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- Table of Contents
IFIT3 / Interferon-induced protein with tetratricopeptide repeats 3 · Western blot design guide
Design a Western Blot for IFIT3
Real validated IFIT3 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-IFIT3 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
IFIT3 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 | ~56 kDa | |
| Gel | 12–15% (standard starting point) | |
| Positive control | Colon (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 | 1 isoform(s) |
Section 1
Real Curated IFIT3 Western Blot Protocols
The A03920 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | HepG2 cell lysate (catalog A03920) |
| 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 | A03920 · 1 μg/ml (catalog A03920) |
| 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 IFIT3 Western Blot Band Size?
IFIT3 is predicted at 56 kDa; phosphorylation could affect mobility, but no empirical band or demonstrated migration shift is supplied.
| Band near 56 kDa | Compatible with the predicted size of IFIT3; confirm identity with a control |
| Single sharp band near 56 kDa | Consistent with the one annotated IFIT3 isoform |
| Band with slightly altered mobility | Could reflect phosphorylation, but the annotated sites do not establish a visible shift |
| Band in a cytoplasmic or mitochondrial fraction | Consistent with the annotated locations of IFIT3 |
💡Expected IFIT3 appearanceIFIT3 has a predicted mass of 56 kDa and no supplied empirical band size; a band near 56 kDa is plausible, but confirm its identity with appropriate controls.
| Predicted molecular mass | Places the expected unmodified band near 56 kDa |
| Phosphoserine 203 | May affect mobility; no visible shift is established |
| Phosphoserine 237 | May affect mobility; no visible shift is established |
| Phosphoserine 478 | May affect mobility; no visible shift is established |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | IFIT3 expression may be low in the tested cells | Check an interferon-stimulated positive control and loading |
| Band higher than expected | Identity or modification state is unresolved | Compare stimulated and unstimulated samples and confirm with an independent antibody |
| Band lower than expected | Identity or sample degradation is unresolved | Check sample handling and confirm with an independent antibody |
| Multiple bands | Phosphorylation is annotated, but distinct bands are not established | Use a phosphatase-treated aliquot and an independent antibody to assess identity |
| Weak or no signal | IFIT3 expression may be low without interferon stimulation | Include a stimulated positive control and verify antibody performance |
Experiment evidence
Western blot images showing IFIT3 band
A03920 · IFIT3
A03920-1 · IFIT3
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for IFIT3 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for IFIT3 in Western blot, you can use colon tissue, which HPA scores High.
Positive control: Colon (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: IFIT3 is intracellular, so tissue lysates are suitable; validate the HPA Not-detected adipose tissue as a negative control.
HPA tissue expression evidence for IFIT3
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 |
|---|---|---|---|---|
| Colon | endothelial cells | High | Protein (IHC) | HPA → |
| Epididymis | glandular cells | High | Protein (IHC) | HPA → |
| Kidney | cells in tubules | High | Protein (IHC) | HPA → |
| Seminal vesicle | glandular cells | High | Protein (IHC) | HPA → |
| Skin | melanocytes | 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 → |
| Cerebellum | cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Esophagus | squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
| Hippocampus | glial cells | Not detected | Protein (IHC) | HPA → |
| Oral mucosa | squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced IFIT3 Western Blot Tips
Deeper troubleshooting and optimisation questions for IFIT3, answered from its protein features.
How should IFIT3 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 IFIT3 isoforms explain multiple bands?
Isoforms · Only one isoform is listed, with no alternative sequence. The supplied features therefore do not support assigning additional bands to IFIT3 isoforms.
Which phosphorylation sites should I consider when interpreting IFIT3 bands?
PTM · UniProt lists phosphoserines at positions 203, 237 and 478. These are UniProt coordinates; paper or antibody numbering may differ. Their presence alone does not demonstrate a visible shift or explain a mass discrepancy.
Does this guide establish induction of IFIT3?
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 IFIT3?
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 A03920 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 IFIT3 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.
Should IFIT3 migrate at exactly 56 kDa?
Interpretation · The predicted mass is 56 kDa, but no observed band position is supplied. Use 56 kDa as a reference, not an exact migration requirement; the listed features do not establish a visible shift.
IFIT3 is identified as interferon-induced and as a component of an interferon-dependent complex. Compare exposed and matched untreated samples, but determine the sampling time experimentally; the supplied features specify no induction kinetics.
IFIT3 interacts with IFIT1 and IFIT2 in an interferon-dependent multiprotein complex. That interaction alone does not establish that a higher Western blot band is the complex; verify the band before assigning it.
IFIT3 is listed in the cytoplasm and mitochondrion. Consider both locations when interpreting fractionated samples, and compare equivalent fractions when measuring changes in IFIT3 abundance.
IFIT3 is identified as interferon-induced and as a component of an interferon-dependent complex. Compare exposed and matched untreated samples, but determine the sampling time experimentally; the supplied features specify no induction kinetics.
IFIT3 interacts with IFIT1 and IFIT2 in an interferon-dependent multiprotein complex. That interaction alone does not establish that a higher Western blot band is the complex; verify the band before assigning it.
IFIT3 is listed in the cytoplasm and mitochondrion. Consider both locations when interpreting fractionated samples, and compare equivalent fractions when measuring changes in IFIT3 abundance.
Boster reagents
IFIT3 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-IFIT3 Antibody
Cat # A03920
Real WB data
Anti-IFIT3 Antibody
Cat # A03920-1
Both listed anti-IFIT3 antibodies have Western blot images. A03920 was shown with HepG2 lysate at 1 μg/ml; A03920-1 was shown with extracts from unspecified cell lines. The supplied evidence does not establish performance across all listed species.
Which to pick: For human HepG2 lysate, A03920 has the more specific reported Western blot example. A03920-1 lists Human, Mouse, and Rat reactivity, but its image caption does not identify the cell lines or species tested.
Source: BosterBio IFIT3 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry O14879.
- Human Protein Atlas. IFIT3 tissue expression.
- PMC7330856 — target-verified WB comparison
- PMC8561380 — target-verified WB comparison
- PMC11415598 — target-verified WB comparison