CDH18 Antibody

What is CDH18 and why is it important in research?

CDH18 (Cadherin 18, Type 2) is a calcium-dependent adhesion protein primarily localized to the cell membrane. It plays significant roles in cellular adhesion processes and has emerged as a promising biomarker in cancer research, particularly in uterine corpus endometrial carcinoma (UCEC). Studies have demonstrated that CDH18 expression correlates with tumor progression, immune cell infiltration patterns, and response to chemotherapeutic agents. This multifaceted involvement in cancer biology makes CDH18 an important research target for understanding disease mechanisms and developing potential therapeutic strategies. Recent findings indicate that CDH18 expression increases with higher WHO grades of UCEC, suggesting its potential utility as a prognostic indicator .

What types of CDH18 antibodies are available for research applications?

Several types of CDH18 antibodies are available for research, varying in their target regions, host species, clonality, and applications:

When selecting an antibody, researchers should consider the specific target region of interest, required applications, and species reactivity based on their experimental design .

How should CDH18 antibodies be stored and handled to maintain optimal activity?

For optimal maintenance of CDH18 antibody activity, follow these methodological guidelines: Store antibodies at -20°C for long-term storage and at 4°C for short-term use (typically up to two weeks). Avoid repeated freeze-thaw cycles by aliquoting the antibody into smaller volumes before freezing. When working with the antibody, maintain cold chain conditions by keeping it on ice. Dilute antibodies in appropriate buffers containing stabilizers such as BSA (0.1-1%) or carrier proteins. For applications requiring higher sensitivity, prepare fresh working dilutions on the day of the experiment. Always centrifuge the antibody vial briefly before opening to ensure all liquid is at the bottom of the vial. Follow manufacturer-specific recommendations for each antibody, as storage conditions may vary based on formulation. Monitor antibody performance regularly by including positive controls in experiments to detect any potential degradation over time .

What are the common applications of CDH18 antibodies in research?

CDH18 antibodies are versatile tools employed across multiple research applications. In Western Blotting (WB), they detect endogenous levels of CDH18 protein, allowing quantitative analysis of expression across different samples. For Immunohistochemistry (IHC), these antibodies visualize CDH18 localization in tissue sections, revealing its predominantly membranous expression pattern that intensifies with increasing cancer grades. Immunofluorescence (IF) applications provide high-resolution imaging of CDH18 distribution in both cell lines and tissue samples, enabling co-localization studies with other proteins of interest. In ELISA assays, CDH18 antibodies facilitate sensitive quantification of protein levels in various sample types. Additionally, researchers utilize these antibodies in immunoprecipitation experiments to study protein-protein interactions involving CDH18. The selection of application should align with research objectives, and optimization of antibody concentration is essential for each specific application to achieve optimal signal-to-noise ratios .

How can CDH18 antibodies be utilized to study its role as a biomarker in cancer research?

CDH18 antibodies can be strategically employed to investigate its biomarker potential in cancer through multiple methodological approaches. For prognostic evaluation, researchers should perform immunohistochemistry on tissue microarrays containing samples with linked survival data to correlate CDH18 expression with clinical outcomes. Importantly, quantitative analysis should incorporate both staining intensity and percentage of positive cells, ideally using digital pathology algorithms for standardization. For mechanistic studies, combine immunofluorescence with markers of tumor progression (Ki-67, p53) to evaluate co-expression patterns. Western blotting analysis across cell lines representing different stages of malignancy can establish expression patterns correlated with aggressiveness. To understand CDH18's clinical utility, researchers should design studies comparing antibody performance across multiple tumor grades and correlate findings with patient survival data. Recent research demonstrates that CDH18 expression increases with higher WHO grades of uterine corpus endometrial carcinoma (UCEC) and associates with poorer prognosis, making it a valuable prognostic indicator. Additionally, incorporating CDH18 into predictive nomogram models has shown improved accuracy in forecasting patient outcomes, suggesting its potential integration into clinical risk assessment tools .

What approaches should be used to validate CDH18 antibody specificity for research applications?

Validating CDH18 antibody specificity requires a multi-faceted approach to ensure reliable research outcomes. First, conduct Western blotting with positive and negative control samples, where positive controls should show a single band at the expected molecular weight (~88 kDa for full-length CDH18), while negative controls should show no significant bands. Implement peptide competition assays by pre-incubating the antibody with the immunizing peptide prior to immunostaining; specific binding will be blocked, resulting in signal reduction. Utilize genetic approaches by performing immunostaining in CDH18 knockout models or CRISPR-edited cell lines lacking CDH18 expression; absence of staining in these samples confirms specificity. Compare staining patterns across multiple antibodies targeting different epitopes of CDH18; consistent localization patterns enhance confidence in specificity. For polyclonal antibodies like ABIN3183613, which was affinity-purified using epitope-specific immunogen derived from the internal region of human Cadherin-18, verify cross-reactivity with intended species through comparative analysis. Additionally, perform immunoprecipitation followed by mass spectrometry to confirm that the antibody pulls down CDH18 and not other cadherin family members. Document all validation procedures thoroughly for publication and reproducibility purposes .

How can researchers correlate CDH18 expression with immune cell infiltration in tumor microenvironments?

To effectively correlate CDH18 expression with immune cell infiltration, researchers should implement a comprehensive methodological approach combining various techniques. Begin with multiplex immunofluorescence using CDH18 antibodies alongside markers for key immune cell populations (CD8+ T cells, NK cells, M2 macrophages) on serial tissue sections to visualize spatial relationships. Quantify co-localization patterns using digital image analysis software that can measure both proximity and density relationships. For higher-throughput analysis, perform flow cytometry on dissociated tumor samples to quantitatively assess CDH18 expression in relation to immune cell populations. When analyzing public datasets, employ computational tools like CIBERSORT to estimate immune cell proportions from gene expression data and correlate with CDH18 expression levels. Recent research has revealed significant correlations between CDH18 expression and immune cell populations in the tumor microenvironment, specifically a negative correlation with CD8+ T cells and positive correlations with resting NK cells and M2 macrophages. Additionally, CDH18 shows important associations with immune checkpoint molecules, correlating positively with VTCN1 (associated with poor prognosis) and negatively with several tumor-inhibiting genes (TNFSF14, TNFSF15, PDCD1, TNFRSF14, and CD44). These findings suggest CDH18's potential role in immune modulation within the tumor microenvironment, which has significant implications for immunotherapy approaches .

What experimental designs are optimal for investigating the relationship between CDH18 expression and drug resistance?

To rigorously investigate CDH18's relationship with drug resistance, researchers should implement a comprehensive experimental framework. Begin by establishing stable cell lines with CDH18 overexpression or knockdown using lentiviral vectors or CRISPR-Cas9 systems, followed by IC50 determination across multiple drug classes. Perform dose-response curves using at least 7-8 concentrations in triplicate to accurately calculate IC50 values. Complement in vitro studies with patient-derived xenograft models comparing treatment responses between high and low CDH18-expressing tumors. For clinical correlation, analyze archival tumor samples from patients with known treatment outcomes using CDH18 immunohistochemistry with quantitative image analysis. Recent research has identified significant correlations between CDH18 expression and drug sensitivity; notably, elevated IC50 values for multiple agents including (5Z)-7-Oxozeaenol, AG-014699, CEP-701, Mitomycin C, PD-0325901, PD-0332991, PHA-665752, SL 0101-1, and SN-38 were observed in high CDH18-expressing samples. These findings suggest that patients with elevated CDH18 levels may exhibit reduced sensitivity to these therapeutic agents. To further elucidate mechanisms, perform RNA-seq on matched sensitive and resistant cell models to identify altered pathways associated with CDH18-mediated resistance. Additionally, integrate computational prediction tools like pRRophetic for preliminary drug response assessment based on expression profiles before experimental validation .

What controls should be included when using CDH18 antibodies in various experimental applications?

When designing experiments with CDH18 antibodies, comprehensive controls are essential for result validation and troubleshooting. For Western blotting, include a positive control (cell line or tissue with known CDH18 expression, such as HEC-1-B endometrial cancer cells), a negative control (cell line with minimal CDH18 expression, such as some HEEC non-cancer cells), and a loading control (β-actin or GAPDH) to normalize protein levels. In immunohistochemistry and immunofluorescence studies, incorporate serial sections stained with isotype control antibodies matching the CDH18 antibody's host species to identify non-specific binding. Include tissue samples with graded expression levels (as seen in WHO grades of UCEC) to verify staining sensitivity. For ELISA applications, prepare standard curves using recombinant CDH18 protein and include blank wells (no primary antibody) to establish background signal levels. When studying functional aspects, utilize CDH18 knockout models (such as those generated by GemPharmatech) as biological negative controls. For cross-reactivity verification, test the antibody on samples from multiple species if working with an antibody like ABIN3183613 that claims multi-species reactivity. Additionally, when evaluating CDH18 in relation to disease progression, include samples representing different disease stages, as CDH18 expression has been shown to increase with higher grades of cancer .

What are the optimal sample preparation methods for detecting CDH18 using immunological techniques?

For optimal CDH18 detection using immunological techniques, sample preparation methodology should be tailored to preserve both protein integrity and cellular architecture. For tissue samples in immunohistochemistry and immunofluorescence: Fix tissues in 10% neutral buffered formalin for 24-48 hours, followed by paraffin embedding. Perform antigen retrieval using citrate buffer (pH 6.0) or EDTA buffer (pH 9.0) at 95-98°C for 15-20 minutes, as CDH18 is a membrane-bound protein that may require enhanced epitope exposure. For cell lines: Culture cells on glass coverslips or chamber slides until 70-80% confluent, fix with 4% paraformaldehyde for 15 minutes at room temperature, and permeabilize with 0.1-0.5% Triton X-100 for intracellular epitopes. For Western blotting: Extract proteins using RIPA buffer supplemented with protease inhibitors, maintaining cold temperatures throughout to prevent degradation. Include 1mM calcium in extraction buffers as CDH18 is calcium-dependent. Denature samples at 95°C for 5 minutes in Laemmli buffer containing β-mercaptoethanol before gel loading. For RNA analysis: Extract total RNA using Trizol reagent as demonstrated in recent studies, where 2μg RNA was reverse transcribed using High-Capacity cDNA Reverse Transcription Kit, with β-actin serving as an internal control. These methodological considerations are critical for accurate detection of CDH18, particularly given its membrane localization observed in cancer progression studies .

What considerations should be made when designing CDH18 knockout or knockdown experiments?

When designing CDH18 knockout or knockdown experiments, researchers should implement a comprehensive methodological framework to ensure valid outcomes. For CRISPR-Cas9 knockout strategies, design multiple guide RNAs targeting conserved functional domains of CDH18, particularly within early exons to ensure complete protein disruption. Validate knockouts through genomic sequencing, protein expression analysis (Western blot, immunofluorescence), and functional assays relevant to cellular adhesion properties. For transient knockdown approaches using siRNA, design at least 3-4 different siRNA sequences targeting different regions of the CDH18 transcript to control for off-target effects, and optimize transfection conditions for each cell type to achieve >80% reduction in expression. When using shRNA for stable knockdown, employ inducible systems (tetracycline-regulated) to distinguish between acute and chronic effects of CDH18 depletion. Include appropriate controls in all experimental designs: non-targeting gRNA/siRNA/shRNA controls, wild-type parental cells, and rescue experiments where the knockdown phenotype is complemented by re-expression of CDH18. For in vivo studies, consider generating conditional knockout models rather than constitutive knockouts, as CDH18 may have developmental roles. Previous research has successfully employed CDH18 knockout mice generated by GemPharmatech for reproductive biology studies, demonstrating the feasibility of genetic manipulation approaches. Monitor potential compensatory mechanisms by other cadherin family members (particularly type II cadherins) that may mask phenotypes in chronic depletion models .

How can researchers address common issues with CDH18 antibody specificity and sensitivity?

To overcome specificity and sensitivity challenges with CDH18 antibodies, researchers should implement a systematic troubleshooting approach. For weak or absent signals in Western blotting, gradually increase antibody concentration (starting from 1:1000 and adjusting up to 1:250 if necessary) while extending primary antibody incubation to overnight at 4°C. Optimize antigen retrieval methods for immunohistochemistry and immunofluorescence by testing multiple buffer systems (citrate pH 6.0 versus EDTA pH 9.0) and retrieval times, as CDH18 epitopes may require specific unmasking conditions. To reduce background staining, implement additional blocking steps using 5% normal serum from the secondary antibody host species, and include 0.1-0.3% Triton X-100 in blocking buffers to reduce non-specific binding. For cross-reactivity concerns, conduct peptide competition assays by pre-incubating the antibody with immunizing peptide; specific signals should be significantly reduced or eliminated. If working with polyclonal antibodies like ABIN3183613 that may show batch-to-batch variability, consider affinity purification against the target epitope to enhance specificity. When interpreting results across different applications, note that antibody performance may vary; for instance, an antibody performing well in Western blotting may require different optimization for immunofluorescence. Recent research successfully employed CDH18 antibody (No:13091-1-AP from Proteintech) for immunofluorescence analysis of both cell lines and tissue samples, demonstrating proper membrane localization consistent with cadherin family proteins .

What approaches should be used to differentiate between CDH18 and other cadherin family members in experimental analyses?

To effectively differentiate CDH18 from other cadherin family members, researchers should implement a multi-faceted approach focusing on specificity validation and careful experimental design. Begin by selecting antibodies targeting unique regions of CDH18 that show minimal sequence homology with other cadherins, particularly those targeting the EC4-EC5 domains or the cytoplasmic domain which exhibit greater sequence divergence. Perform comprehensive pre-validation using Western blotting against recombinant proteins of multiple cadherin family members to confirm absence of cross-reactivity. For genetic approaches, design PCR primers spanning intron-exon boundaries unique to CDH18, and validate specificity through sequencing of amplicons. When performing knockout/knockdown validation, monitor expression of closely related cadherins (particularly type II cadherins like CDH7, CDH11) to detect potential compensatory upregulation. For mass spectrometry-based studies, focus on unique peptides derived from CDH18 for protein identification. In immunohistochemistry applications, compare staining patterns with other cadherins in serial sections to identify distinctive distribution patterns; CDH18 shows characteristic membrane localization with increasing intensity in higher grades of cancer. Additionally, leverage bioinformatic approaches to design experiments targeting CDH18-specific regulatory elements or splicing variants. Researchers should rigorously document these validation steps in publications to establish reproducibility and reliability of CDH18-specific findings .

How should researchers interpret contradictory results between different detection methods for CDH18?

When encountering contradictory results between detection methods for CDH18, researchers should implement a systematic analytical framework to resolve discrepancies. Begin by examining method-specific limitations: Western blotting detects denatured proteins and may miss conformational epitopes recognized in immunohistochemistry/immunofluorescence; conversely, antibodies optimized for native conditions may perform poorly in denaturing applications. Consider epitope accessibility differences - CDH18 is a membrane-bound protein, and certain epitopes may be masked in specific sample preparation methods. Evaluate whether discrepancies arise from methodology sensitivities; qRT-PCR measures mRNA levels which may not directly correlate with protein abundance due to post-transcriptional regulation. Implement orthogonal validation approaches using alternative antibodies targeting different CDH18 epitopes to confirm findings. When differences persist between in vitro and in vivo results, recognize that microenvironmental factors influence CDH18 expression and function. For technical reconciliation, systematically optimize each method's parameters including sample preparation, antibody concentration, and detection systems. In recent research comparing CDH18 expression across platforms, investigators validated TCGA database findings with both immunofluorescence and RT-PCR, finding concordant results that confirmed increased expression in cancer samples. This methodological triangulation approach strengthens confidence in results despite platform differences. Additionally, when integrating public database information with laboratory findings, consider batch effects and normalization methods that might contribute to apparent contradictions .

What emerging technologies might enhance CDH18 antibody applications in research?

Emerging technologies are poised to significantly advance CDH18 antibody applications across multiple research domains. Single-cell proteomics using mass cytometry (CyTOF) will enable high-dimensional analysis of CDH18 expression in heterogeneous tumor samples, revealing previously undetectable subpopulations and their unique microenvironmental interactions. Spatial transcriptomics platforms can be integrated with CDH18 immunofluorescence to correlate protein expression with local gene expression profiles, providing unprecedented insights into tumor heterogeneity and microenvironmental influences. Advanced super-resolution microscopy techniques (STORM, PALM) will enhance visualization of CDH18 membrane dynamics at nanoscale resolution, potentially revealing novel interaction partners and subcellular localization patterns. For therapeutic development, CDH18 antibody-drug conjugates represent a promising approach for targeted delivery of cytotoxic agents to CDH18-overexpressing tumor cells. Intravital imaging with fluorescently labeled CDH18 antibodies could enable real-time monitoring of tumor progression and treatment response in preclinical models. Computationally, machine learning algorithms can be trained on CDH18 immunohistochemistry images to develop automated scoring systems with improved reproducibility and predictive power. Additionally, CRISPR screening approaches paired with CDH18 antibody-based detection will facilitate systematic identification of genes regulating CDH18 expression and function. These technological advances will collectively enhance our understanding of CDH18's role in disease processes and its potential as a therapeutic target or biomarker .

What are the current limitations in CDH18 antibody research and how might they be addressed in future studies?

Current CDH18 antibody research faces several methodological limitations that require strategic approaches to overcome in future investigations. Epitope specificity remains challenging due to sequence homology with other cadherin family members; future studies should implement comprehensive cross-reactivity testing against multiple cadherins and develop monoclonal antibodies targeting CDH18-unique regions. Reproducibility issues arise from antibody lot-to-lot variations; researchers should establish standardized validation protocols including Western blot, immunohistochemistry, and knockout controls for each new lot. The field lacks consensus on quantification methods for immunohistochemistry results; development of automated digital pathology algorithms specifically calibrated for CDH18 would enhance standardization. Current functional understanding is limited by predominantly correlative studies; mechanistic investigations using CRISPR-engineered cell lines and animal models are needed to establish causality in CDH18-associated phenotypes. Translational applications are hindered by insufficient prospective clinical validation; future multicenter studies should evaluate CDH18 as a biomarker in prospectively collected cohorts with comprehensive clinical annotation. Additionally, existing research demonstrates geographic and demographic gaps, focusing primarily on specific populations; expansion to diverse ethnic backgrounds would enhance generalizability. Knowledge of post-translational modifications affecting CDH18 function and antibody recognition is currently limited; mass spectrometry studies mapping these modifications would improve interpretation of antibody-based detection results. Addressing these limitations through methodological innovations and collaborative research will significantly advance the utility of CDH18 antibodies in both basic science and clinical applications .

How can researchers effectively integrate CDH18 antibody data with other -omics approaches for comprehensive analyses?

Effective integration of CDH18 antibody data with multi-omics approaches requires systematic methodological strategies to generate meaningful insights. Researchers should implement antibody-based proteomics through reverse phase protein arrays (RPPA) or mass spectrometry-based approaches to correlate CDH18 protein levels with broader proteomic landscapes. For transcriptomic integration, perform immunohistochemistry or Western blotting on samples with matched RNA-seq data to identify relationships between CDH18 protein expression and global gene expression patterns. Recent studies utilized this approach to correlate CDH18 expression with immune checkpoint molecules and tumor-infiltrating immune cells, revealing significant associations with CD8+ T cells, NK cells, and M2 macrophages. For pathway analysis, implement Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses on genes correlated with CDH18 expression, using methods like Gene Set Enrichment Analysis (GSEA) with defined cutoffs (p < 0.05, FDR < 0.25) to identify enriched biological processes. To evaluate genomic correlations, analyze CDH18 mutation patterns using platforms like cBioPortal, which has been used to identify 13 CDH18 mutation sites with prognostic significance in UCEC. For clinical data integration, develop multivariate models incorporating CDH18 expression alongside clinical parameters to improve prognostic accuracy. Computational approaches should leverage machine learning algorithms to identify complex patterns across multi-omics datasets, potentially revealing novel CDH18-associated signatures. Additionally, single-cell multi-omics techniques combining protein and transcript detection can reveal cell type-specific CDH18 expression patterns within heterogeneous samples, providing unprecedented resolution of CDH18 biology in complex tissues .