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PYCR1 / Pyrroline-5-carboxylate reductase 1, mitochondrial · Western blot design guide
Design a Western Blot for PYCR1
Source-linked PYCR1 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-PYCR1 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
PYCR1 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 | ~33.4 kDa | |
| Observed band | ~35 kDa | |
| Gel | 5–20% (catalog A06018-1) | |
| Positive control | Bronchus (IHC candidate; verify WB) +4 more | |
| Negative control | Oral mucosa (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 | 3 isoform(s) |
Section 1
Source-Linked PYCR1 Western Blot Protocol Options
The A06018-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human SW620, human PANC-1, human 293T (catalog A06018-1) |
| Gel % | 5–20% (catalog A06018-1) |
| Load | 30 ug; reducing conditions (catalog A06018-1) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A06018-1) |
| Membrane | nitrocellulose membrane (catalog A06018-1) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A06018-1) |
| Primary antibody | A06018-1 · 0.5 μg/mL (catalog A06018-1) |
| Primary incubation | overnight at 4°C (catalog A06018-1) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A06018-1) |
| Secondary incubation | 1.5 hour at RT (catalog A06018-1) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A06018-1) |
| Detection | ECL (catalog A06018-1) |
Section 2
What Is the Expected PYCR1 Western Blot Band Size?
PYCR1 is predicted at 33.4 kDa and observed at ~35 kDa; the cause of the difference is not established.
| Band at ~35 kDa | Empirical PYCR1 band in reducing whole-cell lysates; predicted mass is 33.4 kDa. |
| Single band near 33.4 kDa | Consistent with predicted PYCR1 mass; confirm identity with a control. |
| Additional bands at different positions | Isoforms 1, 2 and 3 are annotated, but their migration and resolution are unknown. |
| Little or no band in a mitochondrial-poor fraction | PYCR1 is mitochondrial; check a mitochondria-enriched fraction. |
💡Expected PYCR1 appearancePYCR1 has a predicted mass of 33.4 kDa and an empirical band at ~35 kDa in reducing whole-cell lysates; the cause of the difference is unestablished, so confirm identity with appropriate controls.
| Predicted PYCR1 mass | 33.4 kDa provides the sequence-based reference; the empirical band is ~35 kDa. |
| Isoforms 1, 2 and 3 | Sequence differences may affect size, but their masses and migration are not supplied. |
| N-acetylserine at residue 2 | Annotated modification with no demonstrated visible band shift. |
| Phosphoserine at residue 278 | Annotated modification with no demonstrated visible band shift. |
| Phosphoserine at residue 301 | Annotated modification with no demonstrated visible band shift. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Mitochondrial PYCR1 may be underrepresented in the preparation. | Check extraction and compare with a mitochondria-enriched fraction and a positive lysate. |
| Band higher than expected | The supplied features do not establish a cause for a higher band. | Compare with the ~35 kDa control band and verify identity with PYCR1 depletion. |
| Band lower than expected | An annotated isoform could differ in size, but its migration is unknown. | Verify identity with isoform-specific evidence or PYCR1 depletion. |
| Multiple bands | Isoforms 1, 2 and 3 are annotated, but distinct bands are unconfirmed. | Compare bands after PYCR1 depletion and assess isoform expression. |
| Weak or no signal | Recovery of mitochondrial PYCR1 may be low. | Check mitochondrial protein recovery and run a positive lysate. |
Experiment evidence
Western blot images showing PYCR1 band
A06018-1 · PYCR1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for PYCR1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for PYCR1 in Western blot, you can use bronchus tissue, which HPA rates High.
Positive control: Bronchus (IHC candidate; verify WB)
Negative control: Oral mucosa (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: A tissue negative is feasible because HPA reports PYCR1 as not detected in oral mucosa.
HPA tissue expression evidence for PYCR1
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 |
|---|---|---|---|---|
| Bronchus | basal cells | High | Protein (IHC) | HPA → |
| Cerebral cortex | neuronal cells | High | Protein (IHC) | HPA → |
| Cervix | glandular cells | High | Protein (IHC) | HPA → |
| Hippocampus | neuronal cells | High | Protein (IHC) | HPA → |
| Pancreas | exocrine glandular cells | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Oral mucosa | squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
| Ovary | ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Low | Protein (IHC) | HPA → |
| Liver | hepatocytes | Low | Protein (IHC) | HPA → |
| Salivary gland | glandular cells | Low | Protein (IHC) | HPA → |
Section 3
Advanced PYCR1 Western Blot Tips
Deeper troubleshooting and optimisation questions for PYCR1, answered from its protein features.
How should PYCR1 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 PYCR1 isoforms affect band interpretation?
Isoforms · UniProt lists three isoforms. Isoform 3 has an altered N terminus, while isoform 2 replaces residues 290–319 of the canonical sequence. These sequence differences could affect migration, but the supplied features do not establish distinct band positions. Identify which isoform an antibody recognizes before assigning bands.
Which PYCR1 phosphorylation sites matter when comparing bands?
PTM · UniProt lists phosphoserine at canonical positions 278 and 301. Position 301 lies within the region replaced in isoform 2, so do not assume that coordinate applies to isoform 2. Phosphorylation alone does not establish a visible band shift; confirm any proposed phospho-dependent band experimentally.
UniProt lists N-acetylserine at position 2 in its sequence numbering. Keep that coordinate explicit when comparing papers or antibody annotations, which may use different numbering. This modification alone does not demonstrate a detectable mass shift.
UniProt lists N-acetylserine at position 2 in its sequence numbering. Keep that coordinate explicit when comparing papers or antibody annotations, which may use different numbering. This modification alone does not demonstrate a detectable mass shift.
Does this guide establish induction of PYCR1?
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 PYCR1?
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 A06018-1 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 PYCR1 bands be quantified across samples?
Quantitation · Quantify a consistently identified band across samples, using the approximately 35 kDa observed band as a reference. If multiple bands appear, assess them separately before combining signals: three isoforms and two listed phosphoserines complicate assignment, but the supplied features do not establish which forms each band represents.
Why might PYCR1 appear near 35 kDa instead of 33.4 kDa?
Interpretation · The supplied apparent band is about 35 kDa, versus a predicted mass of 33.4 kDa. UniProt lists modifications and alternative isoforms, but their presence alone does not explain this difference or establish a visible shift. Use the observed band as a reference and assess additional bands separately.
PYCR1 is mitochondrial, but the supplied features list no signal peptide or propeptide and give no cleavage site. Do not assign a lower band to mitochondrial processing from location alone. Compare its reproducibility and antibody recognition before identifying it as PYCR1.
UniProt describes PYCR1 as a homodecamer composed of five homodimers. That supports oligomerization as a possibility to investigate, but does not establish that an oligomer survives Western blot sample preparation. Do not assign a higher band solely by matching a multiple of 33.4 kDa.
PYCR1 is mitochondrial, but the supplied features list no signal peptide or propeptide and give no cleavage site. Do not assign a lower band to mitochondrial processing from location alone. Compare its reproducibility and antibody recognition before identifying it as PYCR1.
UniProt describes PYCR1 as a homodecamer composed of five homodimers. That supports oligomerization as a possibility to investigate, but does not establish that an oligomer survives Western blot sample preparation. Do not assign a higher band solely by matching a multiple of 33.4 kDa.
Boster reagents
PYCR1 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-PYCR1 Antibody Picoband®
Cat # A06018-1
The catalog reports one anti-PYCR1 antibody for Western blotting, A06018-1, with reported human reactivity. Its WB image shows an approximately 35 kDa band in SW620, PANC-1, and 293T whole-cell lysates. No publication evidence or testing in other species is supplied.
Which to pick: A06018-1 is the only listed option. Choose it for human PYCR1 WB when its documented whole-cell lysate conditions fit your experiment; the supplied image covers SW620, PANC-1, and 293T samples at 30 µg per lane.
Source: BosterBio PYCR1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P32322.
- Human Protein Atlas. PYCR1 tissue expression.
- PMC11134742 — target-verified WB comparison
- PMC10918153 — target-verified WB comparison
- PMC7475402 — target-verified WB comparison