This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
CENPE / Centromere-associated protein E · Western blot design guide
Design a Western Blot for CENPE
Real validated CENPE Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-CENPE 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
CENPE 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 | ~316.4 kDa | |
| Gel | 4–12% gradient (standard starting point) | |
| Positive control | Bronchus (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 + Methylated | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 2 isoform(s) |
Section 1
Real Curated CENPE Western Blot Protocols
The M04553-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | HepG2 cell lysate (catalog M04553-1) |
| Gel % | 4–12% gradient (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 | M04553-1; 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 CENPE Western Blot Band Size?
CENPE is predicted at 316.4 kDa; propeptide cleavage and isoforms could affect migration, but no distinct band size is demonstrated.
| Band near 316.4 kDa | Consistent with the predicted CENPE precursor, pending band-identity controls |
| Band marginally below 316.4 kDa | Could reflect cleavage of the 2699–2701 propeptide, though a resolvable shift is not established |
| Several bands near the predicted size | Isoforms 1 and 3 are possible contributors, but distinct migration is not established |
| One band without a resolved doublet | Compatible with isoforms whose migration is not known to differ |
💡Expected CENPE appearanceCENPE has a predicted precursor mass of 316.4 kDa; propeptide cleavage and isoforms could affect migration, but no empirical band size is supplied, so confirm band identity with appropriate controls.
| Predicted precursor mass | Places the full-length CENPE reference near 316.4 kDa |
| Propeptide at residues 2699–2701 | Cleavage would remove three residues; a resolvable shift is not established |
| Isoform 1 | Its apparent size relative to isoform 3 is not supplied |
| Isoform 3 | Its apparent size relative to isoform 1 is not supplied |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | CENPE is associated with kinetochores and the mitotic spindle | Check sample quality and compare mitotic enrichment with an appropriate positive control |
| Band higher than expected | No supplied feature establishes an upward shift | Check size markers and verify band identity by CENPE depletion |
| Band lower than expected | Propeptide cleavage removes only three residues; a substantial shift has no established cause | Check band identity by CENPE depletion and assess sample degradation |
| Multiple bands | Isoforms 1 and 3 are annotated, but distinct migration is unproven | Test which bands respond to CENPE depletion |
| Weak or no signal | CENPE localization to kinetochores and spindle may limit signal in the sampled cells | Check loading, antibody performance, and a mitotic positive control |
| Fragments below expected size | Protein breakdown is possible; fragment masses are not supplied | Use protease inhibitors and verify CENPE identity by depletion |
Experiment evidence
Western blot images showing CENPE band
M04553-1 · CENPE
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for CENPE Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for CENPE in Western blot, you can use bronchus tissue, which HPA scores High.
Positive control: Bronchus (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: CENPE signal may depend on mitotic-cell abundance because it localizes to kinetochores and spindles.
HPA tissue expression evidence for CENPE
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 | respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Cervix | squamous epithelial cells | High | Protein (IHC) | HPA → |
| Epididymis | glandular cells | High | Protein (IHC) | HPA → |
| Kidney | distal tubules | High | Protein (IHC) | HPA → |
| Nasopharynx | respiratory epithelial cells | 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 → |
| Adrenal gland | glandular cells | Not detected | Protein (IHC) | HPA → |
| Caudate | glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Cerebral cortex | endothelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CENPE Western Blot Tips
Deeper troubleshooting and optimisation questions for CENPE, answered from its protein features.
How should CENPE 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.
Could CENPE isoforms produce different bands?
Isoforms · UniProt lists isoforms 1 and 3. Relative to the canonical sequence, isoform 3 lacks residues 549..573 and replaces residues 1972..2068 with Q. These changes affect predicted size and may affect antibody recognition; they do not establish where either isoform runs on a blot.
Which CENPE phosphorylation sites matter when interpreting bands?
PTM · UniProt annotates phosphoserines at positions 611, 2083, 2389, 2639, 2647 and 2651. If testing phosphorylation, compare appropriately treated samples and specify that these are UniProt coordinates; antibody or paper numbering may differ. These annotations alone do not demonstrate a visible band shift.
Does this guide establish induction of CENPE?
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 CENPE Western blot?
Transfer · CENPE is predicted to be 316.4 kDa. Optimize transfer for a protein of this size and check transfer with a high-mass marker and membrane stain. The supplied features do not prescribe a specific transfer method.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the M04553-1 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 CENPE be quantified across samples?
Quantitation · CENPE is annotated with cell-cycle and mitosis keywords and localizes to kinetochores and the spindle. Compare samples with matched cell-cycle conditions, and use consistent lysate preparation and normalization. These annotations do not establish a specific change in abundance during the cell cycle.
Should CENPE run exactly at its predicted mass?
Interpretation · The predicted mass is 316.4 kDa, but no observed band position is supplied. Use a high-mass marker to assess migration. The listed modifications and sequence variants alone do not establish a visible shift or explain any difference from the predicted mass.
UniProt annotates a cysteine methyl ester at position 2698 and a propeptide at 2699..2701. Check whether a C-terminal antibody targets this region and whether its epitope is compatible with the annotated features. The supplied data do not establish how these features affect the blot.
First compare their positions with the 316.4 kDa prediction and consider the two annotated isoforms. Confirm antibody recognition and examine whether sample preparation or transfer affects the high-mass band. The supplied features do not identify any observed band or prove that a smaller band is an isoform or cleavage product.
UniProt annotates a cysteine methyl ester at position 2698 and a propeptide at 2699..2701. Check whether a C-terminal antibody targets this region and whether its epitope is compatible with the annotated features. The supplied data do not establish how these features affect the blot.
First compare their positions with the 316.4 kDa prediction and consider the two annotated isoforms. Confirm antibody recognition and examine whether sample preparation or transfer affects the high-mass band. The supplied features do not identify any observed band or prove that a smaller band is an isoform or cleavage product.
Boster reagents
CENPE 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-CENPE Monoclonal Antibody
Cat # M04553-1
The catalog reports one anti-CENPE antibody for Western blot: M04553-1, a monoclonal antibody with stated human reactivity. Its WB image shows CENPE expression in HepG2 cell lysate; evidence for other samples is not supplied.
Which to pick: M04553-1 is the only listed option. It has a WB image from HepG2 cell lysate and stated human reactivity; assess suitability for other sample types separately.
Source: BosterBio CENPE gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q02224.
- Human Protein Atlas. CENPE tissue expression.
- PMC8799097 — target-verified WB comparison
- PMC12341428 — target-verified WB comparison
- PMC10427915 — target-verified WB comparison