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- Table of Contents
IFITM3 / Interferon-induced transmembrane protein 3 · Western blot design guide
Design a Western Blot for IFITM3
Real validated IFITM3 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-IFITM3 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
IFITM3 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 | ~14.6 kDa | |
| Gel | 15% (standard starting point) | |
| Positive control | Bone marrow (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 + Ubl conjugation | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 1 isoform(s) |
Section 1
Real Curated IFITM3 Western Blot Protocols
The M02265 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | HeLa cell lysate (catalog M02265) |
| Gel % | 15% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Short semi-dry transfer; verify retention (standard starting point) |
| Membrane | 0.2 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | M02265; use the WB datasheet starting dilution (standard starting point) |
| 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 IFITM3 Western Blot Band Size?
IFITM3 has a predicted mass of 14.6 kDa; annotated phosphorylation and lipid modification have no demonstrated migration effect here, and no empirical band size is supplied.
| Band near 14.6 kDa | Consistent with the predicted IFITM3 sequence mass; confirm identity with antibody controls |
| Weak band in whole-cell lysate | Membrane-associated IFITM3 may be poorly recovered during extraction |
| Band enriched in a membrane fraction | Consistent with IFITM3 localization at cell and endosome membranes |
| Band at a different apparent size | The supplied features do not establish a specific migration shift; verify band identity |
💡Expected IFITM3 appearanceIFITM3 has a predicted sequence mass of 14.6 kDa, but no empirical band size is supplied; confirm any candidate band with ordinary antibody specificity controls.
| Predicted sequence mass | Sets a 14.6 kDa reference, not a validated apparent band size |
| Phosphotyrosine at Tyr20 | Is annotated, but a visible mobility shift is not established |
| Signal-anchor membrane localization | May affect recovery from lysate; no size change is established |
| Palmitate annotation | Indicates lipid modification, but its effect on apparent size is not established |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Membrane-associated IFITM3 may be poorly extracted | Check membrane recovery and compare with a membrane-enriched fraction |
| Band higher than expected | The supplied features do not establish a higher migrating species | Check antibody specificity with an IFITM3 depletion control |
| Band lower than expected | The supplied features do not establish a cleaved product | Check antibody specificity with an IFITM3 depletion control |
| Multiple bands | Only one isoform is listed; identities of additional bands are uncertain | Compare bands before and after IFITM3 depletion |
| Weak or no signal | Membrane-associated IFITM3 may be underrepresented after extraction | Check extraction efficiency with a membrane protein control |
Experiment evidence
Western blot images showing IFITM3 band
M02265 · IFITM3
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for IFITM3 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for IFITM3 in Western blot, you can use bone marrow, which HPA reports as highly expressed.
Positive control: Bone marrow (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: As a membrane protein, IFITM3 may require effective membrane protein extraction for a clear signal.
HPA tissue expression evidence for IFITM3
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 |
|---|---|---|---|---|
| Bone marrow | hematopoietic cells | High | Protein (IHC) | HPA → |
| Endometrium | cells in endometrial stroma | High | Protein (IHC) | HPA → |
| Epididymis | glandular cells | High | Protein (IHC) | HPA → |
| Fallopian tube | glandular cells | High | Protein (IHC) | HPA → |
| Kidney | cells in glomeruli | 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 → |
| Appendix | glandular cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Duodenum | glandular cells | Not detected | Protein (IHC) | HPA → |
| Gallbladder | glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced IFITM3 Western Blot Tips
Deeper troubleshooting and optimisation questions for IFITM3, answered from its protein features.
How should IFITM3 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.
Does a second band indicate an IFITM3 isoform?
Isoforms · Only one isoform is listed, with no alternative sequence supplied. Do not assign a second band to an isoform from these features alone; check its specificity experimentally.
Which phosphorylation site matters when interpreting IFITM3 bands?
PTM · UniProt lists phosphotyrosine at position 20. Keep UniProt numbering explicit when comparing antibodies or papers, which may use another convention. This site does not by itself establish a visible band shift.
Can interferon treatment help assess IFITM3 induction?
Induction · IFITM3 is identified as interferon-induced. Compare treated and untreated samples under matched loading conditions to assess its response. These features do not specify an interferon type, dose, treatment time, or expected fold change.
What transfer method to use for IFITM3 Western blot?
Transfer · IFITM3 is a predicted 14.6-kDa membrane protein. Choose conditions that retain small proteins and verify transfer and retention. The supplied features do not establish a specific membrane, buffer, or transfer time.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the M02265 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 IFITM3 abundance be quantified?
Quantitation · Measure bands within the assay's linear range and normalize consistently. IFITM3 is listed at cell and late-endosome membranes, so use comparable sample fractions across conditions when interpreting changes in abundance.
Should IFITM3 migrate exactly at its predicted mass?
Interpretation · The predicted mass is 14.6 kDa, but no observed band position is supplied. Use 14.6 kDa as a starting reference, not a required migration position. The listed modifications alone do not establish a visible shift.
Check band specificity before assigning an identity. The supplied features list phosphotyrosine at UniProt position 20 and keywords for palmitate and ubiquitin-like conjugation, but give no corresponding mass shifts or observed band positions. No glycosylation sites or alternative sequence are listed.
Check band specificity before assigning an identity. The supplied features list phosphotyrosine at UniProt position 20 and keywords for palmitate and ubiquitin-like conjugation, but give no corresponding mass shifts or observed band positions. No glycosylation sites or alternative sequence are listed.
Boster reagents
IFITM3 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-Fragilis IFITM3 Monoclonal Antibody
Cat # M02265
M02265 is a human-reactive anti-IFITM3 monoclonal antibody with a Western blot image showing Fragilis expression in HeLa cell lysate. The supplied evidence does not establish performance in other samples or species.
Which to pick: M02265 is the only listed option. Its reported human reactivity and HeLa cell lysate Western blot image make it the documented choice for that context; other contexts are not shown.
Source: BosterBio IFITM3 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q01628.
- Human Protein Atlas. IFITM3 tissue expression.
- PMC6719045 — target-verified WB comparison
- PMC12478411 — target-verified WB comparison
- PMC13386968 — target-verified WB comparison