and 5 lack residue 102; and isoforms 2, 3, 4 and 7 lack residue 221. These differences make isoforms worth checking, but do not establish which, if any, produces either observed band.
Check the antibody’s epitope against the listed sequence differences. An epitope within canonical residues 32–54 or 336–374 would be absent from isoform 7; one within 102–116 may be affected in isoforms 3–6. UniProt coordinates here refer to the canonical sequence and may differ from antibody numbering.
Check the antibody’s epitope against the listed sequence differences. An epitope within canonical residues 32–54 or 336–374 would be absent from isoform 7; one within 102–116 may be affected in isoforms 3–6. UniProt coordinates here refer to the canonical sequence and may differ from antibody numbering.
Which NUSAP1 modifications should guide band analysis?
PTM · In canonical UniProt numbering, phosphosites are S124 (by ATM), S135, T182, S240, T244, S247, S255, S269, S276, S311, T314, T338, T349, S352 and S363; K411 is acetylated. Compare bands across relevant conditions or with a phosphorylation-sensitive assay. The annotations alone do not demonstrate a visible shift.
Does this guide establish induction of NUSAP1?
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 NUSAP1?
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 A06066-2 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 cell-cycle state affect NUSAP1 quantitation?
Quantitation · NUSAP1 is annotated with cell-cycle and mitosis functions and redistributes from the cytoplasm and nucleolus during interphase to the mitotic spindle in prometaphase. Match cell-cycle conditions when comparing samples, and use a consistent fraction or whole-cell preparation. Location changes can affect comparisons between fractions; these features do not establish an abundance change.
Do the 52 and 60 kDa bands match NUSAP1’s predicted mass?
Interpretation · The supplied Western blots show bands near 52 and 60 kDa, versus a predicted mass of 49.5 kDa. NUSAP1 has annotated modifications and seven isoforms, but those features alone do not explain either apparent mass or establish that both bands are NUSAP1. Compare each band with a NUSAP1-specific control.
UniProt annotates canonical Ser124 as phosphorylated by ATM. If samples differ in ATM activity, compare their NUSAP1 band patterns and assess Ser124 phosphorylation directly where possible. The site annotation does not predict that ATM activity will change total NUSAP1 abundance or produce the 60 kDa band.
For bands beyond the observed approximately 52 and 60 kDa signals, consider the seven annotated isoforms and modification sites as possibilities, then test band identity with a NUSAP1-specific control. UniProt lists no signal peptide, propeptide or glycosylation sites, so the supplied features do not support assigning an extra band to those forms.
UniProt annotates canonical Ser124 as phosphorylated by ATM. If samples differ in ATM activity, compare their NUSAP1 band patterns and assess Ser124 phosphorylation directly where possible. The site annotation does not predict that ATM activity will change total NUSAP1 abundance or produce the 60 kDa band.
For bands beyond the observed approximately 52 and 60 kDa signals, consider the seven annotated isoforms and modification sites as possibilities, then test band identity with a NUSAP1-specific control. UniProt lists no signal peptide, propeptide or glycosylation sites, so the supplied features do not support assigning an extra band to those forms.
Boster reagents
NUSAP1 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-NUSAP1 Antibody Picoband®
Cat # A06066-2
The catalog reports A06066-2, a rabbit polyclonal anti-NUSAP1 antibody with a Western blot image from human 293T, HeLa, Jurkat, and U251 whole-cell lysates. The reported band is approximately 52 kDa versus an expected 49 kDa; no independent specificity validation is supplied.
Which to pick: A06066-2 is the only listed option. Its reported human reactivity and Western blot image support considering it for human lysates, using the shown samples and conditions as a starting point.
Source: BosterBio NUSAP1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q9BXS6.
- Human Protein Atlas. NUSAP1 tissue expression.
- PMC11489181 — target-verified WB comparison
- PMC12824744 — target-verified WB comparison
- PMC8667124 — target-verified WB comparison