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ELK1 / ETS domain-containing protein Elk-1 · Western blot design guide
Design a Western Blot for ELK1
Real validated ELK1 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-ELK1 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
ELK1 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 | ~44.9 kDa | |
| Observed band | ~50 kDa | |
| Gel | 5–20% (catalog A01426-1) | |
| Positive control | Cerebellum (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 | Glycosylated + Phosphorylated | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 2 isoform(s) |
Section 1
Real Curated ELK1 Western Blot Protocols
The A01426-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human placenta, human PC-3, human SiHa (catalog A01426-1) |
| Gel % | 5–20% (catalog A01426-1) |
| Load | 30 ug; reducing conditions (catalog A01426-1) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A01426-1) |
| Membrane | nitrocellulose membrane (catalog A01426-1) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A01426-1) |
| Primary antibody | A01426-1 · 0.5 μg/mL (catalog A01426-1) |
| Primary incubation | overnight at 4°C (catalog A01426-1) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A01426-1) |
| Secondary incubation | 1.5 hour at RT (catalog A01426-1) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A01426-1) |
| Detection | ECL (catalog A01426-1) |
Section 2
What Is the Expected ELK1 Western Blot Band Size?
ELK1 is predicted at 44.9 kDa and observed near 50 kDa; the cause of the difference is not established.
| Band near 50 kDa | Empirical ELK1 band reported in reducing whole-cell and tissue lysates; confirm identity with controls |
| Band near 44.9 kDa | Near the predicted sequence mass; identity still requires controls |
| Two nearby bands | Could reflect ELK1 phosphorylation, but a resolved shift is not established |
| Several bands at different positions | Isoforms 1 and 2 are annotated, but distinct migration is not established |
💡Expected ELK1 appearanceELK1 has a predicted mass of 44.9 kDa and an empirical band near 50 kDa; the cause of the difference is unestablished, so confirm identity with appropriate controls.
| Predicted sequence mass | 44.9 kDa is the sequence-based reference; an empirical band appears near 50 kDa |
| O-linked GlcNAc at Thr381 | A visible mobility change from this single modification is not established |
| Isoform 1 | Its separate apparent size relative to isoform 2 is not supplied |
| Isoform 2 | Its separate apparent size relative to isoform 1 is not supplied |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Nuclear ELK1 may be underrepresented in the sample | Check nuclear enrichment and a positive-control lysate |
| Band higher than expected | The reported ELK1 band near 50 kDa exceeds its 44.9 kDa predicted mass; the cause is unestablished | Compare with the reported band and confirm identity by ELK1 depletion |
| Band lower than expected | An isoform is possible, but its migration is not supplied | Check antibody epitope coverage and confirm the band by ELK1 depletion |
| Multiple bands | Annotated isoforms or phosphorylation may contribute, but distinct bands are not established | Confirm which bands track with ELK1 depletion |
| Weak or no signal | ELK1 may be scarce in the sampled whole-cell lysate | Check loading, antibody performance, and a nuclear or positive-control sample |
Experiment evidence
Western blot images showing ELK1 band
A01426-1 · ELK1
M01426 · ELK1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for ELK1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for ELK1 in Western blot, you can use cerebellum lysate, which has high HPA expression.
Positive control: Cerebellum (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: ELK1 is nuclear, so nuclear-enriched lysate may improve detection; adipose tissue is a reported not-detected control.
HPA tissue expression evidence for ELK1
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 |
|---|---|---|---|---|
| Cerebellum | granular cells - nucleus | High | Protein (IHC) | HPA → |
| Ovary | ovarian stroma cells | High | Protein (IHC) | HPA → |
| Placenta | syncytiotrophoblasts - cell body | High | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | 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 → |
| Bone marrow | hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Duodenum | glandular cells | Not detected | Protein (IHC) | HPA → |
| Liver | cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Lymph node | germinal center cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced ELK1 Western Blot Tips
Deeper troubleshooting and optimisation questions for ELK1, answered from its protein features.
What should be checked when unexpected ELK1 bands appear?
Band shift · Check antibody epitope coverage against both isoforms and compare bands with the observed approximately 50 kDa reference. Isoform 2 has a changed 91–95 segment and lacks residues 96–428. ELK1 also has listed phosphorylation and Thr381 O-linked GlcNAc features, but those annotations alone cannot identify an unexpected band.
How could ELK1 isoforms affect band interpretation?
Isoforms · UniProt lists isoforms 1 and 2. In isoform 2, residues 91–95 change from VAGCS to SHCAP, and residues 96–428 are missing. Check whether the antibody epitope is retained in isoform 2 before interpreting additional bands or comparing isoform signals.
Which ELK1 phosphorylation sites should a phospho-specific assay target?
PTM · In UniProt numbering, MAPK1 phosphorylates Ser324, Thr336, Thr353, Thr363, Thr368, Ser383, Ser389, Thr417 and Ser422; MAPK8 also phosphorylates Ser383. Match the antibody’s stated site and numbering convention to these coordinates. A listed site does not by itself predict a visible band shift.
The listed ELK1 sites include eight attributed to MAPK1 alone and Ser383 attributed to MAPK1 and MAPK8. Measure the relevant phosphosite when assessing phosphorylation associated with either enzyme, and compare it with total ELK1. These features do not specify a treatment or predict the size of a signal change.
UniProt lists O-linked GlcNAc at Thr381. Consider this modification when investigating band behavior, but do not assign a band or apparent mass difference to it without supporting evidence. Thr381 is a UniProt coordinate; check numbering if an antibody or paper uses another convention.
The listed ELK1 sites include eight attributed to MAPK1 alone and Ser383 attributed to MAPK1 and MAPK8. Measure the relevant phosphosite when assessing phosphorylation associated with either enzyme, and compare it with total ELK1. These features do not specify a treatment or predict the size of a signal change.
UniProt lists O-linked GlcNAc at Thr381. Consider this modification when investigating band behavior, but do not assign a band or apparent mass difference to it without supporting evidence. Thr381 is a UniProt coordinate; check numbering if an antibody or paper uses another convention.
Does this guide establish induction of ELK1?
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 ELK1?
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 A01426-1 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 ELK1 Western blot signals be quantified?
Quantitation · For phosphorylation measurements, compare a site-specific phospho-ELK1 signal with total ELK1 measured in the same samples. Confirm that the total-ELK1 antibody recognizes the isoforms being compared: isoform 2 lacks residues 96–428, which include every listed phosphorylation site.
Why might ELK1 appear near 50 kDa rather than 44.9 kDa?
Interpretation · The predicted mass is 44.9 kDa, while the supplied observed band is about 50 kDa. ELK1 has phosphorylation and O-linked GlcNAc features, but their presence alone does not establish the cause of this difference or a visible shift. Use the observed band as a reference and verify band identity experimentally.
Boster reagents
ELK1 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-ELK1 Antibody Picoband®
Cat # A01426-1
Real WB data
Anti-ELK1 Rabbit Monoclonal Antibody
Cat # M01426
The catalog reports two anti-ELK1 antibodies with WB images: polyclonal A01426-1 and rabbit monoclonal M01426. The captions show human, rat, and mouse lysates. Reported bands are approximately 50 and 60 kDa, respectively, versus an expected 45 kDa; no independent validation is supplied.
Which to pick: Choose by the lysates shown in each WB image: A01426-1 includes placenta, PC-3, SiHa, K562, and rat and mouse brain and ovary; M01426 includes Jurkat, Raji, Daudi, rat C6, mouse Neuro-2a, and rat and mouse brain. Consider the differing reported band sizes.
Source: BosterBio ELK1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P19419.
- Human Protein Atlas. ELK1 tissue expression.
- PMC8469350 — target-verified WB comparison
- PMC3511420 — target-verified WB comparison
- PMC7894012 — target-verified WB comparison