PLC1 Antibody
Description
Definition and Biological Significance
PLCγ1 is a 148–155 kDa enzyme encoded by the PLCG1 gene, which hydrolyzes phosphatidylinositol 4,5-bisphosphate (PIP₂) into secondary messengers (IP₃ and DAG) to regulate calcium signaling, cell proliferation, and immune responses . Antibodies targeting PLCγ1 are essential for investigating its role in:
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T cell receptor (TCR) signaling: PLCγ1 promotes phase separation of LAT adaptor proteins, enhancing signal transduction .
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Disease mechanisms: Dysregulation of PLCγ1 is linked to cancer metastasis, autoimmune disorders, and neurological conditions .
Role in Immune Signaling
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Phase separation mechanism: PLCγ1 cross-links LAT proteins via its SH2 domains, stabilizing microclusters that amplify TCR signaling . At concentrations as low as 5 nM, PLCγ1 enhances LAT recruitment of Sos1, critical for ERK activation .
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Regulation of phosphorylation: PLCγ1 protects LAT from CD45-mediated dephosphorylation, ensuring sustained signaling .
Disease Associations
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Cancer: Elevated PLCγ1 activity correlates with tumor progression and metastasis. Antibodies like #5690 (Cell Signaling Tech) are used to detect PLCγ1 in cancer cell lines (e.g., HeLa, HepG2) .
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Autoimmunity: PLCγ1 inhibition reduces aberrant T cell activation, suggesting therapeutic potential .
Validation and Specificity
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Cross-reactivity: Most antibodies (e.g., MAB3288 , #2822 ) are validated for human, mouse, and rat samples.
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Specificity: CAB21545 and CG1461 target amino acids 1202–1291 and synthetic peptides, respectively, minimizing cross-reactivity with other PLC isoforms.
Recommended Protocols
Applications in Therapeutic Development
PLCγ1 antibodies are pivotal in preclinical studies:
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Drug discovery: Used to screen inhibitors targeting PLCγ1’s SH2 domains, which disrupt oncogenic signaling .
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Biomarker analysis: Detect PLCγ1 overexpression in tumor biopsies, aiding patient stratification .
Limitations and Future Directions
Product Specs
**Constituents:** 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
- Aluminum-induced alterations in plasma membrane physical properties can impact the activity of non-specific phospholipase C. PMID: 26024014
Q&A
Here’s a structured FAQ collection for researchers working with PLCγ1 antibodies, organized by complexity and grounded in experimental methodology:
Advanced Research Questions
How do I resolve discrepancies in PLCγ1 detection across experimental models?
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Analysis framework:
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Species reactivity: Confirm cross-reactivity (e.g., rabbit monoclonal M00677 works for human/rat ; clone B-6-4 reacts with mouse/rabbit ).
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Post-translational modifications: PLCγ1 is tyrosine-phosphorylated during signaling . Use phospho-specific antibodies or treat lysates with phosphatases.
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Subcellular localization: Fractionate lysates (cytosolic vs. membrane) to account for activation-dependent translocation.
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What strategies enhance PLCγ1 signal in low-abundance samples?
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Optimization steps:
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Signal amplification: Use high-affinity antibodies (e.g., serplulimab-like engineering ) or tyramide-based systems.
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Multiplex assays: Pair PLCγ1 detection with co-stimulatory markers (e.g., CD28, AKT ) using fluorescent secondary antibodies.
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Pre-absorption controls: Pre-incubate antibody with immunogen peptide to confirm target specificity .
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| Challenge | Solution | Source |
|---|---|---|
| Low protein yield | Load ≥30 µg lysate; extend transfer time to 90 min | |
| Non-specific bands | Include 0.1% BSA in blocking buffer; reduce primary antibody concentration |
Technical Troubleshooting
How to interpret PLCγ1 knockdown phenotypes in signaling studies?
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Validation pipeline:
Can PLCγ1 antibodies distinguish isoforms or splice variants?
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Approach:
