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- Table of Contents
CLK1 / Dual specificity protein kinase CLK1 · Western blot design guide
Design a Western Blot for CLK1
Source-linked CLK1 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-CLK1 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
CLK1 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 | ~57.3 kDa | |
| Gel | 12–15% (standard starting point) | |
| Positive control | Appendix (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 | |
| Caveat | Peptide-blocking control | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 3 isoform(s) |
Section 1
Source-Linked CLK1 Western Blot Protocol Options
The A03271 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | various cells (catalog A03271) |
| Gel % | 12–15% (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 | A03271 · 1:500 (catalog A03271) |
| 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 CLK1 Western Blot Band Size?
CLK1 is predicted at 57.3 kDa; splice isoforms and phosphorylation could affect migration, but no empirical band size or visible shift is established.
| Band near 57.3 kDa | Compatible with predicted CLK1 size; confirm identity with band controls |
| Nearby doublet | Could reflect phosphorylation at Ser61, Thr138 or Ser140; a visible shift is unproven |
| Several bands | Could include CLK1 isoforms 1, 2 and 3; their migration is unknown |
| Weak lysate band with stronger nuclear-fraction signal | Consistent with CLK1 nuclear localization |
💡Expected CLK1 appearanceUniProt predicts CLK1 at 57.3 kDa, but no empirical band size or resolvable isoform pattern is supplied; verify a candidate band with nuclear enrichment and peptide competition.
| UniProt predicted mass | Places the reference size near 57.3 kDa |
| Splice isoform 1 | May migrate differently from other isoforms; its mass is unspecified |
| Splice isoform 2 | May migrate differently from other isoforms; its mass is unspecified |
| Splice isoform 3 | May migrate differently from other isoforms; its mass is unspecified |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Nuclear CLK1 may be weak in whole-cell lysate | Check a nuclear fraction and fractionation controls |
| Band higher than expected | The band may be an unverified CLK1 form or nonspecific signal | Compare nuclear enrichment and peptide-blocked lanes |
| Band lower than expected | Its identity is uncertain because isoform masses are unavailable | Check peptide competition and compare antibodies recognizing different epitopes |
| Multiple bands | CLK1 has three splice isoforms and documented phosphorylation sites, but distinct band migration is unproven | Check which bands enrich in nuclear fractions and disappear with peptide competition |
| Weak or no signal | CLK1 is localized to the nucleus | Enrich the nuclear fraction and verify the fractionation |
Experiment evidence
Western blot images showing CLK1 band
A03271 · CLK1
A03271 · CLK1
A03271 · CLK1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for CLK1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for CLK1 in Western blot, you can use Appendix tissue, which HPA reports as a medium positive.
Positive control: Appendix (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: CLK1 is nuclear, and HPA reports only medium staining in the positive tissues, so the signal may be modest in whole-tissue lysate.
HPA tissue expression evidence for CLK1
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 |
|---|---|---|---|---|
| Appendix | glandular cells | Medium | Protein (IHC) | HPA → |
| Caudate | neuronal cells | Medium | Protein (IHC) | HPA → |
| Cerebellum | Purkinje cells | Medium | Protein (IHC) | HPA → |
| Cerebral cortex | glial cells | Medium | Protein (IHC) | HPA → |
| Epididymis | 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 → |
| Heart muscle | cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Hippocampus | glial cells | Not detected | Protein (IHC) | HPA → |
| Oral mucosa | squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CLK1 Western Blot Tips
Deeper troubleshooting and optimisation questions for CLK1, answered from its protein features.
How should CLK1 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 might CLK1 isoform 2 affect the blot?
Isoforms · Isoform 2 replaces canonical residues 131–136 and lacks residues 137–484. It may produce a smaller band if the antibody recognizes a retained region. An antibody targeting the missing region would not detect it; check the antibody epitope.
Isoform 3 has an altered, extended N terminus relative to canonical CLK1. It could migrate differently, but no observed position is supplied. Check whether the antibody recognizes a region shared with isoform 3.
Isoform 3 has an altered, extended N terminus relative to canonical CLK1. It could migrate differently, but no observed position is supplied. Check whether the antibody recognizes a region shared with isoform 3.
Which CLK1 phosphorylation sites matter when interpreting bands?
PTM · UniProt lists phosphoserine 61, phosphothreonine 138, and phosphoserine 140 in canonical sequence numbering. Residues 138 and 140 are absent from isoform 2. Phosphorylation at these sites does not by itself establish a visible band shift; use a suitable phosphatase comparison to investigate one. Numbering may differ for isoform 3 because its N terminus changes.
Does this guide establish induction of CLK1?
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 CLK1 Western blot?
Transfer · Canonical CLK1 is predicted at 57.3 kDa, so a routine transfer suitable for proteins around that size is a starting point. Check transfer with a total protein stain. The supplied features do not establish an optimal membrane, transfer setting, or band position.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A03271 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 CLK1 bands be quantified?
Quantitation · CLK1 has three annotated isoforms, and phosphorylation is recorded at three canonical positions. Quantify consistently defined bands across samples and use a loading measure for normalization. If multiple bands appear, report them separately unless their identities have been established.
Should CLK1 run at exactly 57.3 kDa?
Interpretation · 57.3 kDa is the predicted mass of canonical CLK1; no observed band position is supplied. Compare the band with a molecular weight marker, but do not attribute any difference to phosphorylation or splicing without further evidence.
Consider the annotated isoforms: isoform 2 lacks canonical residues 137–484, while isoform 3 has an altered N terminus. Check antibody epitope coverage and compare the band with the predicted 57.3 kDa canonical protein. The listed phosphorylation sites alone cannot identify an unexpected band.
CLK1 is annotated in the nucleus. Include a nuclear fraction when assessing its detection, and verify fraction quality with appropriate markers. A weak signal in another fraction alone does not establish that CLK1 is absent from the sample.
Consider the annotated isoforms: isoform 2 lacks canonical residues 137–484, while isoform 3 has an altered N terminus. Check antibody epitope coverage and compare the band with the predicted 57.3 kDa canonical protein. The listed phosphorylation sites alone cannot identify an unexpected band.
CLK1 is annotated in the nucleus. Include a nuclear fraction when assessing its detection, and verify fraction quality with appropriate markers. A weak signal in another fraction alone does not establish that CLK1 is absent from the sample.
Boster reagents
CLK1 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-CLK1 Antibody
Cat # A03271
A03271 is an anti-CLK1 antibody listed as reactive with human and mouse. Its WB image caption reports a 1:500 dilution on nuclear extracts from various cells, but does not identify the cells or establish species-specific WB performance.
Which to pick: A03271 is the only listed antibody and has a WB image. Consider its reported human and mouse reactivity and the caption’s nuclear-extract conditions when assessing it for your sample.
Source: BosterBio CLK1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P49759.
- Human Protein Atlas. CLK1 tissue expression.
- PMC11326178 — target-verified WB comparison
- PMC3537621 — target-verified WB comparison
- PMC9498672 — target-verified WB comparison