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PTK2B / Protein-tyrosine kinase 2-beta · Western blot design guide
Design a Western Blot for PTK2B
Real validated PTK2B Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-PTK2B 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
PTK2B 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 | ~115.9 kDa | |
| Observed band | ~116 kDa | |
| Gel | 8% (catalog A01066-1) | |
| Positive control | Fallopian tube (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 | Phosphorylated | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 2 isoform(s) |
Section 1
Real Curated PTK2B Western Blot Protocols
The A01066-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human A549, rat brain, mouse brain (catalog A01066-1) |
| Gel % | 8% (catalog A01066-1) |
| Load | 30 ug; reducing conditions (catalog A01066-1) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A01066-1) |
| Membrane | nitrocellulose membrane (catalog A01066-1) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A01066-1) |
| Primary antibody | A01066-1 · 0.5 μg/mL (catalog A01066-1) |
| Primary incubation | overnight at 4°C (catalog A01066-1) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A01066-1) |
| Secondary incubation | 1.5 hour at RT (catalog A01066-1) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A01066-1) |
| Detection | ECL (catalog A01066-1) |
Section 2
What Is the Expected PTK2B Western Blot Band Size?
PTK2B is predicted at 115.9 kDa and observed at ~116 kDa; isoforms and homodimers are possible sources of additional bands, but their migration is unconfirmed.
| Band at ~116 kDa | Matches the observed PTK2B band and its 115.9 kDa predicted mass. |
| Single band near ~116 kDa | Isoforms 1 and 2 may not resolve into separate bands. |
| Additional band at a different size | Could reflect isoform 1 or 2 if their migration differs; confirm its identity. |
| Band near ~232 kDa | Could reflect a homodimer if it remains intact during electrophoresis; confirm its identity. |
💡Expected PTK2B appearancePTK2B has a predicted mass of 115.9 kDa and an observed band at ~116 kDa in reducing whole-cell and tissue lysates; use appropriate band-identity controls for additional bands.
| Predicted PTK2B mass | 115.9 kDa, consistent with the observed ~116 kDa band. |
| Homodimer formation | Could yield a band near twice the monomer mass if the complex survives electrophoresis. |
| Isoform 1 | Its size relative to isoform 2 is unspecified; separate migration is unconfirmed. |
| Isoform 2 | Its size relative to isoform 1 is unspecified; separate migration is unconfirmed. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | PTK2B may be underrepresented in the sampled fraction because it also localizes to membranes and focal adhesions. | Check a positive-control lysate and compare soluble with membrane-associated fractions. |
| Band higher than expected | A PTK2B homodimer could persist during electrophoresis. | Compare denaturing conditions and confirm the band with a second PTK2B antibody or knockdown. |
| Band lower than expected | An alternative isoform is possible, but its mass is unspecified. | Confirm the band with a second PTK2B antibody or knockdown. |
| Multiple bands | Isoforms 1 and 2 could migrate differently, although separation is unconfirmed. | Compare isoform expression and confirm band identity with knockdown. |
| Weak or no signal | The sampled fraction may contain little PTK2B. | Check loading and a positive-control lysate; examine a membrane-associated fraction. |
| Fragments below expected size | Proteolysis during sample preparation is possible; no PTK2B cleavage product is specified. | Prepare fresh lysate with protease inhibitors and confirm fragment identity with another antibody. |
Experiment evidence
Western blot images showing PTK2B band
A01066-1 · PTK2B
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for PTK2B Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for PTK2B in Western blot, you can use fallopian tube tissue, where HPA reports high expression.
Positive control: Fallopian tube (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: HPA reports high expression in fallopian tube and no detection in adipose tissue, providing a feasible tissue control pair.
HPA tissue expression evidence for PTK2B
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 |
|---|---|---|---|---|
| Fallopian tube | ciliated cells (cell body) | High | Protein (IHC) | HPA → |
| Nasopharynx | basal cells | High | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | glandular cells | Medium | Protein (IHC) | HPA → |
| Breast | 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 → |
| Bone marrow | hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Liver | cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Oral mucosa | squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
| Ovary | follicle cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced PTK2B Western Blot Tips
Deeper troubleshooting and optimisation questions for PTK2B, answered from its protein features.
How should PTK2B 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 PTK2B isoforms produce different bands?
Isoforms · UniProt lists isoforms 1 and 2. Isoform 2 lacks residues 739–780 of the canonical sequence, so it could migrate differently. The feature does not establish where either isoform will appear on a blot.
No. Isoform 2 lacks canonical residues 739–780, which include phosphoserine 762 and phosphothreonine 765. An antibody directed at either canonical site would not measure that site in isoform 2. Check the antibody’s numbering convention and target sequence.
No. Isoform 2 lacks canonical residues 739–780, which include phosphoserine 762 and phosphothreonine 765. An antibody directed at either canonical site would not measure that site in isoform 2. Check the antibody’s numbering convention and target sequence.
Which phosphorylation sites are useful for interpreting PTK2B activation?
PTM · UniProt lists phosphotyrosine 402 by autocatalysis, phosphotyrosine 580 by SRC, FYN and LCK, and phosphotyrosine 881 by SRC. Choose a site-specific antibody according to the event being measured, and compare its signal with total PTK2B. These are UniProt canonical coordinates; antibody numbering may differ.
Measure the selected phospho-site signal alongside total PTK2B in matched samples. Report which site was measured: the listed phosphoserine, phosphothreonine and phosphotyrosine sites represent distinct modifications. A change at one site does not establish a change at every site.
Measure the selected phospho-site signal alongside total PTK2B in matched samples. Report which site was measured: the listed phosphoserine, phosphothreonine and phosphotyrosine sites represent distinct modifications. A change at one site does not establish a change at every site.
Does this guide establish induction of PTK2B?
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 PTK2B?
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 A01066-1 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.
How should PTK2B be quantified?
Quantitation · Standard workflow guidance: quantify only a validated target band within the linear exposure range. Use consistent sample preparation and loading, retain biological replicates, and avoid interpreting saturation or loading differences as regulation.
Does the observed PTK2B band indicate a mass shift?
Interpretation · The predicted mass is 115.9 kDa, and the reported apparent band is about 116 kDa. These values agree within rounding. PTK2B has listed phosphorylation sites, but their presence alone does not establish a visible shift.
Yes. UniProt places PTK2B in the cytoplasm, at the cytoplasmic side of the cell membrane, at focal adhesions and in the nucleus. Its interaction with NPHP1 induces membrane association. Compare like fractions when interpreting changes in PTK2B abundance.
Check whether a lower band is consistent with isoform 2, which lacks canonical residues 739–780, and whether the antibody recognizes both isoforms. UniProt also describes homodimers or homooligomers, but these features alone do not identify an unexpected band or establish a visible mass shift.
Yes. UniProt places PTK2B in the cytoplasm, at the cytoplasmic side of the cell membrane, at focal adhesions and in the nucleus. Its interaction with NPHP1 induces membrane association. Compare like fractions when interpreting changes in PTK2B abundance.
Check whether a lower band is consistent with isoform 2, which lacks canonical residues 739–780, and whether the antibody recognizes both isoforms. UniProt also describes homodimers or homooligomers, but these features alone do not identify an unexpected band or establish a visible mass shift.
Boster reagents
PTK2B 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-PYK2/PTK2B Antibody Picoband®
Cat # A01066-1
The catalog reports one anti-PTK2B antibody, A01066-1. Its Western blot image shows an approximately 116 kDa band in human A549 cells and rat and mouse brain lysates. Evidence is limited to the supplied product image and caption.
Which to pick: A01066-1 is the only listed option. It reports human, mouse, and rat reactivity and has a Western blot image for the specified lysates, using 30 µg per lane and 0.5 µg/mL primary antibody. Match your sample and conditions to those shown.
Source: BosterBio PTK2B gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q14289.
- Human Protein Atlas. PTK2B tissue expression.
- PMC4998215 — target-verified WB comparison
- PMC4133292 — target-verified WB comparison
- PMC9535427 — target-verified WB comparison