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- Table of Contents
LEFTY1 / Left-right determination factor 1 · Western blot design guide
Design a Western Blot for LEFTY1
Source-linked LEFTY1 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-LEFTY1 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
LEFTY1 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 | ~40.9 kDa | |
| Gel | 12–15% (standard starting point) | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Glycosylated + Cleaved | |
| Caveat | Processing-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked LEFTY1 Western Blot Protocol Options
The A05889-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | extracts of various cell lines, (catalog A05889-1) |
| 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 | Blocking buffer: 3% nonfat dry milk in TBST (catalog A05889-1) |
| Primary antibody | A05889-1 · 1:1000 (catalog A05889-1) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | Goat Anti-Rabbit IgG, 1:10000 (catalog A05889-1) |
| Secondary incubation | 1 h at room temperature (standard starting point) |
| Wash | 3 × 5 min in TBST (standard starting point) |
| Detection | ECL (catalog A05889-1) |
Section 2
What Is the Expected LEFTY1 Western Blot Band Size?
LEFTY1 has a predicted precursor mass of 40.9 kDa; cleavage and Asn158 glycosylation could affect migration, but no empirical band demonstrates their effects.
| Band near 40.9 kDa | Compatible with the predicted precursor size; identity needs confirmation |
| Band below 40.9 kDa | May reflect removal of the signal peptide or propeptide |
| Band above 40.9 kDa | Asn158 N-linked glycosylation could affect migration, but the site alone does not establish a shift |
| Little or no band in whole-cell lysate | LEFTY1 is secreted and may be found in conditioned medium |
💡Expected LEFTY1 appearanceUniProt predicts a 40.9 kDa precursor; secretion, cleavage and Asn158 N-linked glycosylation may affect what appears on a blot, but no empirical band size is supplied, so confirm identity with appropriate controls.
| UniProt predicted mass | The 40.9 kDa value describes the precursor, not a measured band |
| Signal peptide at residues 1–21 | Removal can yield a smaller form than the precursor |
| Propeptide at residues 22–76 | Removal can further reduce the mature protein's mass |
| N-linked glycosylation site at Asn158 | Glycosylation may alter apparent size, but a visible shift is not established |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Secreted LEFTY1 may be depleted from cells | Check concentrated conditioned medium alongside lysate |
| Band higher than expected | Asn158 glycosylation may affect migration, though its effect is unverified | Compare with a deglycosylated sample and verify band identity |
| Band lower than expected | Signal peptide and propeptide removal can reduce mass | Check whether the antibody recognizes the processed form and verify band identity |
| Multiple bands | Precursor and processed forms may coexist | Compare lysate and conditioned medium and verify each band independently |
| Weak or no signal | Secreted protein may be scarce in the tested fraction | Check conditioned medium and a suitable positive control |
| Fragments below expected size | Processing or sample degradation may produce smaller species | Use protease inhibitors during collection and verify fragment identity |
Experiment evidence
Western blot images showing LEFTY1 band
A05889-1 · LEFTY1
A05889 · LEFTY1
A05889 · LEFTY1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for LEFTY1 Western blot
🧪For positive controls for LEFTY1 in Western blot, you can use a separately validated LEFTY1-positive sample, since the supplied HPA evidence identifies no positive tissue or cell line.
Positive control: No high/medium HPA tissue identified
Negative control: Suggested KO / knockdown lysate
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: LEFTY1 is secreted, so conditioned medium may provide a stronger signal than whole-cell lysate.
HPA tissue expression evidence for LEFTY1
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 |
|---|---|---|---|---|
| No high/medium HPA tissues identified in the supplied evidence. | ||||
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| No lower-expression tissue rows available in the supplied evidence. | ||||
Section 3
Advanced LEFTY1 Western Blot Tips
Deeper troubleshooting and optimisation questions for LEFTY1, answered from its protein features.
How should LEFTY1 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.
Are multiple LEFTY1 bands evidence of different isoforms?
Isoforms · The supplied record lists one isoform and no alternative sequence. Do not assign additional bands to splice isoforms from this evidence. Compare their apparent sizes with the annotated signal peptide, propeptide, and Asn158 glycosylation site, but verify band identities experimentally.
Where is LEFTY1 glycosylated?
PTM · UniProt lists one N-linked glycosylation site, Asn158, using coordinates of the supplied full-length sequence. Keep that numbering explicit when comparing antibody or literature positions. The site annotation alone does not establish glycan occupancy or a visible band shift.
Does this guide establish induction of LEFTY1?
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 LEFTY1?
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 A05889-1 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 LEFTY1 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 LEFTY1 differ from its predicted 40.9 kDa mass?
Interpretation · 40.9 kDa refers to the full-length protein. LEFTY1 has a signal peptide at residues 1..21, a propeptide at 22..76, and an N-linked glycosylation site at Asn158. These features are relevant when interpreting apparent mass, but they do not establish a visible shift or predict an observed band size.
Check whether the antibody recognizes a region within the signal peptide (1..21), propeptide (22..76), or remaining sequence. The annotated regions may affect which forms retain the epitope. UniProt coordinates refer to the supplied 366-residue sequence; antibody numbering may use another convention.
LEFTY1 is annotated as secreted and has a signal peptide at residues 1..21. Consider measuring the extracellular fraction as well as cell lysate when choosing samples. Record the fraction used so bands from different sample types are interpreted consistently.
UniProt lists four disulfide bonds. Record whether samples were run under reducing conditions when comparing blots, since disulfide-dependent structure may affect migration. The annotation does not specify the size or direction of any band change.
Start with the 40.9 kDa full-length prediction, then consider the signal peptide (1..21), propeptide (22..76), and N-linked site at Asn158. These annotations offer possibilities to investigate, not identities for extra bands. Confirm an unexpected band's identity before using it for quantitation.
Check whether the antibody recognizes a region within the signal peptide (1..21), propeptide (22..76), or remaining sequence. The annotated regions may affect which forms retain the epitope. UniProt coordinates refer to the supplied 366-residue sequence; antibody numbering may use another convention.
LEFTY1 is annotated as secreted and has a signal peptide at residues 1..21. Consider measuring the extracellular fraction as well as cell lysate when choosing samples. Record the fraction used so bands from different sample types are interpreted consistently.
UniProt lists four disulfide bonds. Record whether samples were run under reducing conditions when comparing blots, since disulfide-dependent structure may affect migration. The annotation does not specify the size or direction of any band change.
Start with the 40.9 kDa full-length prediction, then consider the signal peptide (1..21), propeptide (22..76), and N-linked site at Asn158. These annotations offer possibilities to investigate, not identities for extra bands. Confirm an unexpected band's identity before using it for quantitation.
Boster reagents
LEFTY1 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-LEFTY1 Antibody
Cat # A05889-1
Real WB data
Anti-Lefty LEFTY1 Antibody
Cat # A05889
Both anti-LEFTY1 antibodies list Human and Mouse reactivity and include Western blot images. A05889-1 was used on various cell-line extracts at 1:1000; A05889 on HepG2-UV cells at 1:500. These captions document specific test contexts, not performance across all samples.
Which to pick: For HepG2-UV cells, A05889 has the specifically named WB example at 1:500. A05889-1 has a WB example using various cell-line extracts at 1:1000. Both list Human and Mouse reactivity; choose based on the closer sample and protocol match.
Source: BosterBio LEFTY1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry O75610.
- Human Protein Atlas. LEFTY1 tissue expression.
- PMC7765495 — target-verified WB comparison
- PMC4652748 — target-verified WB comparison
- PMC5609950 — target-verified WB comparison