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- Table of Contents
AMOTL2 / Angiomotin-like protein 2 · Western blot design guide
Design a Western Blot for AMOTL2
Source-linked AMOTL2 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-AMOTL2 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
AMOTL2 Western Blot Experimental Design Guide
Expected bands, source-linked protocol options, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before running
| Expected band | ~85.8 kDa | |
| Observed band | ~130–150 kDa | |
| Gel | 10% (catalog A06852) | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Ubl conjugation | |
| Caveat | Band identity controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 4 isoform(s) |
Section 1
Source-Linked AMOTL2 Western Blot Protocol Options
The A06852 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human 293T, human HepG2 (catalog A06852) |
| Gel % | 10% (catalog A06852) |
| Load | 30 ug; reducing conditions (catalog A06852) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A06852) |
| Membrane | nitrocellulose membrane (catalog A06852) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A06852) |
| Primary antibody | A06852 · 0.5 μg/mL (catalog A06852) |
| Primary incubation | overnight at 4°C (catalog A06852) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A06852) |
| Secondary incubation | 1.5 hour at RT (catalog A06852) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A06852) |
| Detection | ECL (catalog A06852) |
Section 2
What Is the Expected AMOTL2 Western Blot Band Size?
AMOTL2 is predicted at 85.8 kDa but observed at ~130–150 kDa in reducing lysates; the cause of this difference is not established.
| Band at ~130–150 kDa | Empirical AMOTL2 band in reducing whole-cell lysate; confirm identity with controls |
| Band near 85.8 kDa | Near the predicted mass, but identity requires confirmation |
| Several bands | AMOTL2 has four isoforms, though distinct migration is unestablished |
| Lower band alongside ~130–150 kDa | A shorter isoform is possible; verify band identity |
💡Expected AMOTL2 appearanceUniProt predicts 85.8 kDa, while antibody QC detects ~130–150 kDa in reducing whole-cell lysate; the cause of the difference is unestablished, so confirm band identity with appropriate controls.
| Predicted 85.8 kDa mass | Antibody QC detects ~130–150 kDa; the migration difference is unexplained |
| Isoform 1 | Its individual apparent size is not supplied |
| Isoform 2 | Its size relative to the other isoforms is not supplied |
| Isoform 3 | Its size relative to the other isoforms is not supplied |
| Isoform 4 | Its size relative to the other isoforms is not supplied |
| Situation | Likely cause | Next action |
|---|---|---|
| Band higher than expected | The observed ~130–150 kDa band exceeds the 85.8 kDa prediction for an unestablished reason | Compare with a validated positive lysate and confirm identity by AMOTL2 depletion |
| Band lower than expected | An alternative isoform is possible, but its migration is unknown | Check antibody epitope coverage and confirm the band by AMOTL2 depletion |
| Multiple bands | Four annotated isoforms may contribute, but separate bands are unproven | Compare isoform expression and test band identity by AMOTL2 depletion |
| No band in lysate | Intracellular AMOTL2 may be absent or poorly recovered from the sampled cells | Use a validated positive lysate and check extraction of junctional and cytoplasmic proteins |
| Weak or no signal | AMOTL2 abundance or antibody detection may be insufficient | Check loading and transfer against a validated positive lysate |
| Fragments below expected size | Sample degradation may produce smaller reactive bands | Prepare fresh lysate with protease inhibitors and confirm specificity by AMOTL2 depletion |
Experiment evidence
Western blot images showing AMOTL2 band
A06852 · AMOTL2
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for AMOTL2 Western blot
🧪For positive controls for AMOTL2 in Western blot, you can use a validated AMOTL2-positive lysate; the supplied HPA evidence identifies no specific tissue or cell line.
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 the samples.
⚠️Feasibility: HPA provides no expression data, so a validated positive sample and a naturally negative tissue cannot be selected.
HPA tissue expression evidence for AMOTL2
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 AMOTL2 Western Blot Tips
Deeper troubleshooting and optimisation questions for AMOTL2, answered from its protein features.
What should be checked when multiple AMOTL2 bands appear?
Band shift · Consider the four listed isoforms and phosphorylation at UniProt Tyr107, Ser759 and Ser762, then check which isoforms the antibody can recognize. The listed features identify possibilities; they do not assign any observed band to a specific isoform or modification.
Could AMOTL2 isoforms produce different bands?
Isoforms · UniProt lists four isoforms. Relative to the canonical sequence, isoform 3 lacks residues 525–526, isoform 2 changes residue 701 from T to TA, and isoform 4 has an N-terminal sequence replacement. Check whether the antibody recognizes each isoform before comparing bands.
Which AMOTL2 phosphorylation sites matter when interpreting bands?
PTM · UniProt lists Tyr107 phosphorylation by FGFR1 and phosphorylation at Ser759 and Ser762. These are UniProt coordinates, which may differ from antibody or paper numbering. Site presence alone does not establish a visible mobility shift.
FGFR1 is listed as the enzyme phosphorylating AMOTL2 Tyr107. If comparing conditions with different FGFR1 activity, assess Tyr107 phosphorylation directly when possible. The feature does not establish that a particular treatment induces AMOTL2 expression or shifts its band.
FGFR1 is listed as the enzyme phosphorylating AMOTL2 Tyr107. If comparing conditions with different FGFR1 activity, assess Tyr107 phosphorylation directly when possible. The feature does not establish that a particular treatment induces AMOTL2 expression or shifts its band.
Does this guide establish induction of AMOTL2?
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 AMOTL2?
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 A06852 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 AMOTL2 bands be quantified across samples?
Quantitation · Define which band is being measured and use the same definition across samples. AMOTL2 has four listed isoforms and occurs in the cytoplasm, recycling endosome, cell junction and podosome; sample preparation and antibody recognition may affect which forms are measured.
Does the 130–150 kDa band match AMOTL2’s predicted mass?
Interpretation · AMOTL2’s predicted mass is 85.8 kDa, below the supplied apparent band of about 130–150 kDa. Its listed phosphorylation sites and isoforms do not, by themselves, explain that difference. Confirm the band’s identity before assigning it to AMOTL2.
AMOTL2 is described as part of a complex with MAGI1 and CDH5, acting as a scaffold. That annotation does not show that the complex survives Western-blot preparation or causes the supplied 130–150 kDa band. Verify the band’s identity independently.
AMOTL2 is described as part of a complex with MAGI1 and CDH5, acting as a scaffold. That annotation does not show that the complex survives Western-blot preparation or causes the supplied 130–150 kDa band. Verify the band’s identity independently.
Boster reagents
AMOTL2 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-AMOTL2 Antibody Picoband®
Cat # A06852
The catalog reports A06852 for AMOTL2 Western blotting, with stated Human, Mouse, and Rat reactivity. Its WB image uses human 293T and HepG2 lysates; the reported band is approximately 130–150 kDa, above the expected 86 kDa.
Which to pick: A06852 is the only listed antibody. Its WB image documents human cell lysates under the reported conditions; Mouse and Rat reactivity is listed, but those species are not shown in the supplied WB caption.
Source: BosterBio AMOTL2 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q9Y2J4.
- Human Protein Atlas. AMOTL2 tissue expression.
- PMC4501364 — target-verified WB comparison
- PMC5572699 — target-verified WB comparison
- PMC11358041 — target-verified WB comparison