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- Table of Contents
ZFX / Zinc finger X-chromosomal protein · Western blot design guide
Design a Western Blot for ZFX
Source-linked ZFX Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-ZFX 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
ZFX Western Blot Experimental Design Guide
Expected bands, source-linked protocol options, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before running
| Expected band | ~90.5 kDa | |
| Gel | 8–10% (standard starting point) | |
| Positive control | Fallopian tube (IHC candidate; verify WB) +4 more | |
| Negative control | Soft 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 | 3 isoform(s) |
Section 1
Source-Linked ZFX Western Blot Protocol Options
The A05947 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | Daudi cell lysate (catalog A05947) |
| Gel % | 8–10% (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 | A05947 · 1 μg/mL (catalog A05947) |
| 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 ZFX Western Blot Band Size?
ZFX is predicted at 90.5 kDa; isoforms and Ser274 phosphorylation are documented, but their effects on gel migration are not demonstrated.
| Band near 90.5 kDa | Consistent with the predicted ZFX mass; confirm identity with antibody controls. |
| Additional band at another position | Could reflect isoforms 1, 2, or 3; their migration is not established. |
| Band near 90.5 kDa without a visible shift | Could still contain ZFX phosphorylated at Ser274; that site does not establish a resolvable shift. |
| Weak or absent band in cytosolic fraction | Consistent with ZFX localization to the nucleus. |
💡Expected ZFX appearanceUniProt predicts ZFX at 90.5 kDa; no empirical band size or isoform migration is supplied, so confirm any candidate band with antibody and depletion controls.
| Predicted ZFX mass | Places the reference band near 90.5 kDa; migration is unverified. |
| Splice isoform 1 | May differ in size from isoforms 2 and 3; its mass is not supplied. |
| Splice isoform 2 | May differ in size from isoforms 1 and 3; its mass is not supplied. |
| Splice isoform 3 | May differ in size from isoforms 1 and 2; its mass is not supplied. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Nuclear ZFX may be poorly recovered from the lysate. | Check nuclear extraction and include a nuclear protein control. |
| Band higher than expected | The identity of a higher band is unestablished. | Check whether it decreases after ZFX depletion. |
| Band lower than expected | An isoform is possible, but its migration is unknown. | Compare with ZFX depletion and an independent antibody. |
| Multiple bands | ZFX has isoforms 1, 2, and 3; distinct bands are not established. | Test which bands decrease after ZFX depletion. |
| Weak or no signal | Nuclear ZFX may be underrepresented in the sample. | Enrich the nuclear fraction and check sample loading. |
Experiment evidence
Western blot images showing ZFX band
A05947 · ZFX
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for ZFX Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for ZFX in Western blot, you can use fallopian tube tissue, which shows high HPA expression.
Positive control: Fallopian tube (IHC candidate; verify WB)
Negative control: Soft tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: Because ZFX is nuclear, a nuclear extract may improve detection.
HPA tissue expression evidence for ZFX
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 |
|---|---|---|---|---|
| Fallopian tube | glandular cells | High | Protein (IHC) | HPA → |
| Gallbladder | glandular cells | High | Protein (IHC) | HPA → |
| Placenta | trophoblastic cells | High | Protein (IHC) | HPA → |
| Tonsil | non-germinal center cells | High | Protein (IHC) | HPA → |
| Appendix | lymphoid tissue | Medium | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Soft tissue | chondrocytes | Not detected | Protein (IHC) | HPA → |
| Adipose tissue | adipocytes | Low | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Low | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | Low | Protein (IHC) | HPA → |
| Caudate | glial cells | Low | Protein (IHC) | HPA → |
Section 3
Advanced ZFX Western Blot Tips
Deeper troubleshooting and optimisation questions for ZFX, answered from its protein features.
How should ZFX 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.
How could ZFX isoforms affect band interpretation?
Isoforms · UniProt lists three isoforms. Isoform 2 lacks residues 1–229 of the canonical sequence; isoform 3 replaces canonical residue 19 with a longer sequence. These changes may affect band positions, but the supplied features do not establish where any isoform migrates.
An antibody recognizing canonical residues 1–229 may miss isoform 2, which lacks that region. Check the antibody epitope against each isoform sequence before interpreting a missing or additional band.
Decide whether the measurement targets one isoform or combined ZFX signal. Isoform 2 lacks canonical residues 1–229, so an antibody against that region would not measure all three isoforms equally. Quantify the same defined band or bands across samples.
An antibody recognizing canonical residues 1–229 may miss isoform 2, which lacks that region. Check the antibody epitope against each isoform sequence before interpreting a missing or additional band.
Decide whether the measurement targets one isoform or combined ZFX signal. Isoform 2 lacks canonical residues 1–229, so an antibody against that region would not measure all three isoforms equally. Quantify the same defined band or bands across samples.
Does ZFX phosphorylation imply a visible band shift?
PTM · UniProt lists phosphoserine at position 274 in the canonical sequence. Feature presence alone does not demonstrate a visible shift. If comparing with an antibody or paper, check its residue-numbering convention before equating its site number with UniProt position 274.
Does this guide establish induction of ZFX?
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 ZFX?
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 A05947 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 ZFX 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 ZFX appear exactly at its predicted 90.5 kDa?
Interpretation · 90.5 kDa is the predicted mass, and no observed band position is supplied. The listed isoforms and phosphoserine do not establish an apparent mass or explain a difference from 90.5 kDa.
ZFX is annotated as nuclear. Include a nuclear-containing sample when assessing its detection, and keep sample preparation consistent across comparisons.
Compare candidate bands with the 90.5 kDa predicted mass and the documented isoform sequence changes. The supplied record has no observed band position, so band size alone cannot establish identity or assign a band to an isoform.
ZFX is annotated as nuclear. Include a nuclear-containing sample when assessing its detection, and keep sample preparation consistent across comparisons.
Compare candidate bands with the 90.5 kDa predicted mass and the documented isoform sequence changes. The supplied record has no observed band position, so band size alone cannot establish identity or assign a band to an isoform.
Boster reagents
ZFX 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-ZFX Antibody
Cat # A05947
A05947 is an anti-ZFX antibody listed for human, mouse, and rat. Its supplied Western blot image shows Daudi cell lysate tested at 1 μg/mL. No publication evidence or additional tested specimens are supplied.
Which to pick: A05947 is the only listed option and has a Western blot image from Daudi cell lysate. Check its listed reactivity against your sample; the supplied image does not establish performance in mouse or rat specimens.
Source: BosterBio ZFX gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P17010.
- Human Protein Atlas. ZFX tissue expression.
- PMC3774613 — target-verified WB comparison
- PMC3259083 — target-verified WB comparison
- PMC5008364 — target-verified WB comparison