This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
EPHX1 / Epoxide hydrolase 1 · Western blot design guide
Design a Western Blot for EPHX1
Source-linked EPHX1 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-EPHX1 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
EPHX1 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 | ~52.9 kDa | |
| Observed band | ~53 kDa | |
| Gel | 5–20% (catalog A00899-1) | |
| Positive control | Adrenal gland (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 | Methylated | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked EPHX1 Western Blot Protocol Options
The A00899-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human HepG2, human HCCT, human HCCP, rat liver, rat RH35, mouse liver, mouse HEPA1-6 (catalog A00899-1) |
| Gel % | 5–20% (catalog A00899-1) |
| Load | 30 ug; reducing conditions (catalog A00899-1) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A00899-1) |
| Membrane | nitrocellulose membrane (catalog A00899-1) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A00899-1) |
| Primary antibody | A00899-1 · 0.5 μg/mL (catalog A00899-1) |
| Primary incubation | overnight at 4°C (catalog A00899-1) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A00899-1) |
| Secondary incubation | 1.5 hour at RT (catalog A00899-1) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A00899-1) |
| Detection | ECL (catalog A00899-1) |
Section 2
What Is the Expected EPHX1 Western Blot Band Size?
EPHX1 is predicted at 52.9 kDa and observed near 53 kDa; the supplied evidence establishes no meaningful difference between them.
| Band near 53 kDa | Matches the reported EPHX1 band and its 52.9 kDa predicted mass. |
| Band near 53 kDa in whole-cell lysate | Consistent with the reported EPHX1 signal in HepG2 and rat and mouse cell lysates. |
| Band near 53 kDa in liver tissue lysate | Consistent with the reported EPHX1 signal in rat and mouse liver. |
| Weak band in a soluble fraction | EPHX1 is a microsomal membrane protein and may be depleted from that fraction. |
💡Expected EPHX1 appearanceEPHX1 has a predicted mass of 52.9 kDa and an observed band near 53 kDa; confirm identity with an appropriate antibody control and a membrane-enriched sample.
| Predicted full-length mass | Places EPHX1 near 52.9 kDa, consistent with the observed ~53 kDa band. |
| 455-residue sequence | Defines the full-length protein used for the predicted mass; no shorter product is specified. |
| Arg295 dimethylation | Is annotated, but no measurable migration effect is established. |
| Single-pass microsomal membrane localization | Can affect recovery during extraction; no band-size change is established. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Membrane-associated EPHX1 may be poorly recovered. | Check extraction conditions and a microsomal fraction. |
| Weak or no signal | The sampled fraction may contain little EPHX1. | Compare whole-cell and membrane-enriched samples with a positive control. |
| Band higher than expected | Its identity is unclear; the supplied features do not establish a larger EPHX1 species. | Check antibody specificity and compare with the ~53 kDa reference band. |
| Band lower than expected | Its identity is unclear; no signal-peptide or propeptide cleavage is annotated. | Check sample integrity and antibody specificity. |
| Multiple bands | The supplied isoform record does not establish distinct EPHX1 bands. | Compare bands with the ~53 kDa reference and use an independent antibody control. |
Experiment evidence
Western blot images showing EPHX1 band
A00899-1 · EPHX1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for EPHX1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for EPHX1 in Western blot, you can use adrenal gland tissue, which HPA rates as highly expressed.
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 such as Ponceau alongside the samples.
⚠️Feasibility: EPHX1 is an ER and microsomal membrane protein, so a membrane-enriched lysate may give a stronger signal.
HPA tissue expression evidence for EPHX1
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 | glandular cells | High | Protein (IHC) | HPA → |
| Liver | hepatocytes | High | Protein (IHC) | HPA → |
| Testis | Leydig cells | High | Protein (IHC) | HPA → |
| Lung | alveolar cells type II | Medium | Protein (IHC) | HPA → |
| Pancreas | exocrine glandular cells | 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 → |
| Caudate | glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced EPHX1 Western Blot Tips
Deeper troubleshooting and optimisation questions for EPHX1, answered from its protein features.
How should EPHX1 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 EPHX1 isoforms explain additional bands?
Isoforms · The supplied record lists one isoform and no alternative sequence. Do not assign additional bands to EPHX1 isoforms based on these features alone.
Do annotated modifications prove a band shift?
PTM · The linked UniProt record describes protein features. A modification annotation alone does not demonstrate a visible shift; retain any condition or experimental qualifier attached to it.
Does this guide establish induction of EPHX1?
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 EPHX1 Western blot?
Transfer · EPHX1 is a single-pass ER and microsome membrane protein with a predicted mass of 52.9 kDa. Check that your transfer recovers a band near 53 kDa. The supplied features do not specify a preferred transfer method.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A00899-1 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 EPHX1 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.
Why is the EPHX1 band near 53 kDa?
Interpretation · EPHX1 has a predicted mass of 52.9 kDa, close to the reported apparent band at ~53 kDa. Its listed features do not establish a modification-driven band shift.
UniProt lists dimethylated arginine at position 295, using UniProt sequence numbering. This modification alone does not establish a visible band shift or explain an unexpected apparent mass.
Compare the ~53 kDa band using the same sample fraction across lanes. EPHX1 is located in ER and microsome membranes, so differences in membrane recovery could affect measured intensity.
Start with the expected ~53 kDa band. The record lists one isoform, no glycosylation sites, and no disulfides; it does not support assigning other bands to those features. Dimethylation at UniProt Arg295 also does not, by itself, establish a visible shift.
UniProt lists dimethylated arginine at position 295, using UniProt sequence numbering. This modification alone does not establish a visible band shift or explain an unexpected apparent mass.
Compare the ~53 kDa band using the same sample fraction across lanes. EPHX1 is located in ER and microsome membranes, so differences in membrane recovery could affect measured intensity.
Start with the expected ~53 kDa band. The record lists one isoform, no glycosylation sites, and no disulfides; it does not support assigning other bands to those features. Dimethylation at UniProt Arg295 also does not, by itself, establish a visible shift.
Boster reagents
EPHX1 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-Epoxide hydrolase/EPHX1 Antibody Picoband®
Cat # A00899-1
The catalog reports one anti-EPHX1 antibody, A00899-1. Its Western blot image shows an approximately 53 kDa band in the named human, rat, and mouse lysates. The supplied evidence is a product image; no independent validation is provided.
Which to pick: A00899-1 is the only listed option. Its WB image includes human HepG2, HCCT and HCCP; rat liver and RH35; and mouse liver and HEPA1-6 samples, using 30 µg lysate per lane and 0.5 µg/mL primary antibody.
Source: BosterBio EPHX1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P07099.
- Human Protein Atlas. EPHX1 tissue expression.
- PMC11848624 — target-verified WB comparison
- PMC12315303 — target-verified WB comparison
- PMC8331186 — target-verified WB comparison