This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
GPRC5A / Retinoic acid-induced protein 3 · Western blot design guide
Design a Western Blot for GPRC5A
Source-linked GPRC5A Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-GPRC5A 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
GPRC5A 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 | ~40.3 kDa | |
| Observed band | ~37–50 kDa | |
| Gel | 10% (catalog A05072-2) | |
| Positive control | Lung (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 | Modification-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked GPRC5A Western Blot Protocol Options
The A05072-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human A375, human MCF-7, human Hela, rat lung, mouse lung (catalog A05072-2) |
| Gel % | 10% (catalog A05072-2) |
| Load | 30 ug; reducing conditions (catalog A05072-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A05072-2) |
| Membrane | nitrocellulose membrane (catalog A05072-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A05072-2) |
| Primary antibody | A05072-2 · 0.5 μg/mL (catalog A05072-2) |
| Primary incubation | overnight at 4°C (catalog A05072-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A05072-2) |
| Secondary incubation | 1.5 hour at RT (catalog A05072-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A05072-2) |
| Detection | ECL (catalog A05072-2) |
Section 2
What Is the Expected GPRC5A Western Blot Band Size?
GPRC5A is predicted at 40.3 kDa and observed at approximately 37–50 kDa; the cause of the migration range is not established.
| Band at approximately 37–50 kDa | Empirical GPRC5A band range; confirm identity with antibody and genetic controls |
| Band near 40.3 kDa | Consistent with the predicted GPRC5A mass |
| Band above 40.3 kDa | Asn158 N-glycosylation could affect migration; the cause requires testing |
| Weak band in whole-cell lysate | Membrane-associated GPRC5A may be poorly extracted |
💡Expected GPRC5A appearanceGPRC5A has a predicted mass of 40.3 kDa and an empirical band at approximately 37–50 kDa; the cause of any migration difference is unestablished, so confirm band identity with antibody and genetic controls.
| Predicted GPRC5A mass | 40.3 kDa is the sequence-based reference; the empirical band spans approximately 37–50 kDa |
| N-linked glycosylation at Asn158 | If occupied, the glycan may change apparent migration by an unquantified amount |
| Unoccupied Asn158 glycosylation site | Would lack any mass contribution from that site |
| Removal of the Asn158 N-glycan | Could lower apparent mass if the site is occupied; no shift is established |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Membrane-associated GPRC5A may be poorly extracted | Check membrane solubilization and probe a membrane-enriched fraction |
| Band higher than expected | Asn158 N-glycosylation is possible, but its effect on migration is unproven | Compare with an appropriate deglycosylation control and verify band identity |
| Band lower than expected | The 37–50 kDa empirical range extends below the 40.3 kDa prediction; the cause is unknown | Compare with a positive control and verify the band by GPRC5A depletion |
| Broad smear instead of sharp band | The cause is unestablished; Asn158 glycan heterogeneity is only a possibility | Check sample quality and compare treated and untreated samples if testing glycosylation |
| Multiple bands | Band identities are unestablished; no distinct isoform masses are supplied | Use GPRC5A depletion and an independent antibody to identify specific bands |
| Weak or no signal | Membrane-associated GPRC5A may be inefficiently solubilized | Optimize membrane protein extraction and include a positive control |
Experiment evidence
Western blot images showing GPRC5A band
A05072-2 · GPRC5A
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for GPRC5A Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for GPRC5A in Western blot, you can use lung tissue, which HPA rates high for expression.
Positive control: Lung (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: As a multi-pass membrane protein, GPRC5A may be easier to detect in membrane-enriched lysates.
HPA tissue expression evidence for GPRC5A
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 |
|---|---|---|---|---|
| Lung | alveolar cells | High | Protein (IHC) | HPA → |
| Caudate | neuronal cells | Medium | Protein (IHC) | HPA → |
| Colon | glandular cells | Medium | Protein (IHC) | HPA → |
| Duodenum | glandular cells | Medium | Protein (IHC) | HPA → |
| Hippocampus | neuronal 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 → |
| Adrenal gland | glandular cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | Not detected | Protein (IHC) | HPA → |
| Cervix | glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced GPRC5A Western Blot Tips
Deeper troubleshooting and optimisation questions for GPRC5A, answered from its protein features.
How should GPRC5A 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 separate GPRC5A bands represent annotated isoforms?
Isoforms · The supplied UniProt record lists one isoform and no alternative sequence. Do not assign separate bands to annotated GPRC5A isoforms on this evidence; assess each band against the expected 40.3 kDa mass and the reported 37–50 kDa range.
Could glycosylation affect GPRC5A band interpretation?
PTM · UniProt lists an N-linked glycosylation site at Asn158, using UniProt sequence numbering. Consider this site when comparing bands, but do not assign an upper band to glycosylation from position alone.
UniProt lists phosphoserine at Ser301 and Ser345, and phosphotyrosine at Tyr317, Tyr320, Tyr347 and Tyr350. These are UniProt sequence coordinates. A site's presence does not establish that phosphorylation causes a visible band shift in your sample.
UniProt lists phosphoserine at Ser301 and Ser345, and phosphotyrosine at Tyr317, Tyr320, Tyr347 and Tyr350. These are UniProt sequence coordinates. A site's presence does not establish that phosphorylation causes a visible band shift in your sample.
Does this guide establish induction of GPRC5A?
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 GPRC5A Western blot?
Transfer · GPRC5A is a multi-pass membrane protein with a predicted mass of 40.3 kDa. Check that your transfer conditions recover protein around that size, and verify transfer before interpreting a weak or absent signal.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A05072-2 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 GPRC5A localization guide quantitation?
Quantitation · UniProt places GPRC5A at the cell membrane and cytoplasmic vesicle membranes, including perinuclear vesicles. Keep the sampled fraction and extraction procedure consistent across comparisons, and normalize within that fraction.
Why might GPRC5A appear between 37 and 50 kDa?
Interpretation · Its predicted mass is 40.3 kDa, while reported Western blot bands span about 37–50 kDa. UniProt lists one N-linked glycosylation site and six phosphorylation sites, but those features alone do not establish a visible shift or explain a particular band's position.
Compare them with the predicted 40.3 kDa mass and reported 37–50 kDa range. UniProt lists one isoform, an Asn158 glycosylation site, and six phosphorylation sites, but none identifies a particular unexpected band. Its interaction with EGFR through the transmembrane domain likewise does not identify a band.
Compare them with the predicted 40.3 kDa mass and reported 37–50 kDa range. UniProt lists one isoform, an Asn158 glycosylation site, and six phosphorylation sites, but none identifies a particular unexpected band. Its interaction with EGFR through the transmembrane domain likewise does not identify a band.
Boster reagents
GPRC5A 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-RAI3/GPRC5A Antibody Picoband®
Cat # A05072-2
The catalog reports one anti-GPRC5A Western blot antibody, A05072-2, with stated human, mouse, and rat reactivity. Its WB image shows a 37–50 kDa band in human cell and rat and mouse lung lysates. Evidence is limited to the reported conditions.
Which to pick: A05072-2 is the only listed option. Its WB image includes human A375, MCF-7, and HeLa cells, plus rat and mouse lung lysates, with 30 µg loaded per lane under reducing conditions.
Source: BosterBio GPRC5A gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q8NFJ5.
- Human Protein Atlas. GPRC5A tissue expression.
- PMC9870083 — target-verified WB comparison
- PMC5335824 — target-verified WB comparison
- PMC5355118 — target-verified WB comparison