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- Table of Contents
MUC16 / Mucin-16 · Western blot design guide
Design a Western Blot for MUC16
Real validated MUC16 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-MUC16 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
MUC16 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 | ~1519.2 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 | Glycosylated + Phosphorylated | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Real Curated MUC16 Western Blot Protocols
The M01641 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | Human ovary cancer lysate (catalog M01641) |
| 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 | M01641; 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 MUC16 Western Blot Band Size?
MUC16 has a predicted mass of 1519.2 kDa; glycosylation and cleavage may affect migration, but no empirical band size demonstrates either effect.
| Band near 1519.2 kDa | Could represent the full-length MUC16 polypeptide; identity requires confirmation |
| Band above 1519.2 kDa | N-linked glycosylation could affect migration, but the shift is unmeasured |
| Band below 1519.2 kDa | Could reflect proteolytic cleavage of MUC16; fragment size is unknown |
| Little or no band in whole-cell lysate | MUC16 may have been released into the extracellular space |
💡Expected MUC16 appearanceMUC16 has a predicted full-length polypeptide mass of 1519.2 kDa, but no empirical band size is supplied; glycosylation or cleavage may affect migration, so confirm band identity with appropriate controls.
| Predicted full-length mass | 1519.2 kDa is the calculated polypeptide mass, not a measured band position |
| N-linked glycosylation at Asn139 | Could affect apparent size if occupied; the shift is unknown |
| N-linked glycosylation at Asn434 | Could affect apparent size if occupied; the shift is unknown |
| N-linked glycosylation at Asn787 | Could affect apparent size if occupied; the shift is unknown |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | MUC16 may be released into the extracellular space | Check conditioned medium alongside lysate |
| Band higher than expected | N-linked glycosylation may alter migration | Compare untreated and deglycosylated samples |
| Band lower than expected | Proteolytic cleavage may produce a smaller MUC16 fragment | Compare bands detected by antibodies against different regions |
| Broad smear instead of sharp band | Variable glycosylation across numerous N-linked sites is possible | Compare untreated and deglycosylated samples |
| Multiple bands | Full-length and cleaved MUC16 forms may coexist | Compare lysate and conditioned medium using antibodies against different regions |
| Fragments below expected size | Proteolytic cleavage may release the extracellular domain | Check fragment identity with antibodies against different regions |
Experiment evidence
Western blot images showing MUC16 band
M01641 · MUC16
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for MUC16 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for MUC16 in Western blot, you can use bronchus tissue, which shows high HPA expression.
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: MUC16 can shed its extracellular domain, so whole-cell lysate may miss that signal; consider conditioned medium.
HPA tissue expression evidence for MUC16
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 | goblet cells | High | Protein (IHC) | HPA → |
| Cervix | glandular cells | High | Protein (IHC) | HPA → |
| Endometrium | glandular cells | High | Protein (IHC) | HPA → |
| Fallopian tube | non-ciliated cells | High | Protein (IHC) | HPA → |
| Salivary gland | glandular cells | Medium | 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 → |
| Appendix | glandular cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | adipocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced MUC16 Western Blot Tips
Deeper troubleshooting and optimisation questions for MUC16, answered from its protein features.
How should MUC16 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 MUC16 isoforms explain multiple bands?
Isoforms · The supplied features list one isoform and no alternative sequence. They therefore provide no specific isoform explanation for multiple bands. Compare bands using antibodies against defined regions of MUC16 and consider its documented extracellular-domain release when interpreting different signals.
Which glycosylation sites matter when interpreting a MUC16 band?
PTM · UniProt lists 102 N-linked asparagine sites, including positions 139, 434, and 14423 in its sequence numbering. Check whether the antibody’s recognized region contains listed sites when interpreting a change in band position. The site list does not show which sites are occupied in your sample or prove that glycosylation caused a shift.
UniProt notes that phosphorylation of the intracellular C-terminus may induce proteolytic cleavage and release of the extracellular domain. If the signal changes after a treatment, compare membrane-associated and extracellular samples and identify the antibody’s recognized region. The supplied features do not specify a stimulus or phosphorylation coordinate.
UniProt notes that phosphorylation of the intracellular C-terminus may induce proteolytic cleavage and release of the extracellular domain. If the signal changes after a treatment, compare membrane-associated and extracellular samples and identify the antibody’s recognized region. The supplied features do not specify a stimulus or phosphorylation coordinate.
Does this guide establish induction of MUC16?
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 MUC16?
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 M01641 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.
What should be controlled when quantifying MUC16 by Western blot?
Quantitation · Quantify a defined band in the same sample fraction across lanes and report the antibody’s recognized region. MUC16 can be membrane-associated or released extracellularly, so a change in one fraction need not represent a change in total MUC16. Verify that the chosen signal is within the assay’s linear range.
Should MUC16 appear at its predicted 1519.2 kDa mass?
Interpretation · 1519.2 kDa is the calculated mass of the 14,507-residue sequence, not a measured Western-blot position. MUC16 has 102 listed N-linked glycosylation sites, but those features alone cannot establish its apparent mass or a visible shift. Do not assign a band solely by its distance from 1519.2 kDa.
MUC16 is listed as a single-pass type I membrane protein and as extracellular after possible release of its extracellular domain. A membrane sample may contain membrane-associated MUC16, while an extracellular sample may contain released material. Record the sampled fraction and antibody region before treating different bands as the same species.
Proteolytic release of the extracellular domain is documented as a possibility, so a smaller signal could warrant checking which side of the cleavage the antibody recognizes and where the sample was collected. The supplied features give no fragment sizes or cleavage coordinate; band size alone cannot identify a fragment.
MUC16 is listed as a single-pass type I membrane protein and as extracellular after possible release of its extracellular domain. A membrane sample may contain membrane-associated MUC16, while an extracellular sample may contain released material. Record the sampled fraction and antibody region before treating different bands as the same species.
Proteolytic release of the extracellular domain is documented as a possibility, so a smaller signal could warrant checking which side of the cleavage the antibody recognizes and where the sample was collected. The supplied features give no fragment sizes or cleavage coordinate; band size alone cannot identify a fragment.
Boster reagents
MUC16 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-MUC16/Ca125 Rabbit Monoclonal Antibody
Cat # M01641
The catalog reports one anti-MUC16 antibody for Western blotting: M01641, a rabbit monoclonal with stated human reactivity. Its WB image shows MUC16 expression in human ovary cancer lysate; no other sample contexts are documented here.
Which to pick: M01641 is the only listed option. It has a WB image using human ovary cancer lysate, making it the documented choice for that sample context. Check suitability separately for other samples or species.
Source: BosterBio MUC16 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q8WXI7.
- Human Protein Atlas. MUC16 tissue expression.
- PMC4918949 — target-verified WB comparison
- PMC6492407 — target-verified WB comparison
- PMC4234371 — target-verified WB comparison