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- Table of Contents
C6 / Complement component C6 · Western blot design guide
Design a Western Blot for C6
Real validated C6 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-C6 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
C6 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 | ~104.8 kDa | |
| Gel | 8–10% (standard starting point) | |
| Positive control | Kidney (IHC candidate; verify WB) +2 more | |
| Negative control | Adrenal gland (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Glycosylated + Cleaved | |
| Caveat | Extensive glycosylation | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Real Curated C6 Western Blot Protocols
The A00317 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | various cells (catalog A00317) |
| Gel % | 8–10% (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 | A00317 · 1:1000 (catalog A00317) |
| 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 C6 Western Blot Band Size?
C6 has a 104.8 kDa precursor backbone; signal-peptide cleavage lowers its polypeptide mass, while multiple glycans can shift and broaden the mature band.
| Band near 105 kDa or somewhat higher | C6 monomer with glycosylation; 104.8 kDa is the unglycosylated precursor mass |
| Broad band or smear above 105 kDa | C6 glycoforms with differing carbohydrate content |
| Band slightly below the full-length precursor | Signal peptide removal reduces the C6 polypeptide mass |
| Little or no band in whole-cell lysate | C6 is secreted as a soluble protein |
💡Expected C6 appearanceExpect secreted, signal-peptide-cleaved C6 as a glycosylated band near or above its 104.8 kDa precursor mass, potentially broadened by variable glycosylation; UniProt features do not establish an exact apparent size.
| 104.8 kDa predicted precursor | Provides the full-length polypeptide reference before processing and glycosylation |
| Signal peptide residues 1–21 | Cleavage makes the mature polypeptide smaller than the precursor |
| N-linked glycosylation at Asn324 | Adds carbohydrate and can raise apparent mass |
| O-linked glycosylation at Thr38 and Thr392 | Adds carbohydrate and can broaden or raise the band |
| C-linked mannosylation at Trp29 | Adds carbohydrate to the mature protein |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | C6 is secreted | Examine conditioned medium or another extracellular sample |
| Band higher than expected | C6 carries multiple glycans | Compare with a deglycosylated aliquot where the relevant glycans can be removed |
| Band lower than expected | The 1–21 signal peptide is removed | Compare with the expected mature polypeptide rather than only the precursor mass |
| Broad smear instead of sharp band | Heterogeneous C6 glycosylation | Compare treated and untreated aliquots for glycan-dependent shifts |
| Multiple bands | Distinct glycoforms or precursor and mature C6 | Compare sample compartments and glycan-dependent mobility |
| Weak or no signal | Secreted C6 may be scarce in cellular material | Enrich extracellular material and check sample loading |
| Fragments below expected size | C6 may have undergone proteolysis during sample handling | Check sample integrity and use protease inhibitors during preparation |
Experiment evidence
Western blot images showing C6 band
A00317 · C6
A00317 · C6
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for C6 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for C6 in Western blot, you can use kidney tissue, which has high HPA expression.
Positive control: Kidney (IHC candidate; verify WB)
Negative control: Adrenal gland (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: C6 is secreted, so tissue lysates may give weak signal; consider extracellular protein fractions.
HPA tissue expression evidence for C6
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 |
|---|---|---|---|---|
| Kidney | proximal tubules (microvilli) | High | Protein (IHC) | HPA → |
| Breast | glandular cells | Medium | Protein (IHC) | HPA → |
| Fallopian tube | ciliated cells (ciliary rootlets) | Medium | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adrenal gland | glandular cells | Not detected | Protein (IHC) | HPA → |
| Appendix | glandular cells | Not detected | Protein (IHC) | HPA → |
| Bronchus | basal cells | Not detected | Protein (IHC) | HPA → |
| Caudate | glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | cells in granular layer | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced C6 Western Blot Tips
Deeper troubleshooting and optimisation questions for C6, answered from its protein features.
How should C6 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.
Should multiple C6 bands be assigned to isoforms?
Isoforms · Only one C6 isoform is listed. Multiple bands alone do not establish isoforms; assess processing, glycosylation, or association with other proteins before assigning identities.
Could glycosylation affect C6 band interpretation?
PTM · Yes. Ten glycosylation sites are listed, so glycosylation may contribute to a shifted or broad band. Interpret mobility alongside the 104.8 kDa predicted mass and the signal peptide, rather than assuming an exact match.
Does this guide establish induction of C6?
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 C6 Western blot?
Transfer · C6 has a predicted mass of 104.8 kDa. Check that transfer conditions retain and move proteins near this size, and verify transfer in the relevant region before interpreting a weak or absent C6 band.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A00317 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.
Where should C6 be measured for quantitation?
Quantitation · C6 is secreted as a soluble protein and also participates in the membrane attack complex at target cell membranes. Choose the sample fraction that matches the question, and compare like fractions across samples.
Why might C6 migrate differently from its predicted 104.8 kDa mass?
Interpretation · C6 has a 21-residue signal peptide and 10 glycosylation sites. Signal peptide removal and glycosylation can affect its apparent size. No observed band size is supplied, so use 104.8 kDa as a starting reference, not an exact band position.
C6 has 32 listed disulfide bonds. Keep reduction conditions consistent across samples because disulfide-dependent structure can affect migration. Compare bands prepared under the same conditions.
C6 is a component of the membrane attack complex with C5b, C7, C8, and C9. A higher-mass signal could reflect associated material, but band position alone cannot establish complex identity. Interpret it in light of sample fraction and preparation conditions.
C6 has 32 listed disulfide bonds. Keep reduction conditions consistent across samples because disulfide-dependent structure can affect migration. Compare bands prepared under the same conditions.
C6 is a component of the membrane attack complex with C5b, C7, C8, and C9. A higher-mass signal could reflect associated material, but band position alone cannot establish complex identity. Interpret it in light of sample fraction and preparation conditions.
Boster reagents
C6 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-Complement component C6 C6 Antibody
Cat # A00317
The catalog reports one anti-C6 antibody, A00317. Its catalog includes a Western blot image of various cells using the polyclonal antibody at 1:1000. No publication evidence or additional validation is supplied here.
Which to pick: A00317 is the only listed option for C6 and has a Western blot image. Check its product details for suitability for your sample, since reactivity is not specified in the supplied evidence.
Source: BosterBio C6 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P13671.
- Human Protein Atlas. C6 tissue expression.
- PMC7335182 — target-verified WB comparison