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- Table of Contents
NODAL / Nodal homolog · Western blot design guide
Design a Western Blot for NODAL
Real validated NODAL Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-NODAL 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
NODAL 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 | ~39.6 kDa | |
| Gel | 15% (standard starting point) | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Glycosylated + Cleaved | |
| Caveat | Processing-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Real Curated NODAL Western Blot Protocols
The M07627 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | SH-SY5Y cell lysate (catalog M07627) |
| Gel % | 15% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Short semi-dry transfer; verify retention (standard starting point) |
| Membrane | 0.2 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | M07627; use the WB datasheet starting dilution (standard starting point) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | Species-matched HRP conjugate at validated dilution (standard starting point) |
| 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 NODAL Western Blot Band Size?
NODAL’s full-length precursor is predicted at 39.6 kDa; cleavage, N-linked sites, and disulfide-linked dimerization may affect bands, but migration has not been demonstrated here.
| Band near 39.6 kDa | consistent with the predicted full-length precursor; identity needs confirmation |
| Band below 39.6 kDa | could reflect signal peptide or propeptide cleavage |
| Band near twice the monomer position under non-reducing conditions | could reflect the disulfide-linked homodimer |
| Little or no band in whole-cell lysate | consistent with secretion of NODAL |
💡Expected NODAL appearanceThe full-length precursor has a predicted mass of 39.6 kDa; cleavage, N-linked glycosylation, and disulfide-linked dimerization may alter migration, but no empirical band size is supplied, so confirm identity with appropriate controls.
| Predicted full-length precursor mass | sets a 39.6 kDa reference, not a validated band position |
| N-linked glycosylation at Asn72 | may affect precursor migration; this site lies in the propeptide |
| N-linked glycosylation at Asn199 | may affect precursor migration; this site lies in the propeptide |
| Signal peptide and propeptide cleavage | can produce a mature chain smaller than the full-length precursor |
| Disulfide-linked homodimer | can yield a band near twice the monomer position under non-reducing or incompletely reducing conditions |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | NODAL is secreted | check conditioned medium alongside lysate |
| Band higher than expected | disulfide-linked dimers may persist with incomplete reduction | compare reducing and non-reducing lanes |
| Band lower than expected | signal peptide and propeptide cleavage can produce a smaller chain | compare precursor and mature-chain detection using an appropriate antibody |
| Multiple bands | precursor, processed chain, or disulfide-linked dimer may be present | compare reducing conditions and verify band identity with an independent antibody |
| Weak or no signal | secreted NODAL may be scarce in the sampled lysate | check conditioned medium and include a positive control |
| Fragments below expected size | propeptide cleavage creates a shorter mature chain | verify the antibody epitope and compare with a positive control |
Experiment evidence
Western blot images showing NODAL band
M07627 · NODAL
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for NODAL Western blot
🧪For positive controls for NODAL in Western blot, you can use no HPA-supported positive sample because none was supplied.
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: NODAL is secreted, so conditioned medium may be more informative than whole-cell lysate.
HPA tissue expression evidence for NODAL
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 NODAL Western Blot Tips
Deeper troubleshooting and optimisation questions for NODAL, answered from its protein features.
How should NODAL 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 NODAL bands evidence of isoforms?
Isoforms · UniProt lists one isoform and no alternative sequence. Multiple bands therefore cannot be assigned to listed isoforms from this evidence. Consider processing, glycosylation, or disulfide-linked species, and identify which NODAL region the antibody recognizes.
How can NODAL glycosylation affect band interpretation?
PTM · UniProt lists N-linked sites at Asn72 and Asn199, both within the annotated propeptide at 27–237. Glycosylation may affect precursor mobility; the listed sites do not establish glycosylation of a processed product or predict a particular band shift. Use UniProt residue numbering when comparing site labels.
Does this guide establish induction of NODAL?
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 NODAL Western blot?
Transfer · UniProt provides a 39.6 kDa predicted mass for the full sequence and annotates processing and disulfide-linked homodimers. Choose and verify transfer conditions against the NODAL species your antibody detects, since the feature list does not specify an apparent band size or a transfer method.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the M07627 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.
Where should NODAL be measured for quantitation?
Quantitation · NODAL is annotated as secreted, so assess the relevant extracellular sample as well as any cell-associated fraction your experiment requires. Quantify the same antibody-recognized NODAL species across samples; precursor and processed bands may represent different pools.
Why might NODAL bands differ from its predicted mass?
Interpretation · The 39.6 kDa prediction is for the 347-residue sequence. UniProt lists a signal peptide at 1–26, a propeptide at 27–237, and N-linked glycosylation sites at Asn72 and Asn199. Processing and glycosylation can change the species detected, but these features alone do not establish a visible shift or a specific apparent mass.
NODAL is secreted and has a signal peptide at 1–26 and a propeptide at 27–237. An antibody's epitope determines whether it detects precursor, a processed form, or both. Check the immunogen location before interpreting multiple bands as processing products.
UniProt describes NODAL as a disulfide-linked homodimer and lists four disulfide bonds. Compare reducing and nonreducing preparations when testing whether an unexpected high-mass band reflects disulfide-linked NODAL. Band position alone cannot establish its identity.
NODAL is secreted and has a signal peptide at 1–26 and a propeptide at 27–237. An antibody's epitope determines whether it detects precursor, a processed form, or both. Check the immunogen location before interpreting multiple bands as processing products.
UniProt describes NODAL as a disulfide-linked homodimer and lists four disulfide bonds. Compare reducing and nonreducing preparations when testing whether an unexpected high-mass band reflects disulfide-linked NODAL. Band position alone cannot establish its identity.
Boster reagents
NODAL 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-Nodal Rabbit Monoclonal Antibody
Cat # M07627
The catalog reports one anti-NODAL antibody: M07627, a rabbit monoclonal with reported human reactivity. Its Western blot image shows Nodal expression in SH-SY5Y cell lysate; the supplied evidence does not establish performance in other samples.
Which to pick: M07627 is the only listed option and has a Western blot image using SH-SY5Y cell lysate. It is reported reactive with human samples; suitability for other contexts is not documented here.
Source: BosterBio NODAL gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q96S42.
- Human Protein Atlas. NODAL tissue expression.
- PMC6485227 — target-verified WB comparison
- PMC3139718 — target-verified WB comparison
- PMC6779822 — target-verified WB comparison