SSTR1 Antibody

What is SSTR1 and where is it expressed?

SSTR1 (Somatostatin Receptor 1) belongs to the G protein-coupled receptor family that mediates the effects of somatostatin. Based on mRNA expression data and immunohistochemical studies, SSTR1 shows high expression in several tissues and tumors, including pituitary and gastroenteropancreatic tumors, renal tissue, colorectal and breast cancer, meningioma, glioma, neuroblastoma, and pheochromocytoma . Additionally, SSTR1 has been detected in normal pancreatic islet cells, brain tissue, and small intestine tissue . When designing experiments to study SSTR1, researchers should consider this expression pattern to select appropriate positive controls and experimental models.

What are the typical molecular weights observed for SSTR1 in Western blotting?

The molecular weight of SSTR1 observed in Western blotting varies depending on the tissue source, antibody used, and post-translational modifications. While the calculated molecular weight based on amino acid sequence is approximately 43 kDa (391 amino acids), commercially available antibodies report slightly different observed molecular weights:

These variations may result from post-translational modifications such as glycosylation, phosphorylation, or dimerization. When performing Western blot analysis, researchers should anticipate multiple bands and validate specificity using appropriate controls such as SSTR1-transfected cells versus non-transfected cells.

What sample types can be successfully analyzed with SSTR1 antibodies?

SSTR1 antibodies have demonstrated reactivity with various sample types:

When analyzing new sample types, preliminary validation is recommended through comparison with known positive controls and testing of antibody specificity using techniques such as peptide blocking or knockout/knockdown validation.

How can I optimize immunohistochemistry protocols for SSTR1 detection in fixed tissues?

Optimizing immunohistochemistry for SSTR1 requires careful attention to antigen retrieval and signal detection. Based on published protocols, the following methodology is recommended:

• Dewax paraffin sections with xylene, then rehydrate in a graded series of ethanol

• Perform antigen retrieval by microwaving sections in 10 mM citric acid (pH 6.0) for 20 minutes at 600 W

• Block unspecific binding sites with 3% bovine serum albumin for 30 minutes

• Incubate with SSTR1-specific antisera (typical dilution 1:500) overnight at 4°C

• Incubate with biotinylated anti-rabbit IgG (1:200) for 30 minutes

• Incubate with a preformed complex of biotin-peroxidase/streptavidin for 30 minutes

• Visualize antigen-antibody binding sites using diaminobenzidine hydrochloride/H₂O₂ in Tris HCl

Alternatively, for immunofluorescence detection, follow steps 1-4 and then incubate with Cy3-labeled anti-rabbit IgG (1:200) . When troubleshooting weak signals, consider extending primary antibody incubation time or increasing antibody concentration while monitoring background staining.

How do I distinguish between SSTR1 and other somatostatin receptor subtypes in my experiments?

Distinguishing between somatostatin receptor subtypes requires careful selection of subtype-specific antibodies raised against unique epitopes. The scientific literature describes antibodies generated against specific peptide sequences from each receptor subtype:

These antibodies should be validated using positive controls (tissues or cells known to express specific subtypes) and negative controls (knockout/knockdown models). Western blotting can help confirm specificity by revealing different molecular weights for different receptor subtypes. Additionally, dual immunofluorescence labeling can be used to visualize co-expression or differential expression of receptor subtypes within the same tissue section .

What are the critical considerations when interpreting SSTR1 expression data across different tumor types?

When analyzing SSTR1 expression in different tumor types, several important factors must be considered:

• Heterogeneity of expression: As demonstrated in studies of neuroendocrine tumors, SSTR1 expression patterns vary considerably between different tumor types and between individual patients with the same tumor type . This necessitates individualized analysis rather than relying on general expression patterns.

• Methodology standardization: Compare results only when using standardized immunohistochemical protocols, antibody dilutions, and scoring systems. Different detection methods (PCR, in situ hybridization, immunohistochemistry) may yield different results.

• Correlation with functional data: Expression levels may not directly correlate with receptor functionality. Consider complementing expression studies with functional assays measuring somatostatin binding or downstream signaling.

• Co-expression patterns: Analyze SSTR1 expression in the context of other receptor subtypes, as the ratio between different subtypes may impact therapeutic responses to somatostatin analogs.

• Sample processing effects: Fixation methods and times can affect antigen preservation and detection sensitivity. Compare samples processed using identical protocols whenever possible .

How can I resolve specificity issues with SSTR1 antibodies in Western blotting applications?

When encountering specificity concerns with SSTR1 antibodies in Western blotting, implement these troubleshooting approaches:

• Validate with appropriate controls:

  • Positive controls: Cell lines with known SSTR1 expression (SH-SY5Y, BxPC-3)

  • Negative controls: SSTR1 knockout/knockdown samples

  • Peptide competition: Pre-incubate antibody with immunizing peptide to confirm specificity

• Optimize protein extraction:

  • For membrane proteins like SSTR1, use extraction buffers containing mild detergents (0.5-1% Triton X-100 or NP-40)

  • Include protease inhibitors to prevent degradation

  • Avoid excessive heating of samples before loading

• Adjust running and transfer conditions:

  • Use gradient gels (4-12% or 4-20%) to better resolve multiple molecular weight forms

  • Extend transfer time for high molecular weight forms (80 kDa)

  • Include 0.1% SDS in transfer buffer to improve transfer of hydrophobic proteins

• Dilution optimization:

  • Test multiple antibody dilutions (1:500-1:2000 is the recommended range)

  • Extend primary antibody incubation time (overnight at 4°C)

• Blocking optimization:

  • Test alternative blocking agents (5% milk vs. 5% BSA)

  • Block membranes for longer periods (2 hours at room temperature)

What are the best approaches for validating SSTR1 antibody specificity in immunohistochemistry?

To ensure SSTR1 antibody specificity in immunohistochemistry applications:

• Peptide blocking controls: Pre-absorb the antibody with the immunizing peptide and run parallel staining to demonstrate elimination of specific signals.

• Knockout/knockdown validation: Compare staining between wild-type and SSTR1 knockout or knockdown tissues/cells.

• Multiple antibody approach: Confirm staining patterns using different antibodies targeting different epitopes of the same protein.

• Cross-species validation: If the antibody cross-reacts with multiple species, consistent staining patterns across species strengthen confidence in specificity.

• Correlation with other detection methods: Compare immunohistochemistry results with in situ hybridization or RT-PCR data from the same tissue samples.

• Progressive dilution series: Perform staining with serial dilutions of the primary antibody; specific staining should diminish proportionally with dilution, while nonspecific background should reduce more rapidly .

How can SSTR1 immunohistochemistry be used to guide therapeutic decisions for neuroendocrine tumors?

SSTR1 immunohistochemistry provides valuable information for personalized treatment approaches in neuroendocrine tumors:

What are the considerations for using SSTR1 antibodies in flow cytometry for analyzing circulating tumor cells?

Flow cytometry using SSTR1 antibodies requires specific optimization for detecting circulating tumor cells:

• Cell preparation and fixation:

  • For intracellular SSTR1 detection, proper fixation and permeabilization protocols are essential

  • Recommended usage is 0.80 μg per 10^6 cells in a 100 μl suspension

• Antibody validation:

  • Validate antibody performance using positive control cell lines (SH-SY5Y cells have been confirmed to express SSTR1)

  • Include appropriate isotype controls to establish background staining levels

• Multiparameter analysis:

  • Combine SSTR1 detection with other neuroendocrine markers (chromogranin A, synaptophysin) for increased specificity

  • Include epithelial markers (EpCAM) when analyzing circulating tumor cells

• Signal amplification:

  • Consider secondary antibody labeling strategies for enhancing detection sensitivity

  • Biotin-streptavidin systems can amplify signals for low-expression targets

• Gating strategy optimization:

  • Develop specific gating strategies based on forward/side scatter and expression of tumor-specific markers

  • Account for potential autofluorescence of tumor cells

What are the implications of SSTR1 expression in relation to apoptosis induction in tumors?

Recent research suggests important correlations between somatostatin receptor expression and apoptotic pathways:

• Subtype-specific apoptotic effects: While SSTR3 has been most prominently linked to apoptosis induction, the role of SSTR1 in programmed cell death pathways is an emerging area of investigation. Studies have proposed that high-dose somatostatin analogs may induce apoptosis by binding to somatostatin receptors, particularly SSTR3 .

• Therapeutic implications: Understanding the specific contribution of SSTR1 to apoptotic signaling could inform the development of receptor subtype-targeted therapies designed to enhance tumor cell death rather than simply inhibiting secretion or proliferation.

• Combination therapy opportunities: Identifying the mechanisms by which SSTR1 activation influences apoptotic pathways may reveal synergistic opportunities with conventional chemotherapeutics or targeted therapies.

• Expression pattern significance: The heterogeneous expression of SSTR1 across tumor types and patients may partially explain differential responses to somatostatin analog therapy. Comprehensive profiling of receptor subtypes could help predict apoptotic responses.

• Experimental considerations: When investigating SSTR1-mediated apoptosis, researchers should employ multiple complementary methods to detect programmed cell death, including annexin V binding, TUNEL assays, caspase activation, and changes in mitochondrial membrane potential.