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- Table of Contents
BACH1 / Transcription regulator protein BACH1 · Western blot design guide
Design a Western Blot for BACH1
Real validated BACH1 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-BACH1 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
BACH1 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 | ~82 kDa | |
| Observed band | ~100 kDa | |
| Gel | 8% (catalog A02347-2) | |
| Positive control | Cervix (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 + Ubl conjugation | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 1 isoform(s) |
Section 1
Real Curated BACH1 Western Blot Protocols
The A02347-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human MCF-7, human HEL, human K562 (catalog A02347-2) |
| Gel % | 8% (catalog A02347-2) |
| Load | 30 ug; reducing conditions (catalog A02347-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A02347-2) |
| Membrane | nitrocellulose membrane (catalog A02347-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A02347-2) |
| Primary antibody | A02347-2 · 0.5 μg/mL (catalog A02347-2) |
| Primary incubation | overnight at 4°C (catalog A02347-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A02347-2) |
| Secondary incubation | 1.5 hour at RT (catalog A02347-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A02347-2) |
| Detection | ECL (catalog A02347-2) |
Section 2
What Is the Expected BACH1 Western Blot Band Size?
BACH1 is predicted at 82 kDa and observed near 100 kDa; the cause of the difference is not established.
| Band near 100 kDa | Empirical BACH1 band reported in reducing whole-cell lysates; confirm identity with antibody controls. |
| Band near 82 kDa | Near the predicted full-length mass, but identity requires confirmation. |
| Little signal in a cytosolic fraction | Consistent with BACH1's nuclear location; check the nuclear fraction. |
| Doublet near the main band | BACH1 has documented phosphorylation sites, but their effect on migration is unestablished. |
💡Expected BACH1 appearanceBACH1 has a predicted mass of 82 kDa, while antibody QC reports a band near 100 kDa in reducing whole-cell lysates; the cause of the difference is unestablished, so confirm band identity with ordinary controls.
| Predicted full-length mass | 82 kDa is the sequence-based reference; the observed band is near 100 kDa. |
| Phosphoserine at residue 196 | A documented modification whose effect on apparent band size is unestablished. |
| Phosphoserine at residue 364 | A documented modification whose effect on apparent band size is unestablished. |
| Phosphoserine at residue 445 | A documented modification whose effect on apparent band size is unestablished. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Nuclear BACH1 may be poorly recovered during extraction. | Check nuclear protein recovery and a positive control. |
| Band higher than expected | The reported ~100 kDa band exceeds the 82 kDa predicted mass; its cause is unestablished. | Compare with the reported band and verify identity using a second antibody or BACH1 depletion. |
| Band lower than expected | A smaller band is not explained by a listed signal peptide or propeptide. | Check sample integrity and verify identity using BACH1 depletion. |
| Multiple bands | Phosphorylation is documented, but distinct migrating forms are unproven. | Compare bands after phosphatase treatment and verify identity with BACH1 depletion. |
| Weak or no signal | Nuclear extraction or low BACH1 abundance may limit detection. | Check loading, nuclear recovery, and a positive-control lysate. |
| Fragments below expected size | Sample degradation or antibody cross-reactivity may produce smaller bands. | Use fresh protected lysate and verify fragments with a second antibody or BACH1 depletion. |
Experiment evidence
Western blot images showing BACH1 band
A02347-2 · BACH1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for BACH1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for BACH1 in Western blot, you can use cervix tissue, which HPA scores High.
Positive control: Cervix (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: Because BACH1 is nuclear, nuclear-enriched lysate may improve detection.
HPA tissue expression evidence for BACH1
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 |
|---|---|---|---|---|
| Cervix | squamous epithelial cells | High | Protein (IHC) | HPA → |
| Esophagus | squamous epithelial cells | High | Protein (IHC) | HPA → |
| Oral mucosa | squamous epithelial cells | High | Protein (IHC) | HPA → |
| Skin | epidermal cells | High | Protein (IHC) | HPA → |
| Testis | elongated or late spermatids | 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 → |
| Adrenal gland | glandular cells | Not detected | Protein (IHC) | HPA → |
| Breast | adipocytes | Not detected | Protein (IHC) | HPA → |
| Caudate | glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | cells in granular layer | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced BACH1 Western Blot Tips
Deeper troubleshooting and optimisation questions for BACH1, answered from its protein features.
How should BACH1 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 a second BACH1 band represent a listed isoform?
Isoforms · The supplied record lists one isoform and no alternative sequence. It does not support assigning a second band to a BACH1 isoform; verify the band before interpreting it.
Which BACH1 phosphorylation sites matter when interpreting bands?
PTM · UniProt lists phosphoserine at positions 196, 364, and 445. These are UniProt sequence coordinates; antibody or paper numbering may differ. Their presence alone does not demonstrate a visible band shift.
Does this guide establish induction of BACH1?
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 BACH1?
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 A02347-2 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 BACH1 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 might BACH1 appear near 100 kDa instead of 82 kDa?
Interpretation · The supplied predicted mass is 82 kDa, while the observed band is approximately 100 kDa. The listed features do not establish the cause of this difference. Confirm band identity before assigning the difference to a modification.
BACH1 is annotated with Ubl conjugation, but no conjugation site or resulting band size is supplied. This annotation alone cannot identify a higher band. Check whether the band behaves consistently across samples before assigning it to modified BACH1.
BACH1 is annotated as nuclear. A nuclear fraction is therefore relevant when assessing its signal; compare fractions consistently when interpreting band intensity.
Quantify the same verified BACH1 band across samples and keep the sample fraction consistent. Because BACH1 is annotated as nuclear, changes in nuclear band intensity should be interpreted as changes in that fraction, not automatically as changes in total BACH1.
BACH1 is annotated with Ubl conjugation, but no conjugation site or resulting band size is supplied. This annotation alone cannot identify a higher band. Check whether the band behaves consistently across samples before assigning it to modified BACH1.
BACH1 is annotated as nuclear. A nuclear fraction is therefore relevant when assessing its signal; compare fractions consistently when interpreting band intensity.
Quantify the same verified BACH1 band across samples and keep the sample fraction consistent. Because BACH1 is annotated as nuclear, changes in nuclear band intensity should be interpreted as changes in that fraction, not automatically as changes in total BACH1.
Boster reagents
BACH1 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-BACH1.3/BACH1 Antibody Picoband®
Cat # A02347-2
A02347-2 is a rabbit polyclonal anti-BACH1 antibody listed for human, mouse, and rat. Its WB caption reports a band near 100 kDa in human MCF-7, HEL, and K562 cells and rat and mouse testis lysates; the expected size is 82 kDa.
Which to pick: A02347-2 is the only listed antibody and has a WB image using the named human cell lines and rodent testis samples. Consider whether those samples fit your experiment, and note the difference between the observed 100 kDa band and expected 82 kDa size.
Source: BosterBio BACH1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry O14867.
- Human Protein Atlas. BACH1 tissue expression.
- PMC5179191 — target-verified WB comparison
- PMC12314328 — target-verified WB comparison
- PMC12130529 — target-verified WB comparison