Phospho-SYN1 (S62) Antibody
Description
Overview
The Phospho-SYN1 (S62) Antibody is a polyclonal antibody specifically designed to detect phosphorylation at serine residue 62 (S62) of Synapsin I (SYN1), a critical presynaptic protein involved in neurotransmitter release and synaptic plasticity . SYN1 phosphorylation at S62 is associated with activity-dependent regulation of synaptic function, particularly in modulating vesicle clustering and release .
Mechanism and Significance
Synapsin I is a neuron-specific phosphoprotein that binds to synaptic vesicles and actin filaments, playing a dual role in stabilizing vesicle pools and facilitating calcium-dependent release . Phosphorylation at S62, mediated by calcium-calmodulin-dependent protein kinase II (CaMKII), enhances Synapsin I’s ability to interact with actin and modulate synaptic activity . This post-translational modification is critical for short-term synaptic plasticity and memory formation .
Validation and Research Applications
Western Blot:
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Abnova’s PAB9659 demonstrates specificity by blocking immunolabeling with the phospho-peptide (rat cortex lysate) .
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Boster Bio’s A03794S62 validates phosphorylation in HEK293T lysates treated with PMA (100nM) .
Immunohistochemistry (IHC):
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Cephamls’ antibody shows localization in human brain tissue (e.g., cerebral cortex) under physiological conditions .
Functional Studies:
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Kao et al. (2002) demonstrated that phosphorylation at S62 enhances Synapsin I’s role in neurite outgrowth and synaptic plasticity .
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Jovanovic et al. (2001) linked S62 phosphorylation to Ca2+-dependent glutamate release in rat cerebrocortical synaptosomes .
Clinical and Therapeutic Implications
Phosphorylation at S62 has been implicated in:
Product Specs
Target Background
- Researchers have identified a c.236 C > G/p.S79W mutation in SYN1 as the cause of non-syndromic intellectual disability in the MRX50 family. In vitro studies of the S79W SynI mutation show that it doesn't affect neurodevelopmental aspects, but it disrupts spontaneous synaptic vesicle (SV) exocytosis, SV clustering, and SV lateral mobility along axons. PMID: 28973667
- These findings complement previous research demonstrating dysregulation of Synapsins, particularly SYN2, in mood disorders. They enhance our understanding of the regulatory mechanisms that contribute to these changes, potentially leading to bipolar disorder (BD) or major depressive disorder (MDD) phenotypes. PMID: 27515700
- Cerebral malaria induces pre-synaptic excitation and ultimately activates synapsin I, resulting in increased neurotransmitter release. PMID: 26823711
- Immunoreactivity patterns with antibodies against SNAP-25, synapsin-I, and synaptophysin align with those observed in the olfactory bulb (OB) of adults at 38-40 weeks of prenatal development. PMID: 26204769
- The development of the AlphaScreen pSYN1 assay and future creation of additional primary neuronal high-throughput screening (HTS) assays provide a promising approach for identifying novel classes of therapeutic candidates for various central nervous system (CNS) disorders. PMID: 24088370
- These findings suggest that PRICKLE1 mutations contribute to autism spectrum disorder (ASD) by disrupting the interaction with SYN1 and the regulation of synaptic vesicles. PMID: 24312498
- Data indicate that in patients carrying the W356x mutation, synapsin I function is significantly impaired, supporting the value of Syn1(-/-) mice as an experimental model mimicking the human pathology. PMID: 23818987
- The epileptogenic Q555X SYN1 mutant triggers imbalances in release dynamics and short-term plasticity. PMID: 23406870
- Histone modification marks were significantly elevated in major depression, and this effect correlated with substantial increases in SYN1b gene expression. PMID: 22571925
- The allelic frequencies of SYN1 are associated with schizophrenia in Korean women. PMID: 22807112
- Loss-of-function mutations in SYN1 have been linked to autism and partial epilepsy, leading to impaired synaptic function. PMID: 21441247
- Nucleocytoplasmic shuttling of dysbindin-1 regulates synapsin I expression and may therefore contribute to the pathogenesis of schizophrenia. PMID: 20921223
- The authors propose that claudin-2 and SYN1 work together to enhance microbial translocation across the intestinal epithelial barrier, contributing to chronic immune activation and CD4 T-cell depletion in HIV-1-infected patients. PMID: 20700059
- Research has shown that synapsin I is significantly decreased in the stratum radiatum of the CA1 subfield and the molecular layer of the dentate gyrus (DG) in Alzheimer's disease (AD) patients. PMID: 14673601
- A SYN1 nonsense mutation is likely responsible for epileptic and other phenotypes. PMID: 14985377
- Synapsins and S100A1 interact in nerve terminals where they are co-expressed. S100A1 cannot bind SV-associated synapsin I and may function as a cytoplasmic store of monomeric synapsin I. Synapsin dimerization and interaction with S100A1 are mutually exclusive. PMID: 15147519
- This study concluded that the human synapsin I gene is positively regulated by nuclear respiratory factor 1 and mediates its function in neurite outgrowth. PMID: 19301426
Q&A
What is Phospho-SYN1 (S62) Antibody and what does it detect?
Phospho-SYN1 (S62) Antibody is a polyclonal antibody that specifically detects endogenous levels of Synapsin I protein when phosphorylated at Serine 62 (Ser62) . Synapsin I is a neuronal phosphoprotein that associates with the cytoplasmic surface of synaptic vesicles and plays critical roles in neurotransmission . The antibody is typically raised in rabbits against synthetic phosphopeptides derived from human Synapsin I around the phosphorylation site of Serine 62 . Some antibodies are designed to detect phosphorylation at both Ser62 and Ser67 sites .
When used in Western blot analysis, this antibody typically detects a protein band at approximately 80 kDa, representing phosphorylated Synapsin I . The antibody is manufactured for research use only and not for diagnostic procedures .
What is the role of Synapsin I in neuronal function?
Synapsin I is a member of the synapsin family of neuronal phosphoproteins that associate with the cytoplasmic surface of synaptic vesicles . This protein plays crucial roles in:
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Serving as a substrate for several different protein kinases, with phosphorylation functioning in the regulation of this protein in nerve terminals
Synapsins are characterized by common protein domains and are implicated in synaptogenesis and the modulation of neurotransmitter release, suggesting potential roles in several neuropsychiatric diseases . Mutations in the Synapsin I gene may be associated with X-linked disorders with primary neuronal degeneration such as Rett syndrome .
What are the optimal storage conditions for Phospho-SYN1 (S62) Antibody?
For maximum stability and activity retention, Phospho-SYN1 (S62) Antibody should be stored according to the following guidelines:
| Storage Term | Temperature | Additional Recommendations |
|---|---|---|
| Short-term | 4°C | In original container |
| Long-term | -20°C | Aliquot to avoid repeated freeze-thaw cycles |
Most manufacturers supply the antibody in a buffer containing PBS with additives such as sodium azide, glycerol (typically 50%), and sometimes BSA to maintain stability . It is critical to avoid repeated freeze-thaw cycles as they can significantly degrade antibody quality and performance .
What are the typical applications and dilutions for Phospho-SYN1 (S62) Antibody?
The primary applications and recommended dilutions for Phospho-SYN1 (S62) Antibody include:
| Application | Recommended Dilution | Notes |
|---|---|---|
| Western Blot (WB) | 1:500 - 1:2000 | Most common application |
| Immunohistochemistry (IHC) | 1:100 - 1:300 | For tissue section analysis |
| ELISA | 1:10000 | For quantitative detection |
The optimal working dilution should be determined by each researcher through titration experiments as it may vary depending on sample type and experimental conditions . Western blot analysis is the most validated application across multiple manufacturers .
What is the specificity of Phospho-SYN1 (S62) Antibody?
The specificity of Phospho-SYN1 (S62) antibody is characterized by its ability to detect Synapsin I only when phosphorylated at Serine 62 . In Western blot applications, the antibody typically detects a doublet band at approximately 80 kDa corresponding to phosphorylated Synapsin I .
Specificity can be validated experimentally through:
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Blocking experiments using the phosphopeptide immunogen (should block signal)
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Testing with the corresponding dephosphopeptide (should not block signal)
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Treatment of samples with PMA (phorbol 12-myristate 13-acetate), which induces phosphorylation at this site
According to manufacturer testing, the antibody has been shown to work with samples from multiple species including human, mouse, and rat .
How does phosphorylation at Ser62 regulate Synapsin I function?
Phosphorylation at Ser62 is a key regulatory mechanism that modulates Synapsin I function in synaptic vesicle dynamics. While the search results don't provide explicit details about the specific effects of Ser62 phosphorylation, we can infer its importance based on several factors:
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The presence of specific antibodies targeting this phosphorylation site indicates its biological significance
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Treatment with PMA (100nM, 30min) induces phosphorylation at this site, suggesting it's regulated by protein kinase C or related signaling pathways
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The phosphorylation likely affects Synapsin I's interactions with synaptic vesicles and/or cytoskeletal elements
Similar to other phosphorylation events, such as c-Myc phosphorylation at Ser62 (which serves as a priming site for subsequent phosphorylation events), Synapsin I phosphorylation may participate in sequential phosphorylation cascades that regulate protein function and stability .
What controls should be included when validating Phospho-SYN1 (S62) Antibody for experimental use?
When validating Phospho-SYN1 (S62) Antibody for experimental use, researchers should include the following controls:
Drawing from learnings with other phospho-specific antibodies like those for phospho-α-synuclein, researchers should be cautious about non-specific binding and diffuse neuropil labeling that may occur with some antibodies . Titration experiments are essential to determine optimal concentrations that maximize specific signals while minimizing background .
How can researchers troubleshoot poor signal or high background when using Phospho-SYN1 (S62) Antibody?
When encountering issues with Phospho-SYN1 (S62) Antibody, consider these troubleshooting approaches:
For Poor Signal:
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Phosphorylation state preservation: Use phosphatase inhibitors in all buffers during sample preparation
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Protein degradation: Add protease inhibitors to all extraction buffers
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Antibody concentration: Try higher concentrations (within manufacturer's recommended range)
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Exposure time: Increase for Western blot detection
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Sample treatment: Consider using PMA or other treatments known to increase Ser62 phosphorylation
For High Background:
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Blocking optimization: Test different blocking agents (BSA, milk, commercial blockers)
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Antibody dilution: Use more diluted antibody solutions
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Washing steps: Increase number and duration of washes
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Secondary antibody: Ensure appropriate dilution and specificity
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Sample preparation: Optimize extraction and purification protocols to reduce interfering components
Experience with other phospho-specific antibodies suggests that some clones may produce non-specific diffuse neuropil labeling, which should be controlled for with appropriate negative controls .
How can Phospho-SYN1 (S62) Antibody be used to investigate neurological disorders?
Phospho-SYN1 (S62) Antibody can be a valuable tool for investigating neurological disorders, particularly those with synaptic dysfunction components:
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Rett Syndrome: Since mutations in Synapsin I may be associated with X-linked disorders including Rett syndrome , the antibody can be used to:
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Compare phosphorylation levels between patient and control samples
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Evaluate effects of therapeutic interventions on Synapsin I phosphorylation
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Study disease progression by monitoring phosphorylation changes over time
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Other Neurodevelopmental Disorders:
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Analyze phosphorylation patterns in animal models of disorders
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Correlate phosphorylation status with electrophysiological measures of synaptic function
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Examine effects of genetic mutations on signaling pathways regulating Synapsin I phosphorylation
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Methodological Approach:
When investigating disorders, researchers should pay particular attention to sample preparation to preserve phosphorylation status, as postmortem interval and tissue processing can significantly affect phosphoprotein detection.
What are the differences between antibodies that detect Phospho-SYN1 (S62) versus those that detect Phospho-SYN1 (S62/S67)?
Researchers should be aware of the distinctions between antibodies targeting single versus multiple phosphorylation sites:
When choosing between these antibodies, researchers should consider:
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The specific phosphorylation site(s) relevant to their research question
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Whether single or multiple phosphorylation events are being investigated
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The need to distinguish between phosphorylation at different sites
Similar to considerations when selecting phospho-α-synuclein antibodies, researchers should be aware that different clones may have varying sensitivities and specificities for detecting phosphorylated epitopes .
How can Phospho-SYN1 (S62) Antibody be integrated with other research techniques for comprehensive neuronal signaling studies?
Integrating Phospho-SYN1 (S62) Antibody with complementary techniques provides a more comprehensive understanding of neuronal signaling:
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Combination with Electrophysiology:
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Correlate phosphorylation levels with synaptic strength measurements
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Examine the temporal relationship between stimulation protocols and phosphorylation changes
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Investigate how phosphorylation affects synaptic plasticity mechanisms
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Integration with Imaging Techniques:
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Use immunofluorescence with Phospho-SYN1 (S62) Antibody alongside synaptic markers
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Employ super-resolution microscopy to localize phosphorylated Synapsin I within synapses
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Combine with live imaging of calcium dynamics to correlate activity with phosphorylation
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Complementary Molecular Approaches:
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Pair with kinase activity assays to identify enzymes responsible for Ser62 phosphorylation
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Use with phospho-proteomic analyses to map comprehensive phosphorylation networks
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Combine with proximity ligation assays to detect protein interactions specific to phosphorylated Synapsin I
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Functional Studies:
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Correlate phosphorylation with neurotransmitter release measurements
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Assess impact of phosphorylation site mutations on synaptic function
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Study activity-dependent changes in phosphorylation state
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This multi-technique approach allows researchers to connect biochemical modifications with functional outcomes at the synapse, providing insight into the regulatory mechanisms controlling synaptic transmission.
What are the key considerations when comparing results across different lots or manufacturers of Phospho-SYN1 (S62) Antibody?
When comparing results obtained with different lots or manufacturers of Phospho-SYN1 (S62) Antibody, researchers should consider:
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Antibody Production Variables:
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Standardization Approaches:
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Technical Harmonization:
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Maintain consistent sample preparation protocols
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Standardize dilution ratios based on antibody concentration rather than dilution factor
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Use consistent detection methods and exposure settings
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Documentation and Reporting:
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Record antibody catalog numbers, lot numbers, and dilutions used
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Document detailed experimental protocols for reproducibility
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Note any deviations from manufacturer recommendations
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Drawing from practices in phospho-specific antibody validation, researchers should consider validating each new lot with blocking peptides and comparing performance across multiple sample types, as demonstrated with phospho-α-synuclein antibodies .
