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- Table of Contents
FGF5 / Fibroblast growth factor 5 · Western blot design guide
Design a Western Blot for FGF5
Real validated FGF5 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-FGF5 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
FGF5 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 | ~29.6 kDa | |
| Gel | 12–15% (standard starting point) | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Glycosylated + Cleaved | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 2 isoform(s) |
Section 1
Real Curated FGF5 Western Blot Protocols
The A03776-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | mouse liver cells (catalog A03776-1) |
| Gel % | 12–15% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Short semi-dry transfer; verify retention (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | A03776-1; use the WB datasheet starting dilution (standard starting point) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | 1:20000 (catalog A03776-1) |
| 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 FGF5 Western Blot Band Size?
FGF5 has a predicted 29.6 kDa full-length mass; secretion, signal-peptide cleavage, glycosylation, and isoforms could affect detection or migration, but no empirical band size is supplied.
| Band near 29.6 kDa | consistent with the predicted full-length FGF5 precursor, pending identity controls |
| Band below 29.6 kDa | could reflect cleavage of the 1–20 signal peptide |
| Band above 29.6 kDa | could reflect N-linked glycosylation at Asn110; a visible shift is unproven |
| More than one band | could reflect Long and Short isoforms or signal-peptide processing; distinct migration is unproven |
| Little or no band in whole-cell lysate | consistent with secretion of FGF5 |
💡Expected FGF5 appearanceFGF5 has a predicted full-length mass of 29.6 kDa, but no empirical band size is supplied; confirm band identity because secretion, signal-peptide cleavage, glycosylation, and isoforms may affect what is detected.
| Predicted full-length mass | 29.6 kDa is the sequence-based precursor reference, not a measured band position |
| Signal peptide at residues 1–20 | cleavage yields a smaller mature protein than the full-length precursor |
| N-linked glycosylation site at Asn110 | glycosylation could change apparent size, but occupancy and a visible shift are unproven |
| Long and Short splice isoforms | may differ in size; their masses and electrophoretic separation are unspecified |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | FGF5 is secreted | test conditioned medium alongside lysate |
| Band higher than expected | possible N-linked glycosylation at Asn110 | compare untreated and deglycosylated samples and confirm band identity |
| Band lower than expected | possible cleavage of the 1–20 signal peptide | check whether the antibody recognizes the mature protein and confirm band identity |
| Multiple bands | Long and Short isoforms or signal-peptide processing may contribute | use isoform-aware or sequence-specific controls to identify the bands |
| Weak or no signal | secreted FGF5 may be scarce in the tested fraction | examine concentrated conditioned medium and include a positive control |
Experiment evidence
Western blot images showing FGF5 band
A03776-1 · FGF5
A03776-1 · FGF5
A03776 · FGF5
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for FGF5 Western blot
🧪For positive controls for FGF5 in Western blot, you can use a validated positive sample once one is identified; the supplied HPA evidence names none.
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 alongside.
⚠️Feasibility: FGF5 is secreted, so conditioned medium may give a clearer signal than whole-cell lysate; HPA provides no control samples.
HPA tissue expression evidence for FGF5
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 FGF5 Western Blot Tips
Deeper troubleshooting and optimisation questions for FGF5, answered from its protein features.
How should FGF5 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.
How do the Long and Short FGF5 isoforms differ?
Isoforms · In the Short isoform, residues 120..123 change from VLEI to QVHR, and residues 124..268 are absent. Interpret bands with these sequence differences in mind; the supplied features give no observed band positions for either isoform.
An epitope within residues 124..268 would be absent from the Short isoform. For detection of both isoforms, consider a shared region outside that deletion, accounting for the 120..123 substitution and the annotated signal peptide at 1..20.
An epitope within residues 124..268 would be absent from the Short isoform. For detection of both isoforms, consider a shared region outside that deletion, accounting for the 120..123 substitution and the annotated signal peptide at 1..20.
How can glycosylation affect FGF5 band interpretation?
PTM · UniProt annotates one N-linked glycosylation site at Asn110, using canonical sequence numbering. Comparing glycosylated and deglycosylated aliquots can help assess whether glycosylation contributes to a band pattern. The annotation does not establish that a particular band shift will be visible.
Does this guide establish induction of FGF5?
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 FGF5?
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 A03776-1 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.
What should be controlled when quantifying FGF5 bands?
Quantitation · Use the same sample fraction and an antibody whose recognized sequence is present in the isoforms being compared. If multiple bands appear, quantify each defined band consistently: the Long and Short isoforms differ substantially in sequence, and the supplied features do not identify an observed band for either.
Why might FGF5 migrate away from its predicted 29.6 kDa?
Interpretation · The 29.6 kDa prediction refers to the full-length sequence. UniProt annotates a signal peptide at residues 1..20 and N-linked glycosylation at Asn110. Processing or glycosylation could affect apparent migration, but these features alone do not establish a visible shift. No observed band size is supplied.
FGF5 is annotated as secreted and has a signal peptide at residues 1..20. Check conditioned medium when evaluating secreted FGF5, and keep the sample fraction consistent across comparisons.
Check whether the bands are consistent with the Long and Short isoform sequences, the annotated 1..20 signal peptide, or N-linked glycosylation at canonical Asn110. These features offer possibilities, but none identifies an unexpected band by itself; no empirical FGF5 band size is supplied.
FGF5 is annotated as secreted and has a signal peptide at residues 1..20. Check conditioned medium when evaluating secreted FGF5, and keep the sample fraction consistent across comparisons.
Check whether the bands are consistent with the Long and Short isoform sequences, the annotated 1..20 signal peptide, or N-linked glycosylation at canonical Asn110. These features offer possibilities, but none identifies an unexpected band by itself; no empirical FGF5 band size is supplied.
Boster reagents
FGF5 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-FGF-5 Antibody
Cat # A03776-1
Real WB data
Anti-Fibroblast growth factor 5 FGF5 Antibody
Cat # A03776
Both listed anti-FGF5 antibodies report human, mouse, and rat reactivity and have WB images. The supplied captions show mouse liver cells for A03776-1 and mouse and rat fetal brain lysates for A03776; no human WB example is provided.
Which to pick: For mouse liver cells, consider A03776-1, which has a WB image in that sample. For mouse or rat fetal brain lysate, consider A03776, shown at 1:500 with 40 µg per lane. Neither supplied image demonstrates human WB performance.
Source: BosterBio FGF5 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P12034.
- Human Protein Atlas. FGF5 tissue expression.
- PMC7417478 — target-verified WB comparison
- PMC9350999 — target-verified WB comparison
- PMC6628182 — target-verified WB comparison