This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
PEF1 / Peflin · Western blot design guide
Design a Western Blot for PEF1
Source-linked PEF1 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-PEF1 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
PEF1 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 | ~30.4 kDa | |
| Gel | 12–15% (standard starting point) | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Ubl conjugation | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked PEF1 Western Blot Protocol Options
The M09591 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | HeLa cell lysate (catalog M09591) |
| Gel % | 12–15% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Standard semi-dry transfer; verify efficiency (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | M09591; 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 PEF1 Western Blot Band Size?
PEF1 is predicted at 30.4 kDa; its listed features do not demonstrate altered Western blot migration, so confirm any candidate band with identity controls.
| Band near 30.4 kDa in whole-cell lysate | Consistent with the predicted size of PEF1; identity still needs confirmation |
| Band near 30.4 kDa in a cytosolic fraction | Consistent with cytoplasmic PEF1 |
| Band near 30.4 kDa in a membrane fraction | Consistent with peripheral membrane-associated PEF1 |
| Band near 30.4 kDa in a COPII vesicle fraction | Consistent with PEF1 at COPII-coated vesicle membranes |
💡Expected PEF1 appearancePEF1 is predicted at 30.4 kDa, with no empirical band size supplied; a band near that size is the starting expectation, subject to confirmation with band-identity controls.
| Predicted molecular mass | Places full-length PEF1 near 30.4 kDa |
| 284-amino-acid sequence | Defines the full-length protein underlying the predicted mass |
| PDCD6 heterodimerization | Does not establish a higher band on a Western blot |
| Calcium-dependent dissociation from PDCD6 | Does not establish a change in apparent band size |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Peripheral membrane or vesicle-associated PEF1 may be poorly recovered | Check extraction and compare soluble and membrane fractions |
| Weak or no signal | Incomplete recovery of cytoplasmic or membrane-associated PEF1 | Check fraction recovery and use a positive control |
| Band higher than expected | The listed features do not establish a higher-migrating PEF1 species | Confirm band identity with knockdown or a second antibody |
| Band lower than expected | No signal-peptide or propeptide cleavage is annotated | Confirm band identity with knockdown or a second antibody |
| Multiple bands | The single annotated isoform does not establish multiple PEF1 bands | Identify the PEF1-dependent band with knockdown |
Experiment evidence
Western blot images showing PEF1 band
M09591 · PEF1
A09591-2 · PEF1
A09591-2 · PEF1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for PEF1 Western blot
🧪For positive controls for PEF1 in Western blot, you can use no HPA-supported sample from the supplied evidence.
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: PEF1 is intracellular, but the missing HPA data prevent selection of tissue-based controls.
HPA tissue expression evidence for PEF1
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 PEF1 Western Blot Tips
Deeper troubleshooting and optimisation questions for PEF1, answered from its protein features.
Where should the main PEF1 band appear?
Band shift · PEF1 has a predicted mass of 30.4 kDa. Use that as a reference, but the supplied features provide no observed band position and cannot establish its apparent mass on a blot.
Could PEF1 isoforms explain multiple bands?
Isoforms · The supplied record lists one isoform and no alternative sequence, so it provides no isoform-based explanation for multiple bands.
Do the modification annotations predict a shifted band?
PTM · The keywords include ubiquitin-like conjugation and isopeptide bond, but the supplied site lists give no modification coordinates or enzyme. These annotations alone do not establish a visible shift or explain a mass difference.
Does this guide establish induction of PEF1?
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 PEF1 Western blot?
Transfer · PEF1 is predicted to be 30.4 kDa. Choose and verify transfer conditions suitable for a protein near that mass, using a molecular weight marker and a transfer check. The supplied features do not specify a transfer method.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the M09591 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 PEF1 be quantified?
Quantitation · Standard workflow guidance: quantify only a validated target band within the linear exposure range. Use consistent sample preparation and loading, retain biological replicates, and avoid interpreting saturation or loading differences as regulation.
Could calcium change the PEF1 signal?
Interpretation · PEF1 heterodimerizes with PDCD6 through EF-hand 5 and dissociates from PDCD6 in the presence of calcium. Keep calcium conditions consistent when comparing samples; the supplied features do not establish that this interaction changes the PEF1 band position.
PEF1 is annotated in the cytoplasm and at membranes, including COPII-coated vesicle membranes. Compare equivalent sample fractions and use a consistent loading normalization method; a change in one fraction may reflect localization rather than total abundance.
PEF1 is annotated as cytoplasmic and as a peripheral membrane protein at the endoplasmic reticulum and COPII-coated vesicles. Check which fractions were collected before interpreting a weak or absent signal.
Compare them with the 30.4 kDa predicted mass, but do not assign them to isoforms or a specific modification from these features. The record lists one isoform, and its modification-related keywords provide no mapped site or evidence of a visible band shift.
PEF1 is annotated in the cytoplasm and at membranes, including COPII-coated vesicle membranes. Compare equivalent sample fractions and use a consistent loading normalization method; a change in one fraction may reflect localization rather than total abundance.
PEF1 is annotated as cytoplasmic and as a peripheral membrane protein at the endoplasmic reticulum and COPII-coated vesicles. Check which fractions were collected before interpreting a weak or absent signal.
Compare them with the 30.4 kDa predicted mass, but do not assign them to isoforms or a specific modification from these features. The record lists one isoform, and its modification-related keywords provide no mapped site or evidence of a visible band shift.
Boster reagents
PEF1 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-PEF1 Rabbit Monoclonal Antibody
Cat # M09591
Real WB data
Anti-PEF1 Antibody Picoband®
Cat # A09591-2
Both listed anti-PEF1 antibodies have WB images and reported human, mouse, and rat reactivity. M09591 is shown with HeLa lysate; A09591-2 shows an approximately 30 kDa band in eight human samples. No mouse or rat WB samples are shown.
Which to pick: For a HeLa example, consider M09591. For documented conditions across several human sample types, consider A09591-2: its image uses 50 µg of sample per lane under reducing conditions and 0.5 µg/mL primary antibody. Both list mouse and rat reactivity.
Source: BosterBio PEF1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q9UBV8.
- Human Protein Atlas. PEF1 tissue expression.
- PMC8884472 — target-verified WB comparison
- PMC8728108 — target-verified WB comparison
- PMC8024055 — target-verified WB comparison