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GABRB3 / Gamma-aminobutyric acid receptor subunit beta-3 · Western blot design guide
Design a Western Blot for GABRB3
Source-linked GABRB3 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-GABRB3 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
GABRB3 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 | ~54.1 kDa | |
| Observed band | ≈54 kDa | |
| Gel | 5–20% (catalog PB9595) | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Glycosylated + Cleaved | |
| Caveat | Glycosylation-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 4 isoform(s) |
Section 1
Source-Linked GABRB3 Western Blot Protocol Options
The PB9595 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | Rat Brain at 50ug, Mouse Brain at 50ug (catalog PB9595) |
| Gel % | 5–20% (catalog PB9595) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog PB9595) |
| Membrane | nitrocellulose membrane (catalog PB9595) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog PB9595) |
| Primary antibody | PB9595 · 0.5 μg/mL (catalog PB9595) |
| Primary incubation | overnight at 4°C (catalog PB9595) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog PB9595) |
| Secondary incubation | 1.5 hour at RT (catalog PB9595) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog PB9595) |
| Detection | ECL (catalog PB9595) |
Section 2
What Is the Expected GABRB3 Western Blot Band Size?
GABRB3 is predicted at 54.1 kDa and observed at approximately 54 kDa; the supplied features establish no additional migration shift.
| Band at approximately 54 kDa | Consistent with the observed GABRB3 band in brain lysate. |
| Band below approximately 54 kDa | Could reflect signal-peptide cleavage; identity needs confirmation. |
| Band above approximately 54 kDa | Could reflect N-linked glycosylation, though a visible shift is not established. |
| Several bands near 54 kDa | Could include isoforms 1, 2, 3, and 4; distinct migration is not established. |
| Weak or absent band in whole-cell lysate | May reflect limited recovery of this membrane protein. |
💡Expected GABRB3 appearanceThe predicted precursor mass is 54.1 kDa, and brain lysate blots show a band at approximately 54 kDa; confirm its identity with an independent antibody or knockdown.
| Predicted precursor mass | Provides a 54.1 kDa reference; the observed band is approximately 54 kDa. |
| N-linked glycosylation at Asn33 | Could affect apparent size; no site-specific shift is established. |
| N-linked glycosylation at Asn105 | Could affect apparent size; no site-specific shift is established. |
| N-linked glycosylation at Asn174 | Could affect apparent size; no site-specific shift is established. |
| Signal peptide at residues 1–25 | Cleavage makes the mature chain smaller than the precursor; its migration is not supplied. |
| Isoforms 1, 2, 3, and 4 | May differ in size, but individual masses and distinct bands are not established. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Membrane localization may limit recovery in whole-cell lysate. | Check membrane solubilization and use brain or membrane-enriched material as a positive control. |
| Band higher than expected | N-linked glycosylation is possible, but its contribution to migration is unproven. | Compare treated and untreated samples by deglycosylation and confirm band identity. |
| Band lower than expected | Signal-peptide cleavage is possible; the mature band's size is not supplied. | Check antibody epitope coverage and confirm identity with an independent antibody or knockdown. |
| Broad smear instead of sharp band | Heterogeneous N-linked glycosylation is possible but not demonstrated. | Compare deglycosylated material and check sample quality. |
| Multiple bands | Isoforms 1, 2, 3, and 4 exist, but distinct migration is not established. | Use an independent antibody or knockdown to identify GABRB3 bands. |
| Weak or no signal | Membrane protein extraction may be incomplete. | Check solubilization, loading, and a brain lysate positive control. |
Experiment evidence
Western blot images showing GABRB3 band
PB9595 · GABRB3
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for GABRB3 Western blot
🧪For positive controls for GABRB3 in Western blot, you can use a validated GABRB3-positive sample; the supplied HPA data identify no specific 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 alongside the samples.
⚠️Feasibility: GABRB3 is a multi-pass membrane protein, and the supplied HPA data identify no tissue controls.
HPA tissue expression evidence for GABRB3
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 GABRB3 Western Blot Tips
Deeper troubleshooting and optimisation questions for GABRB3, answered from its protein features.
Does the signal peptide change the expected GABRB3 band?
Band shift · The canonical sequence has a signal peptide at residues 1–25. Its annotation makes processing relevant to band interpretation, but the supplied features do not establish the processed protein’s apparent mass. Use canonical UniProt numbering when referring to residues downstream of the signal peptide.
Could GABRB3 isoforms produce different bands?
Isoforms · UniProt lists four isoforms. Relative to the canonical sequence, isoform 2 replaces residues 1–26, isoform 3 replaces residues 3–80, and isoform 4 lacks residues 1–85. These sequence differences could affect migration or antibody recognition; a band pattern alone cannot identify an isoform.
How should GABRB3 glycosylation affect band interpretation?
PTM · UniProt annotates N-linked glycosylation at canonical residues 33, 105, and 174. Consider glycosylation when interpreting band positions, but site annotations do not demonstrate that a particular band is glycosylated or predict its apparent shift. State the sequence convention when comparing these coordinates with antibody or paper numbering.
Does this guide establish induction of GABRB3?
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 GABRB3 Western blot?
Transfer · GABRB3 is a multi-pass membrane protein with a predicted mass of 54.1 kDa. The supplied features do not specify a transfer method. Use a method validated for membrane proteins and check transfer around 54 kDa before interpreting a weak signal.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the PB9595 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 GABRB3 bands be quantified?
Quantitation · Quantify a consistently defined band across samples and document whether additional bands are included. The supplied observed band is near 54 kDa, while four isoforms and three canonical N-linked glycosylation sites could complicate band assignment. Do not pool distinct bands without evidence that they represent GABRB3.
Why is the observed GABRB3 band near 54 kDa?
Interpretation · The observed band near 54 kDa is close to the supplied 54.1-kDa predicted mass. UniProt lists a signal peptide at residues 1–25 and N-linked glycosylation sites at 33, 105, and 174, but those features alone do not establish a visible shift or explain any small mass difference.
Check whether the pattern is consistent with the four annotated isoforms, the canonical signal peptide at 1–25, or N-linked glycosylation sites at 33, 105, and 174. These features suggest possibilities, but none identifies an unexpected band by itself. Confirm band identity before assigning a cause.
Check whether the pattern is consistent with the four annotated isoforms, the canonical signal peptide at 1–25, or N-linked glycosylation sites at 33, 105, and 174. These features suggest possibilities, but none identifies an unexpected band by itself. Confirm band identity before assigning a cause.
Boster reagents
GABRB3 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-GABA A Receptor beta 3/GABRB3 Antibody Picoband®
Cat # PB9595
PB9595 is a rabbit polyclonal anti-GABRB3 antibody listed for human, mouse, and rat. Its Western blot image shows an approximately 54 kDa band in rat and mouse brain tissue lysates. The supplied evidence does not show a human Western blot.
Which to pick: PB9595 is the only listed option and has a Western blot image using 50 µg each of rat and mouse brain tissue lysate. For human samples, reactivity is listed, but no human Western blot example is supplied.
Source: BosterBio GABRB3 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P28472.
- Human Protein Atlas. GABRB3 tissue expression.
- PMC12181925 — target-verified WB comparison
- PMC2924885 — target-verified WB comparison
- PMC8179828 — target-verified WB comparison