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NLRP1 / NACHT, LRR and PYD domains-containing protein 1 · Western blot design guide
Design a Western Blot for NLRP1
Real validated NLRP1 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-NLRP1 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
NLRP1 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 | ~165.9 kDa | |
| Gel | 8–10% (standard starting point) | |
| Positive control | Cerebral cortex (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 + Ubl conjugation | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 5 isoform(s) |
Section 1
Real Curated NLRP1 Western Blot Protocols
The A00777 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | U937 cell lysate (catalog A00777) |
| Gel % | 8–10% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Wet/tank transfer; optimize duration (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | A00777 · (A) 1, (B) 2 and (C) 4 μg/mL (catalog A00777) |
| 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 NLRP1 Western Blot Band Size?
Full-length NLRP1 is predicted at 165.9 kDa; isoforms and phosphorylation could affect bands, but their migration effects are not demonstrated here.
| Band near 165.9 kDa | consistent with full-length NLRP1, pending band-identity controls |
| Several bands at different positions | may reflect isoforms 1–5; their separation is not established |
| Doublet near 165.9 kDa | may reflect phosphorylation, but a visible shift is not established |
| Bands in cytosolic and nuclear fractions | consistent with NLRP1's documented nucleocytoplasmic distribution |
💡Expected NLRP1 appearanceFull-length NLRP1 has a predicted mass of 165.9 kDa; no empirical band position is supplied, so confirm any candidate band with antibody and sample controls.
| Predicted full-length mass | sets a 165.9 kDa reference, not a measured band position |
| Isoforms 1, 2, 3, 4 and 5 | may differ in size; their relative masses and band separation are unspecified |
| Phosphoserines at residues 93, 99, 101 and 107 | may affect mobility; no visible shift is established |
| Phosphothreonines at residues 112, 114 and 129 and phosphoserine at residue 113 | may affect mobility; no visible shift is established |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | NLRP1 detection in this sample is unverified | Compare with U937 lysate and check antibody performance and loading |
| Band higher than expected | Phosphorylation could affect mobility, but its effect here is unmeasured | Compare phosphatase-treated and untreated samples and verify band identity |
| Band lower than expected | An alternative isoform is possible; isoform masses are unspecified | Check isoform recognition and verify the band with an independent antibody |
| Multiple bands | Isoforms or phosphorylation are possible, but distinct bands are unconfirmed | Compare isoform recognition and phosphatase treatment with an independent antibody |
| Weak or no signal | NLRP1 has cell-dependent cytoplasmic and nuclear distribution | Check the relevant fractions and compare with U937 lysate |
Experiment evidence
Western blot images showing NLRP1 band
A00777 · NLRP1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for NLRP1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for NLRP1 in Western blot, you can use cerebral cortex lysate, where HPA reports high expression.
Positive control: Cerebral cortex (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: HPA reports undetected expression in adipose tissue, providing a feasible negative tissue control.
HPA tissue expression evidence for NLRP1
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 |
|---|---|---|---|---|
| Cerebral cortex | neuronal cells | High | Protein (IHC) | HPA → |
| Hippocampus | neuronal cells | High | Protein (IHC) | HPA → |
| Skin | melanocytes | High | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Medium | Protein (IHC) | HPA → |
| Colon | mucosal lymphoid cells | Medium | 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 | endocrine cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Cervix | glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced NLRP1 Western Blot Tips
Deeper troubleshooting and optimisation questions for NLRP1, answered from its protein features.
How should NLRP1 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 NLRP1 isoforms could produce different bands?
Isoforms · UniProt lists five isoforms. Isoforms 3 and 4 lack residues 958–987; isoforms 2 and 3 lack 1262–1305. Isoform 5 has changes at 1044 and 1354–1371 and lacks 1372–1473. Check whether the antibody epitope is retained in the isoforms you intend to detect. These are UniProt canonical sequence coordinates.
Which NLRP1 phosphorylation sites should I track?
PTM · UniProt coordinates: MAPK11/MAPK14 phosphorylate S93, S99, S101, S168 and S170; MAPK14 phosphorylates S107 and S163; MAPK11, MAPK14 and MAP3K20 phosphorylate T112. MAP3K20 phosphorylates S113, T114, T129 and S132; MAPK11 phosphorylates S173 and T178; MAPK11/MAP3K20 phosphorylate S179 and T180. Check the numbering convention before comparing these sites with an antibody datasheet.
NLRP1 has 16 listed phosphoserine or phosphothreonine sites, concentrated between UniProt residues 93 and 180. Compare matched samples when assessing a suspected mobility difference. The site annotations establish possible modifications, but do not show that phosphorylation produces a visible Western blot shift.
NLRP1 has 16 listed phosphoserine or phosphothreonine sites, concentrated between UniProt residues 93 and 180. Compare matched samples when assessing a suspected mobility difference. The site annotations establish possible modifications, but do not show that phosphorylation produces a visible Western blot shift.
Does this guide establish induction of NLRP1?
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 NLRP1 Western blot?
Transfer · NLRP1 has a predicted mass of 165.9 kDa. Choose transfer conditions that recover proteins around that size, then check the high-mass markers and whether NLRP1 remains in the gel. The supplied UniProt features do not specify a transfer method or settings.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A00777 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 I quantify NLRP1 across samples?
Quantitation · Quantify the same defined band or set of bands across samples, and record which isoforms the antibody can recognize. NLRP1 has five listed isoforms and multiple phosphorylation sites, so summing bands or comparing different bands could change what the measurement represents. Check signal linearity before comparing intensities.
Should NLRP1 run at its predicted 165.9 kDa?
Interpretation · Use 165.9 kDa as the predicted mass of the canonical protein, not an exact band position. No observed band size is supplied. The listed splice variants and phosphorylation sites are worth considering when assessing a different apparent mass, but their presence alone does not establish a visible shift or explain a particular band.
First compare each band with the 165.9 kDa canonical prediction and the listed isoform sequence changes. Check antibody epitope coverage, particularly across the deleted regions. Phosphorylation is another feature to consider, but these annotations alone cannot identify an unexpected band or establish why it migrates differently.
First compare each band with the 165.9 kDa canonical prediction and the listed isoform sequence changes. Check antibody epitope coverage, particularly across the deleted regions. Phosphorylation is another feature to consider, but these annotations alone cannot identify an unexpected band or establish why it migrates differently.
Boster reagents
NLRP1 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-NALP1 NLRP1 Antibody
Cat # A00777
The catalog reports one anti-NLRP1 antibody, A00777, with Human reactivity. Its Western blot image shows U937 cell lysate tested at 1, 2, and 4 μg/mL. The supplied evidence does not establish performance in other samples.
Which to pick: A00777 is the only listed option. Its U937 Western blot image provides a starting point for Human samples; confirm performance in your own specimen and conditions.
Source: BosterBio NLRP1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q9C000.
- Human Protein Atlas. NLRP1 tissue expression.
- PMC5682047 — target-verified WB comparison
- PMC8417576 — target-verified WB comparison
- PMC5501746 — target-verified WB comparison