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- Table of Contents
FRY / Protein furry homolog · Western blot design guide
Design a Western Blot for FRY
Real validated FRY Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-FRY 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
FRY 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 | ~338.9 kDa | |
| Observed band | ~339 kDa | |
| Gel | 5–20% (catalog A01003) | |
| Positive control | Adrenal gland (IHC candidate; verify WB) +4 more | |
| Negative control | Oral mucosa (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Real Curated FRY Western Blot Protocols
The A01003 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human RT4, human Caco-2, rat brain (catalog A01003) |
| Gel % | 5–20% (catalog A01003) |
| Load | 30 ug; reducing conditions (catalog A01003) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A01003) |
| Membrane | nitrocellulose membrane (catalog A01003) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A01003) |
| Primary antibody | A01003 · 0.5 μg/mL (catalog A01003) |
| Primary incubation | overnight at 4°C (catalog A01003) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A01003) |
| Secondary incubation | 1.5 hour at RT (catalog A01003) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A01003) |
| Detection | ECL (catalog A01003) |
Section 2
What Is the Expected FRY Western Blot Band Size?
FRY is predicted at 338.9 kDa and observed at ~339 kDa; phosphorylation is documented, but its effect on migration is not established.
| Band at ~339 kDa | Matches the empirical FRY band and its 338.9 kDa predicted mass. |
| Single band near 339 kDa | Consistent with full-length FRY; confirm identity with antibody controls. |
| Slightly shifted band near 339 kDa | FRY phosphorylation could affect migration, but a visible shift is unproven. |
| Close doublet near 339 kDa | Different phosphorylation states are possible; band identities require confirmation. |
💡Expected FRY appearanceFRY has a predicted mass of 338.9 kDa and an empirical band at ~339 kDa; confirm band identity with appropriate antibody controls.
| Predicted FRY mass | Places full-length FRY near 338.9 kDa, consistent with the observed ~339 kDa band. |
| Phosphotyrosine at residue 213 | May affect migration; a visible size change is not established. |
| Phosphoserines at residues 1382 and 1383 | May affect migration; a visible size change is not established. |
| Phosphoserines at residues 2419 and 2420 | May affect migration; a visible size change is not established. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Cytoplasmic FRY was not detected in the sample. | Check a positive-control lysate, loading, and transfer of high-mass proteins. |
| Band higher than expected | Phosphorylation is possible, but its effect on FRY migration is unproven. | Compare phosphatase-treated material and verify band identity. |
| Band lower than expected | A smaller band is not explained by a listed cleavage feature. | Check sample integrity and confirm identity with another antibody. |
| Multiple bands | Different phosphorylation states are possible, but band identities are unproven. | Compare phosphatase-treated material and use an independent FRY antibody. |
| Weak or no signal | Low recovery or inefficient transfer of this 338.9 kDa cytoplasmic protein. | Check lysate loading, transfer, and a positive-control sample. |
| Fragments below expected size | Sample degradation is possible; no FRY cleavage product is specified. | Prepare lysate with protease inhibitors and confirm fragments with another antibody. |
Experiment evidence
Western blot images showing FRY band
A01003 · FRY
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for FRY Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for FRY in Western blot, you can use adrenal gland lysate, which HPA scores High.
Positive control: Adrenal gland (IHC candidate; verify WB)
Negative control: Oral mucosa (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: FRY is cytoplasmic, so tissue lysates are suitable; oral mucosa is an HPA Not-detected comparator.
HPA tissue expression evidence for FRY
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 → |
| Gallbladder | glandular cells | High | Protein (IHC) | HPA → |
| Kidney | cells in tubules | High | Protein (IHC) | HPA → |
| Skin | melanocytes | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Oral mucosa | squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
| Smooth muscle | smooth muscle cells | Not detected | Protein (IHC) | HPA → |
| Spleen | cells in red pulp | Not detected | Protein (IHC) | HPA → |
| Adipose tissue | adipocytes | Low | Protein (IHC) | HPA → |
| Cerebellum | cells in granular layer | Low | Protein (IHC) | HPA → |
Section 3
Advanced FRY Western Blot Tips
Deeper troubleshooting and optimisation questions for FRY, answered from its protein features.
How should FRY 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.
Do listed FRY isoforms explain multiple bands?
Isoforms · The supplied UniProt record lists one isoform and no alternative sequence. It therefore provides no isoform-based explanation for multiple bands. Assess additional bands separately rather than assigning them to named FRY isoforms.
Which FRY phosphorylation sites matter when interpreting bands?
PTM · In UniProt numbering, the listed sites are phosphotyrosine 213; phosphoserines 1382, 1383, 1936, 1940, 2419, 2420, 2487, and 2808; and phosphothreonine 2508, modified by CDK1. Compare samples under matched conditions before attributing a band change to phosphorylation.
Yes. CDK1 phosphorylation promotes FRY interaction with PLK1 in mitotic cells, followed by further PLK1 phosphorylation; that interaction is absent in interphase cells. Record cell-cycle conditions when comparing phospho-sensitive signals. These features do not establish that total FRY abundance changes.
Yes. CDK1 phosphorylation promotes FRY interaction with PLK1 in mitotic cells, followed by further PLK1 phosphorylation; that interaction is absent in interphase cells. Record cell-cycle conditions when comparing phospho-sensitive signals. These features do not establish that total FRY abundance changes.
Does this guide establish induction of FRY?
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 FRY Western blot?
Transfer · FRY is large, at approximately 338.9 kDa. Check whether the high-mass region transfers to the membrane and whether FRY remains in the gel before interpreting a weak signal. The supplied features do not specify a transfer method or settings.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A01003 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 FRY be quantified across cell-cycle conditions?
Quantitation · Compare like samples and account for localization: FRY is diffuse in the cytoplasm during interphase and is also listed at centrosomes and spindle poles. Its CDK1-dependent interaction with PLK1 occurs in mitotic cells. A change in a subcellular fraction may reflect redistribution, so distinguish fraction signals from total-protein measurements.
Should FRY migrate above its predicted mass?
Interpretation · FRY is 3,013 amino acids with a predicted mass of 338.9 kDa; the reported band near 339 kDa is consistent with that value. Its listed phosphorylation sites alone do not establish a visible shift or explain any mass difference.
Use the approximately 339 kDa observed band and 338.9 kDa predicted mass as reference points. The record lists phosphorylation but does not show that it produces a resolvable shift. It lists one isoform and no signal peptide, propeptide, or glycosylation sites, so these features do not explain an additional band.
Use the approximately 339 kDa observed band and 338.9 kDa predicted mass as reference points. The record lists phosphorylation but does not show that it produces a resolvable shift. It lists one isoform and no signal peptide, propeptide, or glycosylation sites, so these features do not explain an additional band.
Boster reagents
FRY 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-FRY Antibody Picoband®
Cat # A01003
The catalog reports one anti-FRY antibody, A01003, with reported human, mouse, and rat reactivity. Its WB image shows a band near the expected 339 kDa in the listed lysates. No publication evidence is supplied.
Which to pick: A01003 is the only listed option. Its WB image includes human RT4 and Caco-2 cells and rat and mouse brain and lung tissue; use those tested samples to judge its fit for your experiment.
Source: BosterBio FRY gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q5TBA9.
- Human Protein Atlas. FRY tissue expression.
- PMC3759375 — target-verified WB comparison