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- Table of Contents
NCOA2 / Nuclear receptor coactivator 2 · Western blot design guide
Design a Western Blot for NCOA2
Real validated NCOA2 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-NCOA2 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
NCOA2 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 | ~159.2 kDa | |
| Observed band | ~160–170 kDa | |
| Gel | 8% (catalog A01706-2) | |
| 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 C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Real Curated NCOA2 Western Blot Protocols
The A01706-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human HEL, human RT4 (catalog A01706-2) |
| Gel % | 8% (catalog A01706-2) |
| Load | 30 ug; reducing conditions (catalog A01706-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A01706-2) |
| Membrane | nitrocellulose membrane (catalog A01706-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A01706-2) |
| Primary antibody | A01706-2 · 0.5 μg/mL (catalog A01706-2) |
| Primary incubation | overnight at 4°C (catalog A01706-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A01706-2) |
| Secondary incubation | 1.5 hour at RT (catalog A01706-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A01706-2) |
| Detection | ECL (catalog A01706-2) |
Section 2
What Is the Expected NCOA2 Western Blot Band Size?
NCOA2 is predicted at 159.2 kDa and observed at ~160–170 kDa; the cause of the difference is not established.
| Band at ~160–170 kDa | Empirical NCOA2 band in whole-cell lysates; confirm identity with appropriate controls |
| Band near 159 kDa | Close to the 159.2 kDa predicted mass; identity still requires confirmation |
| Nearby band or subtle doublet | Could reflect different modification states, but a visible effect is unproven |
| Stronger band in nuclear extract | Consistent with NCOA2 nuclear localization |
💡Expected NCOA2 appearanceNCOA2 has a predicted mass of 159.2 kDa and an empirical band at ~160–170 kDa; confirm band identity with appropriate controls because the cause of the difference is unestablished.
| UniProt predicted mass | Places unmodified NCOA2 near 159.2 kDa; the empirical band is ~160–170 kDa |
| Phosphoserine at residue 29 | May affect apparent migration, but no shift is established |
| Phosphoserines at residues 487, 493 and 499 | May affect apparent migration, but no distinct band pattern is established |
| N-acetylserine at residue 2 | Adds a small modification without an established visible shift |
| Asymmetric dimethylarginine at residue 338 | Adds a small modification without an established visible shift |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Nuclear NCOA2 may be poorly recovered | Check nuclear extraction and a nuclear marker |
| Band higher than expected | Identity or migration of the higher band is uncertain | Compare with the ~160–170 kDa reference band and verify specificity |
| Band lower than expected | Possible proteolysis or nonspecific binding | Use protease inhibitors and verify band identity |
| Multiple bands | Modification states or nonspecific binding are possible; distinct NCOA2 bands are unproven | Compare antibodies or use NCOA2 depletion to identify the specific band |
| Weak or no signal | Poor recovery of nuclear NCOA2 | Check nuclear fraction recovery and sample loading |
| Fragments below expected size | Possible sample proteolysis | Process samples promptly with protease inhibitors and compare fresh lysate |
Experiment evidence
Western blot images showing NCOA2 band
A01706-2 · NCOA2
P01706 · NCOA2
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for NCOA2 Western blot
🧪For positive controls for NCOA2 in Western blot, you can use an HPA-IHC candidate positive sample once identified; none is supplied here.
Positive control: No high/medium HPA tissue identified
Negative control: Suggested KO / knockdown lysate
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: HPA provides no tissue data, so a positive tissue cannot be selected and a knockdown or KO line is needed for a negative control.
HPA tissue expression evidence for NCOA2
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 |
|---|---|---|---|---|
| No high/medium HPA tissues identified in the supplied evidence. | ||||
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| No lower-expression tissue rows available in the supplied evidence. | ||||
Section 3
Advanced NCOA2 Western Blot Tips
Deeper troubleshooting and optimisation questions for NCOA2, answered from its protein features.
How should NCOA2 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 NCOA2 isoforms explain multiple bands?
Isoforms · The supplied UniProt record lists one isoform and no alternative sequence. It therefore provides no specific isoform assignment for additional bands. Confirm any extra band independently before labeling it an NCOA2 isoform.
Which NCOA2 modifications matter when comparing bands?
PTM · UniProt lists phosphorylation, acetylation and arginine methylation, including phosphoserines at positions 487, 493 and 499 and acetyllysines at 636 and 640. These are UniProt sequence coordinates; antibody or paper numbering may differ. Keep sample conditions consistent when comparing bands. A listed modification does not by itself establish a visible shift.
Does this guide establish induction of NCOA2?
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 NCOA2 Western blot?
Transfer · NCOA2 is approximately 159.2 kDa, so choose and optimize a transfer setup that retains and transfers proteins around 160–170 kDa. Check transfer in that size range with a molecular weight marker or total protein stain before interpreting a weak signal.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A01706-2 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 NCOA2 bands be quantified?
Quantitation · NCOA2 is listed as nuclear. Compare equivalent nuclear samples, keep sample preparation consistent, and normalize to an appropriate loading measure. If more than one band appears near 160–170 kDa, state which band or bands were quantified; the supplied features do not establish that each represents NCOA2.
How should the observed band compare with predicted NCOA2 mass?
Interpretation · NCOA2 has a predicted mass of 159.2 kDa, while the supplied Western blot observation is approximately 160–170 kDa. Use that range as a guide when identifying the band. The listed modifications alone do not establish why apparent and calculated masses differ.
Start with the predicted 159.2 kDa mass and observed approximately 160–170 kDa range, then verify the identity of bands outside that range. The supplied record lists no signal peptide, propeptide or glycosylation sites, so those features do not provide an explanation. Listed modifications and protein complexes also do not establish the identity or cause of an extra band.
Start with the predicted 159.2 kDa mass and observed approximately 160–170 kDa range, then verify the identity of bands outside that range. The supplied record lists no signal peptide, propeptide or glycosylation sites, so those features do not provide an explanation. Listed modifications and protein complexes also do not establish the identity or cause of an extra band.
Boster reagents
NCOA2 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-NCOA2 Antibody Picoband®
Cat # A01706-2
Real WB data
Anti-Phospho NCoA2 (Ser736) Antibody
Cat # P01706
Two the supplier antibodies are listed: A01706-2 for NCOA2 and P01706 for phospho-NCOA2 (Ser736). Both have WB images. A01706-2 is shown with human HEL and RT4 lysates; P01706 is shown with TSA-treated HeLa lysate and phosphopeptide blocking. These examples do not establish validation in every listed species.
Which to pick: Choose A01706-2 for NCOA2 in human samples; its image reports a 160–170 kDa band in HEL and RT4 lysates. Choose P01706 for Ser736 phosphorylation. It lists human, mouse, and rat reactivity, but its supplied WB example uses HeLa cells.
Source: BosterBio NCOA2 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q15596.
- Human Protein Atlas. NCOA2 tissue expression.
- PMC10054615 — target-verified WB comparison
- PMC6894885 — target-verified WB comparison
- PMC12061476 — target-verified WB comparison