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- Table of Contents
COP1 / E3 ubiquitin-protein ligase COP1 · Western blot design guide
Design a Western Blot for COP1
Real validated COP1 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-COP1 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
COP1 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 | ~80.5 kDa | |
| Gel | 8–10% (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 C7 membership | |
| Isoform | 5 isoform(s) |
Section 1
Real Curated COP1 Western Blot Protocols
The A31677 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | extracts of various cell lines, (catalog A31677) |
| Gel % | 8–10% (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 | Blocking buffer: 3% nonfat dry milk in TBST (catalog A31677) |
| Primary antibody | A31677 · 1:1000 (catalog A31677) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | Goat Anti-Rabbit IgG, 1:10000 (catalog A31677) |
| Secondary incubation | 1 h at room temperature (standard starting point) |
| Wash | 3 × 5 min in TBST (standard starting point) |
| Detection | ECL (catalog A31677) |
Section 2
What Is the Expected COP1 Western Blot Band Size?
COP1 has a predicted monomer size of 80.5 kDa; isoforms and homodimerization could affect migration, but no empirical band size demonstrates these effects.
| Band near 80.5 kDa | Consistent with the predicted COP1 monomer; confirm band identity |
| Band near 161 kDa | Could reflect a homodimer that survives sample denaturation |
| Several bands at different positions | Could reflect COP1 isoforms 1 through 5; their migration differences are unknown |
| Bands in nuclear and cytoplasmic fractions | Consistent with COP1 localization in both compartments |
💡Expected COP1 appearanceUniProt predicts an 80.5 kDa COP1 monomer, but no empirical band size is supplied; confirm any candidate band with appropriate identity controls.
| UniProt predicted mass | Places the predicted monomer near 80.5 kDa |
| Coiled coil mediated homodimer | Could produce a band near twice the monomer mass if the dimer survives sample denaturation |
| Splice isoforms 1, 2 and 3 | May differ in apparent size; individual masses and migration differences are not supplied |
| Splice isoforms 4 and 5 | May differ in apparent size; individual masses and migration differences are not supplied |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | COP1 signal may be below detection in the sampled lysate | Check sample loading and antibody performance with a positive control |
| Band higher than expected | The coiled coil mediated homodimer may persist during sample preparation | Compare thoroughly denatured samples and verify band identity |
| Band lower than expected | A COP1 splice isoform may migrate differently; its mass is unknown | Compare with an independent COP1 antibody or COP1 depleted sample |
| Multiple bands | COP1 has five named splice isoforms, but distinct bands are unproven | Check which bands respond to COP1 depletion |
| Weak or no signal | COP1 is distributed between nuclear and cytoplasmic compartments | Check fraction recovery and sample loading, then compare with a positive control |
Experiment evidence
Western blot images showing COP1 band
A31677 · COP1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for COP1 Western blot
🧪For positive controls for COP1 in Western blot, you can use a validated COP1-positive cell lysate; HPA provides no positive tissue or cell line to name.
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 such as Ponceau alongside the samples.
⚠️Feasibility: HPA provides no tissue data, so a positive sample needs independent validation and siRNA knockdown or a KO line is needed for a negative control.
HPA tissue expression evidence for COP1
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 COP1 Western Blot Tips
Deeper troubleshooting and optimisation questions for COP1, answered from its protein features.
How should COP1 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 COP1 isoforms produce smaller bands?
Isoforms · Yes. Five isoforms are listed. Relative to the canonical sequence, isoform 2 lacks residues 211–214 and 277–296; isoform 4 lacks 277–296. Isoform 3 lacks 1–264 and has a replacement at 342. Isoform 5 has Y157E and lacks 158–731. These differences could affect band size, but do not establish where any isoform migrates.
Check the antibody’s epitope against the listed sequence changes. An epitope within residues 277–296 would be absent from isoforms 2 and 4; one within 158–731 would be absent from isoform 5. UniProt residue numbers here refer to the canonical sequence and may differ from antibody numbering.
Check the antibody’s epitope against the listed sequence changes. An epitope within residues 277–296 would be absent from isoforms 2 and 4; one within 158–731 would be absent from isoform 5. UniProt residue numbers here refer to the canonical sequence and may differ from antibody numbering.
Does COP1 modification explain a band shift?
PTM · The supplied features list no modified residues or glycosylation sites. They therefore do not support assigning a band shift to a specific modification. Compare the band with the 80.5 kDa predicted canonical mass and consider the listed isoform sequences, without treating either as proof of its identity.
Does this guide establish induction of COP1?
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 COP1?
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 A31677 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 can localization affect COP1 quantitation?
Quantitation · COP1 is listed in both the cytoplasm and nuclear speckles. Keep the sample fraction consistent across comparisons, and state whether measurements come from whole cell, nuclear, or cytoplasmic material. Differences between fractions can reflect COP1 localization as well as abundance.
What mass should I expect for COP1?
Interpretation · The supplied predicted mass is 80.5 kDa for canonical COP1. No observed band position is available, so use 80.5 kDa as a reference rather than an expected apparent mass.
Compare its size with the five listed isoforms and check whether the antibody epitope is retained. COP1 is also described as a homodimer and a component of a ubiquitin ligase complex, but those associations alone do not identify an unexpected Western blot band.
Compare its size with the five listed isoforms and check whether the antibody epitope is retained. COP1 is also described as a homodimer and a component of a ubiquitin ligase complex, but those associations alone do not identify an unexpected Western blot band.
Boster reagents
COP1 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-RFWD2 COP1 Antibody
Cat # A31677
A31677 is the listed anti-COP1 antibody for Western blotting, with reported Human, Mouse, and Rat reactivity. Its WB image uses extracts from various cell lines at 1:1000 with 25 µg per lane; the supplied caption does not identify the cell lines.
Which to pick: A31677 is the only listed option and has a WB image. Choose it based on the reported reactivity and the shown 1:1000, 25 µg-per-lane conditions; confirm performance in your own sample.
Source: BosterBio COP1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q8NHY2.
- Human Protein Atlas. COP1 tissue expression.
- PMC12089050 — target-verified WB comparison