GPR142 Antibody

What is GPR142 and where is it primarily expressed?

GPR142 is a G protein-coupled receptor (GPCR) belonging to class "A" of the GPCR family. According to tissue expression studies, GPR142 is predominantly expressed in pancreatic islets, with intense and abundant expression detected through in situ hybridization. Within islets, GPR142 expression has been confirmed in two main cell populations: a subset of insulin-producing β cells and a subset of glucagon-positive α cells . The expression is significantly higher in islet cells compared to exocrine pancreatic tissue, which shows barely detectable expression levels . Additionally, GPR142 has been detected in various cell lines, including monocytic leukemia cell lines (THP-1), T-cell leukemia cell lines (Jurkat), and neuroblastoma cell lines (SH-SY5Y) .

What are the typical molecular weights observed for GPR142 in Western blot analyses?

When working with GPR142 antibodies for Western blot applications, researchers should be aware of the expected molecular weight patterns:

The discrepancy between observed and calculated molecular weights is common for membrane proteins like GPCRs and may reflect post-translational modifications, glycosylation, or the presence of protein complexes . When validating GPR142 antibody specificity, using blocking peptides as negative controls is highly recommended, as demonstrated in the Western blot analyses of mouse and rat tissues .

What applications are GPR142 antibodies validated for?

Based on the available research materials, commercially available GPR142 antibodies have been validated for multiple experimental applications:

For optimal results, researchers should always perform preliminary dilution series optimization for their specific experimental conditions and sample types. Additionally, it's advisable to validate antibody specificity using appropriate positive controls (e.g., GPR142-expressing cell lines) and negative controls (e.g., blocking peptides or GPR142-knockout samples) .

How does GPR142 signaling impact pancreatic islet function and glucose homeostasis?

GPR142 plays a multifaceted role in pancreatic islet function that extends beyond simple insulin secretion. Research has revealed several key mechanisms:

• Insulin secretion pathway: GPR142 agonists function as insulin secretagogues, directly stimulating insulin release from β cells. Importantly, this effect requires intact GLP-1 receptor signaling, indicating a complex signaling interplay .

• Glucagon regulation: Contrary to initial assumptions, GPR142 activation increases glucagon secretion from α cells in both human and mouse islets. This dual effect on both insulin and glucagon suggests a nuanced role in glucose homeostasis regulation .

• GLP-1 production mechanism: Perhaps most significantly, GPR142 activation potentiates glucagon-like peptide-1 (GLP-1) production and release from islets through a mechanism involving upregulation of prohormone convertase 1/3 expression. This represents an intra-islet incretin system that may have important implications for diabetes therapy .

• Cellular protection effects: Beyond hormone secretion, GPR142 agonists increase β cell proliferation and protect both mouse and human islets against stress-induced apoptosis, suggesting a role in preserving islet mass and function .

For researchers studying GPR142 in metabolic disease models, these multiple pathways should be considered when interpreting experimental results, as the net effect on glucose homeostasis will reflect the integrated output of these various mechanisms.

What experimental considerations are important when studying GPR142 with antibody-based techniques?

Researchers investigating GPR142 should consider several methodological factors to ensure reliable results:

• Antibody selection criteria: Choose antibodies with validated reactivity in your species of interest. Available GPR142 antibodies show reactivity with human, mouse, and rat samples, but their performance may vary across applications .

• Epitope considerations: Target selection is critical for GPR142 detection. Available antibodies target:

  • N-terminal region epitopes (NBP2-85004: IMMLPMEQKIQWVPTSLQDITAVLGTEAYTEEDKSMVSHAQKSQHSCLSH)

  • Extracellular domain epitopes (AGR-082: APVHRDWRVHLALD, corresponding to residues 291-304 in the third extracellular loop)

  The choice of epitope may affect detection based on protein conformation or interaction status.

• Storage protocols: For optimal antibody performance, store GPR142 antibodies at 4°C for short-term use (weeks), and aliquot and maintain at -20°C for long-term storage (months to years). Avoid repeated freeze-thaw cycles that can degrade antibody quality .

• Co-localization studies: When determining GPR142 cellular localization, dual-labeling with cell-type-specific markers is essential. In pancreatic tissue, co-staining with insulin or glucagon enables precise identification of GPR142-expressing α and β cells .

• Validation controls:

  • Positive controls: GPR142-overexpressing cell lines or tissues with known high expression

  • Negative controls: Use blocking peptides to confirm signal specificity

  • Secondary-only controls: To assess non-specific binding of secondary antibodies

How can researchers effectively design experiments to study GPR142 agonism and its downstream effects?

When investigating GPR142 activation and its cellular consequences, researchers should consider this experimental workflow:

• Agonist selection: Use selective GPR142 agonists with established pharmacological profiles. Several compounds have been identified through pharmacophore screening approaches .

• Dose-response assessment: Establish complete dose-response curves (typically 10^-9 to 10^-5 M) to determine EC50 values and maximal efficacy of agonists.

• Temporal dynamics: Monitor both acute (minutes to hours) and chronic (days) effects of GPR142 activation, as different cellular pathways may show distinct temporal patterns.

• Readout selection:

  • For insulin secretion: Static incubation or perifusion studies with glucose challenges

  • For GLP-1 production: Analysis of both GLP-1 content in islets and secretion into media

  • For cellular protection: Apoptosis assays following stress induction (e.g., cytokine exposure)

  • For signaling pathways: Analysis of prohormone convertase 1/3 expression changes

• Mechanistic dissection: Use GLP-1 receptor antagonists or GLP-1R-deficient models to distinguish direct GPR142 effects from those mediated through the intra-islet GLP-1 system .

This approach allows for comprehensive characterization of GPR142 function while accounting for the multiple cellular pathways it influences.

What is known about GPR142's role in diabetes and potential therapeutic applications?

GPR142 has emerged as a promising target for diabetes therapy, with several lines of evidence supporting its potential:

• Insulin secretagogue activity: GPR142 agonists stimulate insulin secretion, positioning them as potential therapies for insulin deficiency in diabetes. This effect appears functionally significant in both rodent and human islets .

• GLP-1 augmentation mechanisms: Unlike traditional secretagogues, GPR142 activates both direct insulin release and intra-islet GLP-1 production. This dual mechanism provides a potentially superior therapeutic profile, as GLP-1 offers additional benefits beyond insulin secretion, including β-cell protection and proliferation .

• β-cell preservation effects: GPR142 agonists demonstrate protective effects against stress-induced apoptosis in islets, suggesting potential applications in preserving β-cell mass in early-stage diabetes or following islet transplantation .

• Translational considerations: For researchers developing GPR142-targeting therapies, consideration should be given to species differences in receptor expression patterns and signaling pathways. While core mechanisms appear conserved between rodents and humans, detailed pharmacological profiling in human systems is essential for clinical translation.

How is GPR142 being investigated in cancer research, and what are the emerging findings?

Recent research has begun exploring GPR142's potential role in cancer biology:

• Genomic alterations: Analysis of tumor samples has revealed genomic alterations in GPR142, suggesting possible involvement in oncogenic processes. These findings have prompted investigation of GPR142 as a potential cancer biomarker or therapeutic target .

• Compound screening approaches: Structure-based computational drug design techniques, including pharmacophore development and 3D quantitative structure-activity relationship (QSAR) models, have been employed to identify compounds targeting GPR142 with potential anticancer properties .

• Pan-cancer implications: Intriguingly, compounds identified through GPR142-targeted screening have shown associations with pan-cancer effects. This suggests that GPR142 may be involved in fundamental cellular processes relevant across multiple cancer types .

• Research methodology: For cancer researchers exploring GPR142, methodologies include:

  • Genomic profiling of GPR142 alterations across cancer databases (e.g., The Cancer Genome Atlas)

  • Virtual screening of compound libraries against GPR142 models

  • Validation of hit compounds in cancer cell line panels

  • Analysis of downstream signaling pathways affected by GPR142 modulation

These emerging connections between GPR142 and cancer biology represent an active area of investigation that may reveal novel therapeutic opportunities.

What are the most common technical challenges when working with GPR142 antibodies, and how can they be addressed?

Researchers frequently encounter several challenges when using GPR142 antibodies:

• Specificity verification issues:

  • Challenge: Confirming antibody specificity for GPR142 versus related GPCRs

  • Solution: Employ blocking peptides corresponding to the immunogen sequence, as demonstrated in Western blot analyses of brain and pancreas lysates . Additionally, testing in GPR142 knockout tissues or siRNA-treated cells can provide definitive validation.

• Multiple band detection:

  • Challenge: Western blots often show multiple bands, including the expected ~72 kDa band and additional signals

  • Solution: Perform detailed characterization using reducing/non-reducing conditions, different sample preparation methods (membrane fractionation), and deglycosylation treatments to identify specific GPR142 signals versus artifacts or post-translationally modified forms.

• Low signal-to-noise ratio in immunostaining:

  • Challenge: Detecting GPR142 in tissue sections with high specificity

  • Solution: Optimize antigen retrieval methods (heat-induced versus enzymatic), blocking protocols (BSA versus serum), antibody concentration, and incubation conditions (temperature, duration). For pancreatic tissues specifically, careful handling to prevent digestive enzyme activation is critical.

• Detection in different cellular compartments:

  • Challenge: Distinguishing membrane-localized versus intracellular GPR142

  • Solution: Combine surface labeling techniques (non-permeabilized conditions) with total protein detection (after permeabilization) to assess receptor trafficking. Use confocal microscopy with z-stack analysis for precise localization.

• Cross-species reactivity limitations:

  • Challenge: Variability in antibody performance across species

  • Solution: When working with new species, validate antibody performance using positive controls and consider epitope sequence conservation analysis to predict likely cross-reactivity.

How should researchers interpret discrepancies between observed and calculated molecular weights for GPR142?

A common observation in GPR142 research is the difference between calculated molecular weight (~51 kDa) and observed molecular weight in Western blots (~72 kDa) . When encountering such discrepancies, consider these methodological explanations:

• Post-translational modifications: GPCRs frequently undergo extensive modifications, including:

  • N-linked glycosylation at asparagine residues in extracellular domains

  • Palmitoylation at cysteine residues

  • Phosphorylation at serine/threonine residues

• Verification approach: To determine if glycosylation explains the higher molecular weight:

  • Treat samples with deglycosylation enzymes (PNGase F for N-linked glycans)

  • Compare migration patterns before and after treatment

  • A significant shift toward the calculated molecular weight would confirm glycosylation

• Oligomerization consideration: GPCRs can form dimers or higher-order oligomers that may not fully dissociate in SDS-PAGE conditions. To assess this possibility:

  • Compare reducing versus non-reducing conditions

  • Apply stronger denaturing protocols (higher SDS concentration, urea addition)

  • Use cross-linking approaches to stabilize potential protein-protein interactions

• Hydrophobic protein behavior: As a transmembrane protein, GPR142 contains highly hydrophobic domains that can bind more SDS molecules, potentially altering migration patterns. Sample preparation using specialized protocols for membrane proteins may help address this issue.

Understanding these factors is critical for accurate interpretation of Western blot results and confirming antibody specificity in GPR142 research.

How do GPR142 expression patterns compare between different experimental models and human tissues?

When designing GPR142 research, understanding expression patterns across models is essential for experimental design and data interpretation:

This comparative expression analysis reveals several important considerations:

• Translational relevance: The similar expression patterns between human and rodent islets suggest rodent models may have translational value for studying GPR142 function in diabetes.

• Cell line selection: For in vitro studies, researchers can consider THP-1, Jurkat, or SH-SY5Y cells as potential models expressing endogenous GPR142.

• Tissue specificity: The enriched expression in islets versus exocrine pancreas suggests GPR142-targeted therapies might achieve relatively selective effects on endocrine pancreatic function.

• Unexpected expression sites: The detection in brain tissue and multiple cell types beyond pancreatic islets warrants investigation of GPR142 functions in these contexts.

What analytical approaches can distinguish direct versus indirect effects of GPR142 activation in complex biological systems?

When studying GPR142 function, particularly in pancreatic islets, researchers face the challenge of distinguishing direct receptor effects from secondary signaling cascades. Based on current research, these analytical strategies can help:

• Temporal dissection: Monitor acute (minutes) versus sustained (hours to days) responses to GPR142 agonists. Immediate effects (e.g., calcium mobilization) likely represent direct signaling, while delayed responses may involve transcriptional changes and secondary mediators.

• Pharmacological inhibitor studies: Systematically apply inhibitors of potential downstream mediators:

  • GLP-1 receptor antagonists: To block the contribution of intra-islet GLP-1 signaling

  • PC1/3 inhibitors: To prevent prohormone processing required for GLP-1 production

  • Traditional GPCR pathway inhibitors (PKA, PKC, MAPK inhibitors)

• Genetic manipulation approaches:

  • siRNA knockdown of GPR142 versus downstream mediators

  • CRISPR-based GPR142 knockout models

  • Cell-specific conditional knockout models (e.g., β-cell-specific versus α-cell-specific)

• Single-cell analysis techniques:

  • Single-cell RNA sequencing to identify cell-type-specific responses

  • Single-cell calcium imaging to distinguish direct cellular responders

  • FRET-based biosensors for real-time signaling pathway activation

• Ex vivo versus in vivo comparison: Effects observed in isolated islets that are absent or modified in vivo may indicate systemic compensatory mechanisms or neural/hormonal modulation.

Research has revealed that GPR142's effects on insulin secretion require intact GLP-1 receptor signaling , demonstrating the importance of such mechanistic dissection approaches for accurate functional characterization.

What emerging technologies might advance GPR142 research beyond current methodological limitations?

Several cutting-edge approaches have potential to overcome existing challenges in GPR142 research:

• Structural biology advances:

  • Cryo-electron microscopy for GPR142 structure determination in active and inactive states

  • Hydrogen-deuterium exchange mass spectrometry to map ligand binding sites

  • These approaches could facilitate rational design of highly selective agonists and antagonists

• Biosensor development:

  • GPCR activation sensors based on conformational changes

  • Genetically-encoded indicators for downstream signaling events

  • These tools would enable real-time monitoring of GPR142 activation in living cells

• Single-cell multiomics:

  • Integrated transcriptomic, proteomic, and metabolomic analysis at single-cell resolution

  • Spatial transcriptomics to map GPR142 expression patterns with preserved tissue architecture

  • These methods would reveal cell-type-specific responses and heterogeneity within GPR142-expressing populations

• Advanced tissue models:

  • Organoid systems incorporating multiple islet cell types

  • Microfluidic pancreas-on-chip models with controlled perfusion

  • These platforms would better recapitulate the complex cellular interactions in GPR142 signaling

• In vivo imaging innovations:

  • PET ligands for GPR142 to enable non-invasive receptor visualization

  • Reporter mouse models for real-time monitoring of GPR142-dependent signaling pathways

  • These approaches would facilitate translation between in vitro findings and physiological relevance

How might findings from GPR142 research in different disease contexts be integrated to develop comprehensive therapeutic strategies?

The multifaceted roles of GPR142 across diabetes and potentially cancer research suggest several integrative research approaches:

• Pathway convergence analysis:

  • Systematic comparison of GPR142 signaling pathways activated in different disease contexts

  • Identification of common versus context-specific downstream effectors

  • This would reveal whether similar mechanism-based monitoring approaches apply across diseases

• Translational biomarker development:

  • Assessment of circulating GPR142 ligands in patient cohorts with diabetes, cancer, or comorbidities

  • Correlation with disease progression and treatment response

  • This could enable patient stratification for GPR142-targeted therapies

• Dual-purpose compound screening:

  • Testing GPR142 agonists identified for diabetes in cancer models and vice versa

  • Comprehensive profiling of effects on metabolism and cell proliferation

  • This might identify compounds with favorable dual properties or reveal contraindications

• Comorbidity models:

  • Development of experimental systems modeling diabetes with increased cancer risk

  • Investigation of GPR142 modulation effects on both conditions simultaneously

  • This would address potential benefits or risks in complex patient populations

• Targeted delivery strategies:

  • Cell-type specific targeting approaches for GPR142 modulators

  • Tissue-selective drug delivery systems

  • This could enhance therapeutic index by concentrating effects in target tissues

The emerging connection between GPR142, diabetes, and cancer highlights the importance of such integrative approaches to fully leverage therapeutic opportunities while mitigating potential risks.

What are the most critical validation steps researchers should perform when using a new GPR142 antibody?

When adopting a new GPR142 antibody for research, a systematic validation workflow is essential:

• Initial characterization:

  • Confirm reactivity with recombinant GPR142 protein or overexpression systems

  • Verify expected molecular weight pattern in Western blots (~72 kDa observed, ~51 kDa calculated)

  • Compare staining patterns with previously validated antibodies when possible

• Specificity controls:

  • Test with blocking peptide corresponding to the immunogen sequence

  • Compare signal in GPR142-high versus GPR142-low/negative tissues or cell lines

  • If available, validate in GPR142 knockout/knockdown models

• Application-specific optimization:

  • For Western blot: Optimize sample preparation, blocking, and antibody concentration (1:500-1:2000)

  • For immunostaining: Develop appropriate antigen retrieval protocols and validate co-localization with expected cell-type markers (insulin, glucagon for islet studies)

  • For all applications: Include appropriate positive and negative controls in each experiment

• Cross-reactivity assessment:

  • Test performance across intended species (human, mouse, rat)

  • Consider potential cross-reactivity with closely related GPCRs

  • Verify performance in multiple experimental models relevant to your research

These validation steps are essential for generating reliable and reproducible data in GPR142 research and should be thoroughly documented in research publications.

What interdisciplinary approaches might yield the most significant advances in understanding GPR142 biology?

The complex biology of GPR142 spanning diabetes, potential cancer connections, and other physiological functions suggests that collaborative approaches integrating multiple disciplines will yield the most comprehensive insights:

• Integrated expertise requirements:

  • Molecular pharmacologists: For ligand discovery and receptor characterization

  • Islet biologists: For pancreatic function assessment

  • Medicinal chemists: For optimizing selective GPR142 modulators

  • Computational biologists: For pathway analysis and target identification

  • Clinical researchers: For translational validation in patient samples

• Technology integration opportunities:

  • Combining structural biology with artificial intelligence for drug design

  • Merging single-cell transcriptomics with functional assays

  • Integrating in vivo imaging with precise genetic manipulation tools

• Translational research framework:

  • Bidirectional workflow between basic mechanism discovery and clinical observations

  • Integration of findings from metabolic disease and cancer research contexts

  • Development of biomarkers in parallel with therapeutic candidates