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- Table of Contents
CYC1 / Cytochrome c1, heme protein, mitochondrial · Western blot design guide
Design a Western Blot for CYC1
Real validated CYC1 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-CYC1 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
CYC1 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 | ~35.4 kDa | |
| Observed band | ~33 kDa | |
| Gel | 5–20% (catalog A02958-2) | |
| Positive control | Adrenal gland (IHC candidate; verify WB) +4 more | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 1 isoform(s) |
Section 1
Real Curated CYC1 Western Blot Protocols
The A02958-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human A431, human Hela, human MCF-7 (catalog A02958-2) |
| Gel % | 5–20% (catalog A02958-2) |
| Load | 30 ug; reducing conditions (catalog A02958-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A02958-2) |
| Membrane | nitrocellulose membrane (catalog A02958-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A02958-2) |
| Primary antibody | A02958-2 · 0.5 μg/mL (catalog A02958-2) |
| Primary incubation | overnight at 4°C (catalog A02958-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A02958-2) |
| Secondary incubation | 1.5 hour at RT (catalog A02958-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A02958-2) |
| Detection | ECL (catalog A02958-2) |
Section 2
What Is the Expected CYC1 Western Blot Band Size?
CYC1 is predicted at 35.4 kDa and observed at ~33 kDa on reducing Western blots; the cause of the difference is not established.
| Band at ~33 kDa | Empirical CYC1 band reported in reducing whole-cell lysates; confirm identity with antibody controls |
| Band near 35.4 kDa | Consistent with the predicted CYC1 mass, though migration may differ |
| Weak band in poorly solubilized lysate | CYC1 is a single-pass mitochondrial inner-membrane protein |
| Close doublet near ~33 kDa | Ser182 phosphorylation is documented, but its effect on migration is unestablished |
💡Expected CYC1 appearanceUniProt predicts 35.4 kDa, while antibody QC reports a ~33 kDa band in reducing whole-cell lysates; the difference is unexplained, so confirm band identity with appropriate controls.
| Predicted CYC1 mass | 35.4 kDa is the sequence-based reference, versus the observed ~33 kDa band |
| Phosphoserine at Ser182 | A modification is documented, but no apparent-size change is established |
| Single-pass inner-membrane location | May affect recovery; no apparent-size change is established |
| Complex III membership | CYC1 belongs to a multisubunit complex; no SDS-PAGE size effect is established |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Poor recovery of mitochondrial inner-membrane CYC1 | Check membrane solubilization and use a mitochondrial fraction as a control |
| Band higher than expected | Identity or migration of the higher band is unestablished | Compare reducing samples with the ~33 kDa reference and verify antibody specificity |
| Band lower than expected | The reported ~33 kDa band runs below the 35.4 kDa prediction for an unknown reason | Compare with the reported band and confirm identity using an independent antibody or CYC1 depletion |
| Multiple bands | Additional bands are not explained by listed isoforms | Identify the ~33 kDa band using CYC1 depletion or an independent antibody |
| Weak or no signal | Incomplete extraction of single-pass mitochondrial membrane CYC1 | Optimize membrane solubilization and check sample loading |
Experiment evidence
Western blot images showing CYC1 band
A02958-2 · CYC1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for CYC1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for CYC1 in Western blot, you can use adrenal gland tissue.
Positive control: Adrenal gland (IHC candidate; verify WB)
Negative control: Suggested KO / knockdown lysate
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: No tissue is reported as not detected, so use siRNA knockdown or a KO line for a negative control.
HPA tissue expression evidence for CYC1
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 |
|---|---|---|---|---|
| Adrenal gland | glandular cells | High | Protein (IHC) | HPA → |
| Appendix | glandular cells | High | Protein (IHC) | HPA → |
| Breast | glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Cerebellum | Purkinje cells | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Caudate | glial cells | Low | Protein (IHC) | HPA → |
| Adipose tissue | adipocytes | Medium | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | Medium | Protein (IHC) | HPA → |
| Cerebral cortex | endothelial cells | Medium | Protein (IHC) | HPA → |
| Cervix | glandular cells | Medium | Protein (IHC) | HPA → |
Section 3
Advanced CYC1 Western Blot Tips
Deeper troubleshooting and optimisation questions for CYC1, answered from its protein features.
How should CYC1 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 CYC1 isoforms explain multiple bands?
Isoforms · Only one isoform is listed, with no alternative sequence supplied. The supplied features therefore do not support assigning additional bands to CYC1 isoforms.
Could phosphorylation affect the CYC1 band?
PTM · UniProt lists phosphoserine at position 182, using its sequence numbering. Phosphorylation is a possible source of altered migration, but this site alone does not demonstrate a visible shift. Do not assign a second band to phosphorylated CYC1 without further validation.
Does this guide establish induction of CYC1?
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 CYC1 Western blot?
Transfer · CYC1 is listed as a single-pass mitochondrial inner-membrane protein. Use transfer conditions suited to a membrane protein and check transfer efficiency around the reported 33 kDa band. The supplied features do not specify a membrane type or transfer settings.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A02958-2 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 CYC1 bands be quantified?
Quantitation · Quantify a consistently identified CYC1 band near the reported 33 kDa position, using comparable sample preparation and loading across lanes. CYC1 is a mitochondrial inner-membrane component of complex III, so interpret changes alongside an appropriate measure of mitochondrial material.
Why might CYC1 appear near 33 kDa instead of 35.4 kDa?
Interpretation · The predicted mass is 35.4 kDa, while the reported apparent band is about 33 kDa. CYC1 has a transit-peptide keyword, but no cleavage site or processed mass is supplied. These features do not establish the cause of the difference; use the observed band as a reference and verify its identity.
Compare them with the reported band near 33 kDa before assigning an identity. The supplied record lists one isoform and phosphoserine at UniProt position 182, but neither establishes the identity of another band. CYC1’s membership in multisubunit complex III also does not identify an unexpected band on its own.
Compare them with the reported band near 33 kDa before assigning an identity. The supplied record lists one isoform and phosphoserine at UniProt position 182, but neither establishes the identity of another band. CYC1’s membership in multisubunit complex III also does not identify an unexpected band on its own.
Boster reagents
CYC1 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-CYC1 Antibody Picoband®
Cat # A02958-2
A02958-2 is a human-reactive anti-CYC1 antibody with a Western blot image from A431, HeLa, and MCF-7 whole-cell lysates. The caption reports a band near 33 kDa versus the expected 35 kDa; no independent validation is supplied.
Which to pick: Only A02958-2 is listed. Choose it for human CYC1 Western blots when its documented whole-cell lysate conditions fit your experiment. Performance in other species or specimen types is not established here.
Source: BosterBio CYC1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P08574.
- Human Protein Atlas. CYC1 tissue expression.
- PMC4261936 — target-verified WB comparison
- PMC6202082 — target-verified WB comparison
- PMC9926390 — target-verified WB comparison