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- Table of Contents
FDX1 / Adrenodoxin, mitochondrial · Western blot design guide
Design a Western Blot for FDX1
Real validated FDX1 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-FDX1 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
FDX1 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 | ~19.4 kDa | |
| Observed band | ~14 kDa | |
| Gel | 10% (catalog A05441-1) | |
| Positive control | Adrenal gland (IHC candidate; verify WB) +3 more | |
| Negative control | Adipose tissue (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Acetylated | |
| Caveat | Band identity controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 1 isoform(s) |
Section 1
Real Curated FDX1 Western Blot Protocols
The A05441-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human A549, human 293T, human HepG2 (catalog A05441-1) |
| Gel % | 10% (catalog A05441-1) |
| Load | 30 ug; reducing conditions (catalog A05441-1) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A05441-1) |
| Membrane | nitrocellulose membrane (catalog A05441-1) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A05441-1) |
| Primary antibody | A05441-1 · 1:1000 (catalog A05441-1) |
| Primary incubation | overnight at 4°C (catalog A05441-1) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A05441-1) |
| Secondary incubation | 1.5 hour at RT (catalog A05441-1) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A05441-1) |
| Detection | ECL (catalog A05441-1) |
Section 2
What Is the Expected FDX1 Western Blot Band Size?
FDX1 is predicted at 19.4 kDa, but antibody QC reports approximately 14 kDa; the cause of this difference is not established.
| Band near 14 kDa | Empirical FDX1 band reported in reducing whole-cell and tissue lysates; confirm identity with controls |
| Band near 19.4 kDa | Near the UniProt predicted mass; identity requires confirmation |
| Doublet near the main band | Listed phosphorylation or lysine modifications could affect mobility, but distinct bands are unproven |
| Faint band in whole-cell lysate | FDX1 is located in the mitochondrial matrix; compare with an enriched mitochondrial fraction |
💡Expected FDX1 appearanceFDX1 has a predicted mass of 19.4 kDa, while antibody QC reports a band near 14 kDa; the cause of the difference is unestablished, so confirm identity with appropriate controls.
| UniProt predicted mass | 19.4 kDa provides the sequence-based reference, while the reported band is near 14 kDa |
| Phosphoserine 63 | Could affect mobility; no visible shift is established |
| Phosphoserine 177 | Could affect mobility; no visible shift is established |
| Alternate acetylation or succinylation at lysine 66 | Could affect mobility; no distinct band is established |
| Succinylation at lysine 158 | Could affect mobility; no visible shift is established |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | FDX1 is localized to the mitochondrial matrix | Check an enriched mitochondrial fraction and a positive-control lysate |
| Band higher than expected | Listed modifications might affect mobility, but a higher FDX1 band is unverified | Compare with the reported 14 kDa band and confirm identity with an independent antibody or FDX1 depletion |
| Band lower than expected | The reported 14 kDa band runs below the 19.4 kDa predicted mass; its cause is unestablished | Check a molecular-weight marker and confirm band identity with FDX1 depletion or an independent antibody |
| Multiple bands | Listed modifications could contribute, but distinct FDX1 forms are unproven | Compare phosphatase-treated samples and verify each band with FDX1 depletion |
| Weak or no signal | Mitochondrial matrix FDX1 may be difficult to detect in the tested lysate | Compare whole-cell lysate with an enriched mitochondrial fraction and a positive control |
Experiment evidence
Western blot images showing FDX1 band
A05441-1 · FDX1
M05441 · FDX1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for FDX1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for FDX1 in Western blot, you can use adrenal gland lysate.
Positive control: Adrenal gland (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside.
⚠️Feasibility: FDX1 is mitochondrial, and HPA lists adipose tissue as not detected, providing a candidate negative tissue.
HPA tissue expression evidence for FDX1
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 |
|---|---|---|---|---|
| Adrenal gland | cells in zona fasciculata | High | Protein (IHC) | HPA → |
| Placenta | syncytiotrophoblasts - cell body | High | Protein (IHC) | HPA → |
| Testis | Leydig cells | High | Protein (IHC) | HPA → |
| Kidney | proximal tubules (cell body) | Medium | 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 → |
| Bone marrow | hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | adipocytes | Not detected | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced FDX1 Western Blot Tips
Deeper troubleshooting and optimisation questions for FDX1, answered from its protein features.
How should FDX1 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 FDX1 isoforms explain multiple bands?
Isoforms · Only one isoform is supplied, with no alternative sequence. The supplied features therefore do not support assigning additional bands to FDX1 isoforms.
Which FDX1 modifications matter when interpreting bands?
PTM · The supplied UniProt positions are phosphoserine 63 and 177, alternate N6-acetyllysine or N6-succinyllysine at 66, and N6-succinyllysine at 158. Keep these UniProt coordinates explicit when comparing with antibody or paper numbering. Listed sites alone do not establish a visible band shift.
Position 66 is listed with alternate acetylation and succinylation. A band’s position alone cannot tell which modification is present. Check the antibody’s target and the numbering convention before linking a band to this UniProt site.
Position 66 is listed with alternate acetylation and succinylation. A band’s position alone cannot tell which modification is present. Check the antibody’s target and the numbering convention before linking a band to this UniProt site.
Does this guide establish induction of FDX1?
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 FDX1?
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 A05441-1 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 FDX1 be quantified across samples?
Quantitation · FDX1 is located in the mitochondrial matrix. Use the same sample preparation and loading basis across samples, and account for differences in mitochondrial content when interpreting changes in FDX1 signal. Its interaction with CYP11A1 does not by itself establish a change in FDX1 abundance.
Why might FDX1 appear near 14 kDa instead of 19.4 kDa?
Interpretation · The supplied apparent band is ~14 kDa, while the predicted mass is 19.4 kDa. FDX1 is mitochondrial and has a transit peptide keyword, so processing is a possibility, but no cleavage site is supplied. These features do not establish the cause of the difference; confirm the band’s identity before assigning it to FDX1.
FDX1 interacts with CYP11A1 and has listed phosphorylation, acetylation, and succinylation sites. Those features do not establish that an unexpected band is a complex or a modified FDX1 species. Verify band identity before assigning a mechanism.
FDX1 interacts with CYP11A1 and has listed phosphorylation, acetylation, and succinylation sites. Those features do not establish that an unexpected band is a complex or a modified FDX1 species. Verify band identity before assigning a mechanism.
Boster reagents
FDX1 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-FDX1 Antibody Picoband®
Cat # A05441-1
Real WB data
Anti-ADX Rabbit Monoclonal Antibody
Cat # M05441
Two the supplier FDX1 antibodies have Western blot images. A05441-1 was tested on human cell and rat and mouse ovary lysates; M05441 was tested on human and rat cell lysates. Reported bands were approximately 13–14 kDa, below the stated 19 kDa expected size.
Which to pick: Choose A05441-1 for its documented human cell and mouse ovary examples, or M05441 when listed rat reactivity and rat cell examples matter. Both have WB images; the supplied evidence does not establish which performs better.
Source: BosterBio FDX1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P10109.
- Human Protein Atlas. FDX1 tissue expression.
- PMC12748187 — target-verified WB comparison
- PMC12116747 — target-verified WB comparison
- PMC11366913 — target-verified WB comparison