SLC18A2 Antibody

What is SLC18A2 and why is it important in research?

SLC18A2 (Solute Carrier Family 18 Member A2), also known as VMAT2 (Vesicular Monoamine Transporter 2), is a transmembrane protein with significant roles in neuronal and neuroendocrine systems. The canonical human protein consists of 514 amino acid residues with a molecular weight of approximately 55.7 kDa . SLC18A2 functions as an electrogenic antiporter that exchanges cationic monoamines with intravesicular protons across secretory and synaptic vesicle membranes . This protein is crucial for neurotransmission, as it facilitates the uptake of monoamines into cytoplasmic large dense core and synaptic vesicles in neuronal cells, being particularly responsible for catecholamine and serotonin storage in central synapses . Additionally, SLC18A2 has emerged as a potential biomarker in prostate cancer diagnosis and prognosis, where its hypermethylation and downregulation correlate with disease outcomes .

What are the key structural and functional characteristics of SLC18A2 protein?

SLC18A2 is primarily localized in cytoplasmic vesicles and exists in up to two different isoforms . As a member of the Vesicular transporter protein family, it functions through an electrogenic antiporter mechanism, exchanging one cationic monoamine with two intravesicular protons across vesicular membranes . The protein undergoes post-translational modifications, notably glycosylation, which may influence its functionality . SLC18A2 is predominantly expressed in neuronal and neuroendocrine tissues, where it plays a critical role in monoamine storage and neurotransmission . The protein has a molecular weight of approximately 60 kDa as detected by Western blotting techniques . SLC18A2 has been used as a marker for both Tissue Resident Mucosal Type Mast Cells and Connective Tissue Type Mast Cells in human tissue studies .

What are the common synonyms and orthologs for SLC18A2?

Researchers should be aware of various synonyms when searching literature and resources related to SLC18A2. Common alternative designations include SVAT, SVMT, VAT2, VMAT2, synaptic vesicular amine transporter, and PKDYS2 . This awareness is crucial for comprehensive literature searches and avoiding redundancy in research. SLC18A2 gene orthologs have been identified across multiple species, including mouse, rat, bovine, frog, zebrafish, chimpanzee, and chicken, making it a conserved protein of interest in comparative biology studies . When studying SLC18A2 across species, researchers should note potential structural and functional variations that might affect antibody binding specificity and experimental interpretations.

What are the optimal applications for SLC18A2 antibodies in research?

SLC18A2 antibodies demonstrate versatility across multiple experimental applications. Based on commercial antibody specifications, researchers can effectively employ these antibodies for Western Blot (WB), Immunohistochemistry (IHC), Flow Cytometry (FCM), and Immunofluorescence (IF) techniques . For immunocytochemistry (ICC) and Western blotting, monoclonal antibodies like N449/73 have been specifically validated and purified through Protein A chromatography techniques . When designing experiments, researchers should consider that different antibody clones may have varying optimal applications. For instance, the N449/73 clone has been specifically validated for ICC and WB applications with human, mouse, and rat samples . For detecting cellular localization patterns, immunohistochemistry studies have revealed distinct staining patterns in prostate tissues, including both diffuse and apical cytoplasmic staining patterns that may reflect different cellular activities such as vesicular storage versus active secretion of monoamines .

How should researchers select the appropriate SLC18A2 antibody for cross-species studies?

The selection of SLC18A2 antibodies for cross-species studies requires careful consideration of reactivity profiles. Commercial antibodies demonstrate varying cross-reactivity with SLC18A2 from different species. For instance, some antibodies show reactivity across human, mouse, rat, dog, and monkey samples, while others have broader reactivity profiles including rabbit, bovine, guinea pig, and hamster SLC18A2 . When conducting cross-species research, investigators should:

• Verify species reactivity information provided by manufacturers

• Perform validation tests on target species tissues before proceeding with experiments

• Consider epitope conservation across species when selecting antibody clones

• Be aware that antibodies targeting highly conserved regions (like the first lumenal domain) may provide better cross-species reactivity

For example, a monoclonal antibody derived from fusion protein amino acids 42-132 (first lumenal domain) of mouse SLC18A2 has been validated to work effectively with human, mouse, and rat samples . Importantly, researchers should verify whether the antibody cross-reacts with related proteins like SLC18A1/VMAT1 or SLC18A3/VAChT, as some antibodies (like N449/73) have been specifically tested to confirm they do not cross-react with these related transporters .

What methodologies are recommended for analyzing SLC18A2 expression at different molecular levels?

A comprehensive analysis of SLC18A2 requires examination at multiple molecular levels:

DNA Methylation Analysis:

• Quantitative methylation-specific PCR (qMSP) can effectively assess SLC18A2 promoter methylation

• Illumina 450K methylation microarray provides high-throughput methylation profiling

• These techniques have successfully identified hypermethylation of the SLC18A2 promoter-associated CpG island in prostate cancer

RNA Expression Analysis:

• RNA sequencing (RNAseq) has been successfully employed to assess SLC18A2 transcript levels

• This approach has identified reduced SLC18A2 mRNA expression in cancerous versus normal tissues

Protein Expression Analysis:

• Immunohistochemistry on tissue microarrays (TMAs) allows for high-throughput protein analysis

• Scoring systems differentiating between diffuse and apical cytoplasmic staining patterns provide insights into potential functional differences

• Western blotting can confirm protein molecular weight (approximately 60 kDa)

For optimal results, researchers should implement laser capture microdissection (LMD) to isolate specific cell populations before analysis, as this approach enhances specificity of findings related to SLC18A2 expression in complex tissues .

How can SLC18A2 antibodies be utilized in studying neurodegenerative disorders?

SLC18A2 antibodies offer valuable insights into neurodegenerative disorders due to the protein's critical role in monoamine neurotransmission. Researchers have effectively employed these antibodies to monitor VMAT2 expression during development, aging, and neurodegeneration . As markers for monoamine terminals, SLC18A2 antibodies allow researchers to:

• Track changes in dopaminergic, serotonergic, and noradrenergic systems across disease progression

• Visualize and quantify alterations in synaptic vesicle populations

• Assess the integrity of monoaminergic circuits in models of neurodegeneration

• Monitor therapeutic responses affecting vesicular monoamine transport

For these applications, immunohistochemistry and immunofluorescence techniques are particularly valuable, allowing visualization of SLC18A2 distribution within specific neuronal populations. When designing such studies, researchers should consider region-specific expression patterns and potential alterations in subcellular localization that might accompany pathological states.

What is the significance of SLC18A2 in prostate cancer research and how should antibodies be applied in this context?

SLC18A2 has emerged as a promising biomarker in prostate cancer research with significant diagnostic and prognostic potential. Large-scale evaluations have revealed that SLC18A2 is frequently hypermethylated and downregulated in prostate cancer tissues . The application of SLC18A2 antibodies in this context requires specialized methodological considerations:

Diagnostic Applications:

• Immunohistochemical staining of prostate tissue sections shows reduced SLC18A2 protein in cancerous versus benign tissues (AUC 0.898)

• Different staining patterns (diffuse versus apical cytoplasmic) should be evaluated separately, as both show high diagnostic accuracy (AUCs 0.870 and 0.832, respectively)

Prognostic Applications:

Methodological Considerations:

• Tissue microarray (TMA) construction with adequate representation of both tumor and adjacent normal tissues

• Implementation of standardized scoring systems (0: no/weak staining, 1: moderate staining, 2: strong staining)

• Distinction between predominantly apical versus predominantly diffuse cytoplasmic staining patterns

• Correlation with clinical outcomes using appropriate statistical analyses (multivariate Cox regression models)

How can researchers integrate SLC18A2 antibody staining with other molecular analyses for comprehensive biomarker studies?

A multi-omics approach incorporating SLC18A2 antibody staining with other molecular analyses provides the most comprehensive understanding of its biological significance. Based on research strategies employed in prostate cancer studies, researchers should consider:

What are common challenges in SLC18A2 antibody staining and how can they be addressed?

Researchers working with SLC18A2 antibodies may encounter several technical challenges that require specific troubleshooting approaches:

Variable Staining Patterns:

• The observation of both diffuse and apical cytoplasmic staining patterns might complicate interpretation

• Solution: Score these patterns separately and analyze their diagnostic/prognostic value independently

• Consider that different patterns may reflect distinct biological processes (vesicular storage versus active secretion)

Specificity Concerns:

• Cross-reactivity with related transporters like SLC18A1/VMAT1 or SLC18A3/VAChT may occur

• Solution: Select validated antibodies specifically tested for absence of cross-reactivity with these proteins

• Include appropriate positive and negative control tissues in each experiment

Optimization Requirements:

• Different applications (WB, IHC, IF) may require distinct optimization protocols

• Solution: Perform titration experiments to determine optimal antibody concentration

• For IHC, optimize antigen retrieval methods (heat-induced versus enzymatic) based on fixation conditions

Subcellular Localization Variability:

• SLC18A2 localization may vary depending on cell type and physiological state

• Solution: Include detailed documentation of subcellular distribution patterns

• Consider co-localization studies with vesicular markers for confirmation of proper localization

How should researchers interpret conflicting SLC18A2 data across different molecular levels?

Multi-level molecular analyses of SLC18A2 may occasionally yield seemingly contradictory results. Based on prostate cancer research findings, the following interpretation framework is recommended:

• Methylation-Expression Discrepancies:

  • If hypermethylation is observed without corresponding decrease in expression

  • Consider: Additional regulatory mechanisms, compensatory pathways, or technical limitations

  • Action: Verify methylation status at specific CpG sites near transcription factor binding regions

• Transcript-Protein Inconsistencies:

  • If reduced RNA levels are not reflected in protein abundance

  • Consider: Post-transcriptional regulation, protein stability differences, or antibody sensitivity issues

  • Action: Employ multiple antibodies targeting different epitopes and quantitative protein measurement techniques

• Heterogeneous Tissue Responses:

  • Variable SLC18A2 expression within the same specimen

  • Consider: Cellular heterogeneity, microenvironmental influences, or clonal evolution

  • Action: Implement laser capture microdissection to isolate specific cell populations before analysis

• Cross-Species Variations:

  • Different expression patterns or antibody reactivity between species

  • Consider: Evolutionary differences in protein structure or function

  • Action: Verify epitope conservation and use species-specific positive controls

What statistical approaches are recommended for analyzing SLC18A2 immunohistochemistry data in biomarker studies?

Based on established methodologies in SLC18A2 biomarker research, the following statistical approaches are recommended:

For Diagnostic Performance Assessment:

• Receiver Operating Characteristic (ROC) analysis to determine Area Under the Curve (AUC) values

• Mann-Whitney U tests to compare expression levels between malignant and benign tissues

• Calculation of sensitivity, specificity, positive and negative predictive values at optimal cutoff points

For Prognostic Evaluation:

• Kaplan-Meier survival analyses with log-rank tests for initial survival comparisons

• Univariate Cox regression to calculate hazard ratios for outcomes like biochemical recurrence

• Multivariate Cox regression including standard clinicopathological parameters to determine independent prognostic value

For Multi-level Molecular Correlation:

• Spearman or Pearson correlation coefficients to assess relationships between methylation, RNA, and protein levels

• Multiple regression models to identify primary determinants of protein expression

• Hierarchical clustering to identify patient subgroups with distinct molecular profiles

For Scoring System Validation:

• Inter-observer agreement statistics (kappa values) to ensure reproducibility of staining interpretation

• Intra-class correlation coefficients for continuous scoring data

• Sensitivity analyses using different scoring thresholds to ensure robustness of findings

What are the latest findings regarding SLC18A2 as a biomarker in prostate cancer?

Recent large-scale evaluations have revealed significant potential for SLC18A2 as both a diagnostic and prognostic biomarker in prostate cancer:

Diagnostic Biomarker Potential:

• SLC18A2 promoter hypermethylation demonstrates high cancer-specificity (AUC: 0.923–0.976)

• Protein expression is substantially reduced in prostate cancer tissues compared to benign prostate tissue (AUC 0.898)

• Both diffuse and apical cytoplasmic staining patterns show high diagnostic accuracy (AUCs 0.870 and 0.832, respectively)

Prognostic Biomarker Potential:

These findings suggest that SLC18A2-based molecular tests could have valuable future applications for prostate cancer detection and identification of high-risk patients who might benefit from more aggressive treatment approaches .

How can researchers design validation studies for SLC18A2 as a biomarker?

To effectively validate SLC18A2 as a biomarker, researchers should consider the following design elements derived from successful previous studies:

Multi-cohort Validation Approach:

• Include multiple independent patient cohorts with diverse clinical characteristics

• Analyze fresh-frozen (FF) and formalin-fixed paraffin-embedded (FFPE) tissues to ensure methodology translates across preservation methods

• Incorporate samples from different institutions to account for technical variations in tissue processing

Multi-level Molecular Assessment:

• Evaluate SLC18A2 at DNA (methylation), RNA, and protein levels

• Use complementary technologies (e.g., qMSP, Illumina 450K arrays, RNAseq, IHC)

• Correlate findings across molecular levels to establish mechanistic understanding

Comprehensive Clinical Annotation:

Statistical Validation Framework:

• Pre-specify primary and secondary endpoints

• Perform power calculations to ensure adequate sample sizes

• Include multivariate analyses with established prognostic factors to demonstrate independent prognostic value

• Consider competing risk analyses for more accurate prognostication

What emerging technologies might enhance SLC18A2 antibody applications in research?

Several innovative technologies hold promise for advancing SLC18A2 antibody applications in both basic research and clinical contexts:

Single-cell Antibody-based Technologies:

• Mass cytometry (CyTOF) incorporating SLC18A2 antibodies could enable high-dimensional profiling at single-cell resolution

• Imaging mass cytometry would allow spatial mapping of SLC18A2 expression in tissue contexts

• Single-cell Western blotting might reveal cell-to-cell variability in SLC18A2 protein levels

Digital Pathology Integration:

• Automated image analysis algorithms could standardize SLC18A2 immunohistochemistry interpretation

• Machine learning approaches might identify subtle staining patterns with prognostic significance

• Whole slide imaging would facilitate comprehensive tissue evaluation beyond the constraints of tissue microarrays

Liquid Biopsy Applications:

• Circulating tumor cell (CTC) analysis using SLC18A2 antibodies might enable non-invasive monitoring

• Extracellular vesicle immunocapture using SLC18A2 antibodies could provide novel biomarker opportunities

• Cell-free DNA methylation analysis of the SLC18A2 promoter might complement tissue-based testing

Therapeutic Targeting Opportunities:

• Development of antibody-drug conjugates targeting SLC18A2 in malignancies with retained expression

• Bispecific antibodies linking SLC18A2 to immune effector cells

• CAR-T approaches incorporating SLC18A2 recognition domains