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- Table of Contents
WIPI2 / WD repeat domain phosphoinositide-interacting protein 2 · Western blot design guide
Design a Western Blot for WIPI2
Real validated WIPI2 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-WIPI2 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
WIPI2 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 | ~49.4 kDa | |
| Gel | 12–15% (standard starting point) | |
| Positive control | Testis (IHC candidate; verify WB) | |
| 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 C7 membership | |
| Isoform | 6 isoform(s) |
Section 1
Real Curated WIPI2 Western Blot Protocols
The A06492 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human testis tissue lysate (catalog A06492) |
| Gel % | 12–15% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Standard semi-dry transfer; verify efficiency (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | A06492 · 1 μg/mL (catalog A06492) |
| 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 WIPI2 Western Blot Band Size?
WIPI2 is predicted at 49.4 kDa; isoforms and Ser413 phosphorylation could affect migration, but no empirical band size or visible shift is established.
| Band near 49.4 kDa | Consistent with the predicted WIPI2 mass; confirm identity with antibody controls |
| Several discrete bands | Could reflect WIPI2 isoforms 1–6 if their migration differs; band identities need verification |
| Close doublet | Could reflect phosphorylation at Ser413 if it affects migration; this has not been demonstrated |
| Weak band in a soluble fraction | WIPI2 is a peripheral membrane protein and may partition into the membrane fraction |
💡Expected WIPI2 appearanceWIPI2 has a predicted mass of 49.4 kDa, but no empirical band size is supplied; confirm a candidate band with antibody specificity and fractionation controls.
| Predicted WIPI2 mass | Sets a 49.4 kDa reference, not a measured migration position |
| Splice isoform 1 | Its apparent size relative to other isoforms is not supplied |
| Splice isoforms 2 and 3 | Their apparent sizes may differ, but no relative sizes are supplied |
| Splice isoforms 4, 5, and 6 | Their apparent sizes may differ, but no relative sizes are supplied |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | WIPI2 may be poorly recovered during extraction of membrane-associated proteins | Check the membrane fraction and extraction efficiency |
| Band higher than expected | Isoform-dependent migration is possible but unverified | Compare isoform-specific controls and confirm antibody specificity |
| Band lower than expected | An alternatively spliced isoform is possible, but its size is unknown | Compare isoform-specific controls and confirm band identity |
| Multiple bands | WIPI2 has six annotated isoforms, though distinct bands are not established | Check isoform expression and use an independent WIPI2 antibody |
| Weak or no signal | Membrane association may reduce recovery in the tested fraction | Check membrane-containing fractions and use a positive control |
| Fragments below expected size | Sample degradation or nonspecific antibody binding is possible | Compare fresh lysate with a second WIPI2 antibody |
Experiment evidence
Western blot images showing WIPI2 band
A06492 · WIPI2
A06492-1 · WIPI2
A06492-1 · WIPI2
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for WIPI2 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for WIPI2 in Western blot, you can use testis lysate, which HPA scores high for WIPI2.
Positive control: Testis (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: A high-expression testis sample and an HPA not-detected adipose tissue sample are available, but membrane-associated WIPI2 requires adequate protein extraction.
HPA tissue expression evidence for WIPI2
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 |
|---|---|---|---|---|
| Testis | cells in seminiferous ducts | High | 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 → |
| Breast | adipocytes | 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 WIPI2 Western Blot Tips
Deeper troubleshooting and optimisation questions for WIPI2, answered from its protein features.
How should WIPI2 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.
Which WIPI2 isoforms could produce different bands?
Isoforms · UniProt lists six isoforms. Isoforms 2 and 4 lack residues 26–43; isoform 3 lacks residues 1–59 and replaces 60–71; isoform 5 replaces 1–71. Isoforms 2, 3, 5, and 6 lack 407–417. Check which isoforms your antibody recognizes before assigning bands.
Check the immunogen against the annotated variable regions. An antibody targeting residues 407–417 would miss isoforms 2, 3, 5, and 6; one targeting 26–43 would miss isoforms 2 and 4. Confirm epitope coverage before interpreting absent bands as absent protein.
Check the immunogen against the annotated variable regions. An antibody targeting residues 407–417 would miss isoforms 2, 3, 5, and 6; one targeting 26–43 would miss isoforms 2 and 4. Confirm epitope coverage before interpreting absent bands as absent protein.
Could phosphorylation affect the WIPI2 band?
PTM · UniProt annotates phosphoserine at position 413 in the canonical sequence. The 407–417 segment is absent from isoforms 2, 3, 5, and 6, so that site is absent from those isoforms. Match residue numbers to the UniProt canonical sequence when comparing antibody or paper annotations. This annotation alone does not establish a visible shift.
Does this guide establish induction of WIPI2?
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 WIPI2?
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 A06492 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 WIPI2 bands be quantified across samples?
Quantitation · WIPI2 is annotated as a peripheral protein of the preautophagosomal structure membrane. Keep fractionation and sample preparation consistent across samples, and quantify the same antibody-recognized band or isoform set each time. Changes in membrane association could affect the amount recovered in a given fraction.
Should WIPI2 run at its predicted 49.4 kDa?
Interpretation · 49.4 kDa is the predicted mass for canonical WIPI2. Six annotated isoforms and phosphoserine at UniProt position 413 warrant checking band identity, but these features alone do not establish an apparent band shift. No observed band position is supplied.
Compare each band's size and antibody epitope with the annotated isoform changes, especially the N-terminal variants and the 407–417 deletion. The supplied features do not establish observed band positions, so neither an isoform assignment nor a phosphorylation-dependent shift can be confirmed from size alone.
Compare each band's size and antibody epitope with the annotated isoform changes, especially the N-terminal variants and the 407–417 deletion. The supplied features do not establish observed band positions, so neither an isoform assignment nor a phosphorylation-dependent shift can be confirmed from size alone.
Boster reagents
WIPI2 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-WIPI2 Antibody
Cat # A06492
Real WB data
Anti-WIPI2 Antibody
Cat # A06492-1
Both listed anti-WIPI2 antibodies have Western blot images. A06492 was shown with human testis lysate at 1 μg/mL; A06492-1 was shown with extracts from various cell lines at 1:1000. The cell lines are unspecified, and these images do not establish broader validation.
Which to pick: For mouse samples, choose A06492-1, the only option listing mouse reactivity. Both list human and rat reactivity. For a human testis lysate precedent, consider A06492; for a cell-line blot precedent, consider A06492-1.
Source: BosterBio WIPI2 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q9Y4P8.
- Human Protein Atlas. WIPI2 tissue expression.
- PMC7107944 — target-verified WB comparison
- PMC11817878 — target-verified WB comparison
- PMC10189881 — target-verified WB comparison