FOXN1 / Forkhead box protein N1 · Western blot design guide

Design a Western Blot for FOXN1

Real validated FOXN1 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-FOXN1 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

Western blot protocol sheet for FOXN1: expected band ~68.9 kDa, hero antibody A03369-2, catalog values and labelled standard workflow; separate PMC comparisons on the guide — Printable **FOXN1** Western blot protocol sheet — expected band ~68.9 kDa, antibody A03369-2, controls and PMC citations. Open the full FOXN1 WB guide →

Navigate to: Protocols Band Controls Antibodies

FOXN1 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	~68.9 kDa
	Observed band	~69 kDa
	Gel	5–20% (catalog A03369-2)
	Negative control ⓘ	Suggested KO / knockdown lysate

Important caveats

Reasons your observed band may differ from the expected size.

	PTM	—
	Caveat	—
	Gene-set association	MSigDB C7 membership
	Isoform	1 isoform(s)

Section 1

Real Curated FOXN1 Western Blot Protocols

The A03369-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.

Recommended Western blot protocol parameters
Sample / lysate	human Hela, human Raji, mouse NIH/3T3 (catalog A03369-2)
Gel %	5–20% (catalog A03369-2)
Load	30ug; reducing conditions (catalog A03369-2)
Transfer	a Nitrocellulose membrane at 150mA for 50-90 minutes (catalog A03369-2)
Membrane	Nitrocellulose membrane (catalog A03369-2)
Blocking	5% Non-fat Milk/ TBS for 1.5 hour at RT (catalog A03369-2)
Primary antibody	A03369-2 · 0.5 μg/mL (catalog A03369-2)
Primary incubation	overnight at 4°C (catalog A03369-2)
Secondary antibody	goat anti-rabbit IgG-HRP, 1:5000 (catalog A03369-2)
Secondary incubation	1.5 hour at RT (catalog A03369-2)
Wash	TBS-0.1%Tween 3 times with 5 minutes each (catalog A03369-2)
Detection	ECL (catalog A03369-2)

Section 2

What Is the Expected FOXN1 Western Blot Band Size?

FOXN1 is predicted at 68.9 kDa and observed at ~69 kDa; confirm band identity with appropriate controls.

What am I looking at on my blot?
Band at ~69 kDa	Matches the empirical FOXN1 band and its 68.9 kDa predicted mass
Single sharp band near 69 kDa	Consistent with the one listed FOXN1 isoform; confirm band identity
Faint band near 69 kDa in whole-cell lysate	Nuclear FOXN1 may be underrepresented in the sample
No band near 69 kDa in whole-cell lysate	Nuclear FOXN1 may be absent or below detection in the sample

💡Expected FOXN1 appearanceFOXN1 has a predicted mass of 68.9 kDa, and antibody QC shows a band at ~69 kDa; confirm its identity with appropriate controls.

How each factor affects band size
UniProt predicted mass of 68.9 kDa	Places full-length FOXN1 near 69 kDa
Predicted mass of 68,925 Da	Corresponds to approximately 68.9 kDa
Full-length FOXN1 predicted mass	Closely matches the empirical ~69 kDa band
Predicted mass for the listed FOXN1 sequence	Provides no evidence for a second band size

Why is my band missing or off?
Situation	Likely cause	Next action
No band in lysate	Nuclear FOXN1 may be underrepresented	Check nuclear extraction and include a FOXN1-positive control
Band higher than expected	The supplied features do not establish a larger FOXN1 form	Compare with the ~69 kDa positive-control band and verify antibody specificity
Band lower than expected	The supplied features do not establish a smaller FOXN1 form	Check sample integrity and compare with a positive control
Multiple bands	Only one isoform is listed; additional band identities are uncertain	Use positive and negative controls to identify the ~69 kDa band
Weak or no signal	The sample may contain little nuclear FOXN1	Assess nuclear enrichment and include a positive control

Source: UniProt O15353 Human Protein Atlas product datasheet (A03369-2)

Experiment evidence

Western blot images showing FOXN1 band

A03369-2 · FOXN1

← swipe / scroll for more images →

Real WB data behind the band-size expectations in this section.

Sample controls for FOXN1 Western blot

🧪For positive controls for FOXN1 in Western blot, you can use no HPA-supported positive sample because none is provided.

Positive control: No high/medium HPA tissue identified

Negative control: Suggested KO / knockdown lysate

Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.

⚠️Feasibility: HPA provides no expression data, so tissue controls cannot be validated; the nucleus is the reported location.

HPA tissue expression evidence for FOXN1

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
No high/medium HPA tissues identified in the supplied evidence.

Lower expression tissues · IHC evidence, not confirmed WB-negative controls

Tissue	Cell type	Level	Evidence	Source
No lower-expression tissue rows available in the supplied evidence.

Section 3

Advanced FOXN1 Western Blot Tips

Deeper troubleshooting and optimisation questions for FOXN1, answered from its protein features.

How should FOXN1 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 FOXN1 isoforms explain multiple bands?

Isoforms · The supplied UniProt data lists one isoform and no alternative sequence. Do not assign additional bands to FOXN1 isoforms on this evidence alone.

Do listed modifications explain a FOXN1 band shift?

PTM · No modified residues or glycosylation sites are listed. This does not rule out unlisted modifications, and the supplied features do not establish a visible shift.

Does this guide establish induction of FOXN1?

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.

What transfer method to use for FOXN1 Western blot?

Transfer · FOXN1 is predicted at 68.9 kDa. The supplied features do not favor a particular transfer method; verify that protein around 69 kDa reaches the membrane using a molecular weight marker and a transfer check.

How should blocking be optimized?

Blocking · Standard workflow guidance: follow the A03369-2 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 FOXN1 abundance be quantified?

Quantitation · FOXN1 is annotated as nuclear. For comparisons, use the same sample fraction across lanes and quantify the approximately 69 kDa band against a suitable loading measure for that fraction.

Does FOXN1 show a shift from its predicted molecular mass?

Interpretation · FOXN1 is predicted at 68.9 kDa, and the supplied observed band is approximately 69 kDa. These values agree within the stated precision; no shift is established.

Compare them with the approximately 69 kDa observed band. The supplied record lists one isoform and no modified residues or glycosylation sites, so it does not identify the source of additional bands. Check band specificity before assigning them to FOXN1.

Boster reagents

FOXN1 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

Western blot analysis of FOXN1 using anti-FOXN1 antibody (A03369-2). Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30ug of sample under reducing conditions. Lane 1: human Hela whole cell lysates, Lane 2: human Raji whole cell lysates, Lane 3: mouse NIH/3T3 whole cell lysates, Lane 4: mouse RAW264.7 whole cell lysates. After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/ TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-FOXN1 antigen affinity purified polyclonal antibody (Catalog # A03369-2) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for FOXN1 at approximately 69KD. The expected band size for FOXN1 is at 69KD.

Anti-FOXN1 Antibody Picoband®

Cat # A03369-2

View product →

The catalog reports one anti-FOXN1 antibody for Western blot, A03369-2, with reported human, mouse, and rat reactivity. Its WB image shows an approximately 69 kDa band in human HeLa and Raji and mouse NIH/3T3 and RAW264.7 lysates; no rat sample is shown.

Which to pick: A03369-2 is the only listed option. Its WB image documents human and mouse cell lysates at 0.5 μg/mL antibody; rat reactivity is listed, but the supplied image does not test a rat sample.

Source: BosterBio FOXN1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.

References

UniProt Consortium. UniProt entry O15353.
Human Protein Atlas. FOXN1 tissue expression.
PMC11389453 — target-verified WB comparison
PMC11272594 — target-verified WB comparison
PMC9934747 — target-verified WB comparison