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- Table of Contents
AFF4 · Western blot design guide
AFF4 Western Blot Planning Guide
Plan a AFF4 Western blot around the catalog-observed 127.5 kDa band, image-backed M03824 evidence, HPA controls, and verified protocol records.
Evidence assembled July 2026 · For research use; verify linked source records and product datasheet before use
AFF4 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 | 127.5 kDa observed | |
| Observed band | 127.5 kDa observed | |
| Gel | 8-10% | |
| Positive control | Cerebellum | |
| Negative control | Ovary |
Important caveats
Reasons your observed band may differ from the expected size.
| ⓘ | Calculated mass | 127.5 kDa |
| ⓘ | Localization | Nucleus / Chromosome |
| ⓘ | Processing / PTM | Record-dependent |
| ⓘ | Reactivity | Human |
Section 1
Real Curated AFF4 Western Blot Protocols
Start with the molecular-weight rule, then compare verified publication-derived conditions.
| Sample / lysate | Caudate |
| Gel % | 8-10% |
| Load | 20-30 µg total protein per lane |
| Transfer | Wet/tank, extended transfer |
| Membrane | 0.45 µm PVDF |
| Blocking | 5% non-fat milk or 5% BSA in TBST |
| Primary | M03824 at datasheet starting dilution |
| Primary incubation | Overnight at 4 °C with gentle agitation |
| Secondary | Species-matched HRP conjugate at validated dilution |
| Wash | 3 × 5 min in TBST |
| Detection | Chemiluminescent substrate |
| Exposure | Bracket exposures to avoid saturation |
Section 2
What Is the Expected AFF4 Western Blot Band Size?
Use the product-observed 127.5 kDa band as the primary planning value and retain the UniProt calculated mass as context.
| 127.5 kDa | Matches the authoritative product WB observation. |
| 127.5 kDa calculated | Use as UniProt context, not as a replacement observed band. |
| Unexpected additional signal | Do not assign identity without orthogonal positive/negative controls. |
💡Expected AFF4 appearancePlan around 127.5 kDa and keep the calculated mass as supporting context.
| Catalog-observed band | 127.5 kDa; use this as the primary experimental expectation. |
| Calculated mass | 127.5 kDa from UniProt Q9UHB7; retain as context. |
| Gel selection | 8-10%; shared with the recommended protocol and poster. |
| Specificity check | Compare the lead HPA positive and negative controls with M03824. |
| Situation | Likely cause | Next action |
|---|---|---|
| 127.5 kDa | Matches the authoritative product WB observation. | Confirm with orthogonal controls and the linked product record. |
| Additional band | May reflect processing, modification, or non-specific signal. | Run a dilution series and compare positive/negative controls. |
| Weak signal | Target abundance or transfer may be limiting. | Verify transfer, increase positive-control abundance, and bracket exposure. |
Experiment evidence
Western blot images showing AFF4 band
M03824 · AFF4Observed 127.5 kDa · product-specific WB image
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for AFF4 Western blot
🧪Use Cerebellum as the first positive-control candidate and retain an orthogonal negative control.
Positive control: Cerebellum (High)
Negative control: Ovary (Not detected)
HPA protein score determines control status; other expression data is supporting context only.
HPA tissue expression evidence for AFF4
Comprehensive Human Protein Atlas IHC scoring per tissue. Rows are taken directly from the HPA tissue chart on each tissue page — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Cerebellum | Reported tissue cells | High | Protein (HPA) | HPA → |
| Testis | Reported tissue cells | High | Protein (HPA) | HPA → |
| Caudate | Reported tissue cells | High | Protein (HPA) | HPA → |
| Hippocampus | Reported tissue cells | High | Protein (HPA) | HPA → |
| Appendix | Reported tissue cells | Low | Protein (HPA) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Ovary | Reported tissue cells | Not detected | Protein (HPA) | HPA → |
Section 3
Advanced AFF4 Western Blot Tips
Deeper troubleshooting and optimisation questions for AFF4, answered from its protein features.
Which band should guide the blot?
Use 127.5 kDa, the observation attached to the authoritative M03824 WB record.
How should calculated mass be interpreted?
Treat the UniProt calculated mass as context; it does not replace the catalog-observed 127.5 kDa expectation.
Which positive control should I start with?
Start with Cerebellum, the lead HPA protein-expression candidate.
Which negative control is defensible?
Use Ovary as an orthogonal HPA Not detected candidate.
Which gel should I use?
Use 8-10% consistently across the quick facts, protocol table, and poster.
What transfer method to use for AFF4 Western blot?
Use the transfer method in the recommended protocol and verify transfer before blocking.
How should M03824 be started?
Start at the linked datasheet condition and run a three-point primary-antibody dilution test.
Which publication-derived protocols can I compare?
No verified publication protocol was supplied; use only the deterministic recommended protocol.
Boster reagents
AFF4 Western Blot Reagents
Human-reactive AFF4 Western blot reagents with authoritative product imagery.
Real WB data
Anti-AFF4 Picoband® Antibody (monoclonal, 8G12)
Cat # M03824
Only image-backed, WB-validated Human/Mouse/Rat recommendations from the prepared catalog evidence are shown.
Source: prepared picoband-wb product evidence; each card retains its own SKU, URL, observed band, and authoritative WB image.