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- Table of Contents
NPR3 / Atrial natriuretic peptide receptor 3 · Western blot design guide
Design a Western Blot for NPR3
Source-linked NPR3 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-NPR3 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
NPR3 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 | ~59.8 kDa | |
| Observed band | ~72 kDa | |
| Gel | 10% (catalog M03589) | |
| Positive control | Kidney (IHC candidate; verify WB) | |
| Negative control | Adipose tissue (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Glycosylated + Cleaved | |
| Caveat | Glycosylation-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 3 isoform(s) |
Section 1
Source-Linked NPR3 Western Blot Protocol Options
The M03589 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human Hela, human PC-3, rat kidney, rat heart, mouse kidney, mouse heart (catalog M03589) |
| Gel % | 10% (catalog M03589) |
| Load | 30 ug; reducing conditions (catalog M03589) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog M03589) |
| Membrane | nitrocellulose membrane (catalog M03589) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog M03589) |
| Primary antibody | M03589 · 1:500 (catalog M03589) |
| Primary incubation | overnight at 4°C (catalog M03589) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog M03589) |
| Secondary incubation | 1.5 hour at RT (catalog M03589) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog M03589) |
| Detection | ECL (catalog M03589) |
Section 2
What Is the Expected NPR3 Western Blot Band Size?
NPR3 is predicted at 59.8 kDa and observed near 72 kDa under reducing conditions; the cause of the difference is not established.
| Band near 72 kDa under reducing conditions | Empirical NPR3 band in whole-cell and tissue lysates; UniProt predicts 59.8 kDa |
| Band near twice the monomer size under non-reducing conditions | Consistent with a disulfide-linked NPR3 homodimer |
| Band below the full-length precursor | Could reflect cleavage of the 1–26 signal peptide; identity needs confirmation |
| Several bands at different sizes | Could reflect isoforms 1, 2, and 3, though distinct migration is unestablished |
| Broad band or smear | Could reflect variation in N-linked glycosylation; the three sites alone do not establish a smear |
💡Expected NPR3 appearanceLook for the empirically observed NPR3 band near 72 kDa under reducing conditions, compared with the 59.8 kDa UniProt precursor prediction; confirm its identity with an appropriate control.
| UniProt predicted precursor mass | 59.8 kDa predicted, while the empirical reducing-condition band is near 72 kDa; the cause of the difference is unestablished |
| N86 complex N-linked glycosylation | May alter apparent migration; its contribution to the 72 kDa band is unknown |
| N293 high-mannose N-linked glycosylation | May alter apparent migration; its contribution to the 72 kDa band is unknown |
| N394 complex N-linked glycosylation | May alter apparent migration; its contribution to the 72 kDa band is unknown |
| Disulfide-linked homodimer | Can yield a band near twice the monomer size without complete reduction |
| Isoforms 1, 2, and 3 | May differ in size, but their masses and resolvable migration differences are unspecified |
| Signal peptide at residues 1–26 | Cleavage makes the mature chain smaller than the full-length precursor; its apparent migration is unspecified |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | NPR3 is a single-pass membrane protein and may be poorly extracted | Check membrane-protein recovery and use a positive lysate control |
| Band higher than expected | Incomplete reduction may retain disulfide-linked dimers | Compare reducing and non-reducing samples and confirm band identity |
| Band lower than expected | Signal-peptide cleavage can shorten the precursor | Check antibody epitope and confirm the band with an independent antibody or knockdown |
| Broad smear instead of sharp band | Variation in N-linked glycosylation is possible | Compare matched samples before and after N-glycan removal |
| Multiple bands | Isoforms 1, 2, and 3 or incomplete reduction may contribute | Compare reduction conditions and confirm individual bands with an independent antibody or knockdown |
| Weak or no signal | Membrane-protein extraction may be inefficient | Check extraction and loading with a positive lysate control |
Experiment evidence
Western blot images showing NPR3 band
M03589 · NPR3
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for NPR3 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for NPR3 in Western blot, you can use kidney tissue lysate, which HPA scores High.
Positive control: Kidney (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: As a membrane protein, NPR3 may be easier to detect in a membrane-enriched fraction.
HPA tissue expression evidence for NPR3
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 |
|---|---|---|---|---|
| Kidney | bowman's capsule | 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 → |
| Appendix | glandular cells | Not detected | Protein (IHC) | HPA → |
| Breast | adipocytes | Not detected | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | Not detected | Protein (IHC) | HPA → |
| Caudate | glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced NPR3 Western Blot Tips
Deeper troubleshooting and optimisation questions for NPR3, answered from its protein features.
How should NPR3 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.
How could NPR3 isoforms affect antibody detection?
Isoforms · UniProt lists isoforms 1, 2, and 3. Isoform 3 replaces canonical residues 1–40 and lacks residues 41–256, so an antibody targeting that deleted region would not detect it. Check the antibody epitope before assigning bands to isoforms.
Which NPR3 glycosylation sites matter when interpreting bands?
PTM · UniProt annotates N-linked glycosylation at Asn86 and Asn394 (complex) and Asn293 (high mannose). These are UniProt canonical-sequence coordinates; antibody or paper numbering may differ. The site annotations do not establish that a particular band shift will be visible.
Does this guide establish induction of NPR3?
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 NPR3 Western blot?
Transfer · NPR3 is a single-pass membrane protein and can form disulfide-linked dimers. Check transfer across both the monomer and higher-mass regions, then adjust transfer conditions based on the gel and membrane used. The supplied features do not specify a particular transfer method.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the M03589 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 NPR3 bands be quantified?
Quantitation · Define which band or bands you will measure before comparing samples. NPR3 has three isoforms, annotated glycosylation, and disulfide-linked dimers, so confirm that the measured signal represents the same species under the same sample conditions. Report whether quantitation includes a higher-mass band.
Why might NPR3 appear near 72 kDa instead of 59.8 kDa?
Interpretation · The observed ~72 kDa band is above the predicted 59.8 kDa mass. NPR3 has a signal peptide at residues 1–26 and three annotated N-linked glycosylation sites. These features may affect its apparent size, but the annotations alone do not establish the cause or size of the difference.
NPR3 is annotated as a disulfide-linked homodimer; isoform 2 can also form dimers through its C-terminal cysteine. Compare reducing and nonreducing samples when assessing a higher-mass band. Migration alone cannot identify it as a dimer.
Isoform 3 lacks canonical residues 41–256 and has a different sequence at residues 1–40, making it a candidate to investigate for a smaller band. Its annotated sequence does not establish where it migrates. Check whether the antibody epitope is retained before assigning the band.
NPR3 is annotated as a disulfide-linked homodimer; isoform 2 can also form dimers through its C-terminal cysteine. Compare reducing and nonreducing samples when assessing a higher-mass band. Migration alone cannot identify it as a dimer.
Isoform 3 lacks canonical residues 41–256 and has a different sequence at residues 1–40, making it a candidate to investigate for a smaller band. Its annotated sequence does not establish where it migrates. Check whether the antibody epitope is retained before assigning the band.
Boster reagents
NPR3 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-NPRC Rabbit Monoclonal Antibody
Cat # M03589
The catalog reports one anti-NPR3 antibody, M03589 (labeled anti-NPRC). Its Western blot image shows an approximately 72 kDa band in human cell, rat tissue, and mouse tissue lysates. Evidence is limited to the reported samples and conditions.
Which to pick: M03589 is the only listed option. It reports human, mouse, and rat reactivity and includes a Western blot image using HeLa, PC-3, kidney, and heart lysates. Match your sample and protocol to the reported conditions.
Source: BosterBio NPR3 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P17342.
- Human Protein Atlas. NPR3 tissue expression.
- PMC6885401 — target-verified WB comparison
- PMC5026813 — target-verified WB comparison
- PMC3685298 — target-verified WB comparison