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LGR5 / Leucine-rich repeat-containing G-protein coupled receptor 5 · Western blot design guide
Design a Western Blot for LGR5
Real validated LGR5 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-LGR5 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
LGR5 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 | ~100 kDa | |
| Observed band | ~130 kDa | |
| Gel | 5–20% (catalog A00239-2) | |
| Positive control | Bronchus (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 + Cleaved | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 3 isoform(s) |
Section 1
Real Curated LGR5 Western Blot Protocols
The A00239-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human SH-SY5Y, human HepG2, human RT4 (catalog A00239-2) |
| Gel % | 5–20% (catalog A00239-2) |
| Load | 30 ug; reducing conditions (catalog A00239-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A00239-2) |
| Membrane | nitrocellulose membrane (catalog A00239-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A00239-2) |
| Primary antibody | A00239-2 · 0.5 μg/mL (catalog A00239-2) |
| Primary incubation | overnight at 4°C (catalog A00239-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A00239-2) |
| Secondary incubation | 1.5 hour at RT (catalog A00239-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A00239-2) |
| Detection | ECL (catalog A00239-2) |
Section 2
What Is the Expected LGR5 Western Blot Band Size?
LGR5 has a predicted 100 kDa precursor and a reported band near 130 kDa; the cause of the difference is not established.
| Band near 130 kDa | Empirically reported LGR5-reactive band; confirm identity with an appropriate control |
| Band near 100 kDa | Possible migration near the predicted precursor mass; identity requires confirmation |
| Band below the precursor position | Signal-peptide removal or an isoform could contribute; the resulting migration is unknown |
| Several bands at different positions | Isoforms 1, 2, and 3 could contribute, but distinct migration is not established |
| Broad band or smear | Heterogeneous N-linked glycosylation is possible but has not been shown to cause this pattern |
💡Expected LGR5 appearanceUniProt predicts a 100 kDa LGR5 precursor, while antibody QC reports a band near 130 kDa; the cause of the difference is unestablished, so confirm band identity with appropriate controls.
| Predicted precursor mass | Provides a 100 kDa sequence-based reference, not a measured migration |
| N-linked glycosylation at Asn-63, Asn-77, Asn-208, Asn-500, and Asn-792 | Could alter apparent mass; its contribution to the 130 kDa band is unestablished |
| Signal peptide at residues 1–21 | Cleavage yields a shorter mature chain, but its apparent band size is not supplied |
| Splice isoforms 1, 2, and 3 | Could differ in size; their individual masses and migration patterns are not supplied |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Membrane-localized LGR5 may be poorly recovered during extraction | Check a membrane-enriched fraction and detergent solubilization with a positive control |
| Band higher than expected | The reported 130 kDa band exceeds the 100 kDa predicted mass; its cause is unestablished | Compare with a validated positive control and test whether N-linked deglycosylation changes migration |
| Band lower than expected | Signal-peptide cleavage or an isoform could contribute | Confirm identity with a positive control or an independent LGR5 antibody |
| Broad smear instead of sharp band | Heterogeneous N-linked glycosylation is possible | Compare migration before and after N-linked deglycosylation and verify band identity |
| Multiple bands | Isoforms or different glycosylation states could contribute | Compare a positive control and deglycosylated sample; verify candidate bands independently |
| Weak or no signal | Membrane extraction or low LGR5 recovery may limit detection | Check sample preparation and antibody performance with a validated positive control |
Experiment evidence
Western blot images showing LGR5 band
A00239-2 · LGR5
M00239 · LGR5
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for LGR5 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for LGR5 in Western blot, you can use bronchus tissue, which HPA rates High.
Positive control: Bronchus (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: As a multipass membrane protein, LGR5 may require effective membrane protein extraction for a clear signal.
HPA tissue expression evidence for LGR5
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 |
|---|---|---|---|---|
| Bronchus | ciliated cells (cilia axoneme) | High | Protein (IHC) | HPA → |
| Fallopian tube | ciliated cells (cilia axoneme) | High | Protein (IHC) | HPA → |
| Nasopharynx | ciliated cells (cilia axoneme) | High | Protein (IHC) | HPA → |
| Cerebral cortex | endothelial cells | Medium | Protein (IHC) | HPA → |
| Cervix | 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 LGR5 Western Blot Tips
Deeper troubleshooting and optimisation questions for LGR5, answered from its protein features.
How should LGR5 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 LGR5 isoforms produce different bands?
Isoforms · UniProt lists three isoforms. Relative to the canonical sequence, isoform 3 lacks residues 143–214 and isoform 2 lacks residues 263–286. These deletions could affect migration. Check which isoforms the antibody can recognize; residue 208 lies within the segment missing from isoform 3. Coordinates here use canonical UniProt numbering.
Which glycosylation sites matter when assessing a band shift?
PTM · UniProt lists N-linked sites at Asn-63, Asn-77, Asn-208, Asn-500, and Asn-792 in canonical sequence numbering. Compare treated and untreated samples if testing glycosylation, but do not assume these sites produce a visible shift.
Does this guide establish induction of LGR5?
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 LGR5 Western blot?
Transfer · LGR5 is a 907-residue multipass membrane protein, and the supplied observed band is about 130 kDa. Check transfer efficiency in that region when choosing conditions. The supplied features do not establish one specific transfer method.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A00239-2 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 I quantify LGR5 bands?
Quantitation · Use the same sample preparation and band definition across comparisons. LGR5 occurs at the cell membrane and trans-Golgi network, and is rapidly internalized at steady state; a change in a membrane-enriched fraction may reflect redistribution. Report which fraction and band were measured.
Why might LGR5 run near 130 kDa instead of 100 kDa?
Interpretation · The supplied predicted mass is 100 kDa and the observed band is about 130 kDa. LGR5 has five annotated N-linked glycosylation sites, which could affect migration, but these features alone do not establish the cause or size of the difference.
Consider the annotated splice variants before assigning identity: isoform 3 lacks residues 143–214, while isoform 2 lacks 263–286 in canonical UniProt numbering. The features do not establish where either isoform migrates, so band size alone cannot identify it.
UniProt lists five disulfide bonds for LGR5. Keep reducing conditions consistent when comparing samples, and compare reducing and nonreducing lanes if investigating an unexpected band. The annotation alone does not predict a particular band shift.
Consider the annotated splice variants before assigning identity: isoform 3 lacks residues 143–214, while isoform 2 lacks 263–286 in canonical UniProt numbering. The features do not establish where either isoform migrates, so band size alone cannot identify it.
UniProt lists five disulfide bonds for LGR5. Keep reducing conditions consistent when comparing samples, and compare reducing and nonreducing lanes if investigating an unexpected band. The annotation alone does not predict a particular band shift.
Boster reagents
LGR5 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-LGR5 Antibody Picoband®
Cat # A00239-2
Real WB data
Anti-LGR5/GPR49 Rabbit Monoclonal Antibody
Cat # M00239
Both listed anti-LGR5 antibodies report human, mouse, and rat reactivity and have WB images. A00239-2 shows an approximately 130 kDa band in four human cell lysates, versus an expected 92–100 kDa; M00239 shows human fetal skeletal muscle lysate.
Which to pick: Choose A00239-2 for documented human cell lysate examples with detailed WB conditions, or M00239 for a human fetal skeletal muscle example. Both list mouse and rat reactivity, but neither caption documents WB testing in those species.
Source: BosterBio LGR5 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry O75473.
- Human Protein Atlas. LGR5 tissue expression.
- PMC6050418 — target-verified WB comparison
- PMC4253421 — target-verified WB comparison
- PMC5432274 — target-verified WB comparison