CELF6 Antibody, HRP conjugated

What is CELF6 and why is it important in neurological research?

CELF6 belongs to the CELF family of RNA-binding proteins with established roles in human health and disease. It functions primarily as a repressive RNA-binding protein in the central nervous system (CNS) that downregulates specific mRNAs in vivo . CELF6 mediates exon inclusion and/or exclusion in pre-mRNAs that undergo tissue-specific and developmental regulation .

Recent CLIP-Seq (Cross-Linking Immunoprecipitation-Sequencing) studies have revealed that CELF6 predominantly associates with 3'UTRs of target mRNAs in the brain, many of which code for synaptic proteins . This suggests a critical role in neuronal function and potential implications for neurodevelopmental disorders. The protein's ability to decrease RNA abundance in a sequence-dependent manner, reversible by mutating UGU-rich motifs, represents a significant regulatory mechanism in neuronal gene expression .

What are the key applications for CELF6 antibodies in experimental neuroscience?

CELF6 antibodies serve numerous critical applications in neuroscience research, with Western blotting, immunohistochemistry, and flow cytometry being particularly valuable . In Western blotting, these antibodies enable researchers to detect and quantify CELF6 protein levels across different experimental conditions or tissue samples, providing insights into expression patterns during development or in pathological states .

For immunohistochemistry on paraffin-embedded tissues (IHC-P), CELF6 antibodies allow visualization of protein localization within specific cell types or brain regions . This application is especially relevant given CELF6's cell-specific expression patterns in the CNS. Flow cytometry with CELF6 antibodies facilitates analysis of intracellular protein expression at the single-cell level, enabling more nuanced studies of cell heterogeneity in neural populations . These diverse applications make CELF6 antibodies versatile tools for investigating both normal neurobiological processes and disease mechanisms.

How does HRP conjugation enhance detection sensitivity in CELF6 antibody applications?

HRP (Horseradish Peroxidase) conjugation significantly improves detection sensitivity in CELF6 antibody applications through enzymatic signal amplification. When an HRP-conjugated CELF6 antibody binds to its target protein, the attached enzyme catalyzes the oxidation of substrates (such as TMB or luminol), producing a colorimetric, chemiluminescent, or fluorescent signal that is substantially amplified compared to direct detection methods .

This enzymatic amplification is particularly valuable when studying CELF6 in scenarios where protein expression is dynamically regulated or present at low abundance, as demonstrated in cell cycle studies where CELF6 levels fluctuate significantly throughout different phases . The enhanced sensitivity also reduces the amount of sample required, which is beneficial when working with limited brain tissue specimens. Additionally, HRP conjugation eliminates the need for secondary antibody incubation steps in many protocols, streamlining experimental workflows and reducing potential sources of background signal or cross-reactivity.

What are the optimal storage and handling conditions for maintaining CELF6 antibody activity?

Proper storage and handling of CELF6 antibodies are crucial for maintaining their specificity and sensitivity. HRP-conjugated antibodies are particularly susceptible to activity loss through enzyme denaturation or oxidation. To preserve optimal activity, these antibodies should be stored at -20°C in small aliquots to minimize freeze-thaw cycles, which can significantly compromise antibody performance .

When working with CELF6 antibodies, researchers should avoid exposure to sodium azide, as this preservative inhibits HRP activity. Instead, ProClin or similar preservatives are recommended for long-term storage solutions. Working dilutions should be prepared fresh in buffers containing stabilizing proteins such as BSA (0.5-1%) and used within 24 hours. For immunohistochemistry applications, maintaining proper pH during antigen retrieval is critical, as CELF6 epitope recognition can be pH-dependent . Finally, researchers should validate antibody performance in their specific experimental system, as CELF6 expression patterns and accessibility may vary across different tissue types and cell lines.

How can CELF6 antibodies be effectively utilized in CLIP-Seq experiments to identify RNA targets?

CLIP-Seq represents a powerful approach for identifying the in vivo RNA targets of CELF6, though this application presents unique challenges requiring methodological optimization. Successful CLIP-Seq protocols for CELF6 have employed epitope-tagged versions (such as CELF6-YFP/HA) when direct antibodies prove problematic for immunoprecipitation of the endogenous protein . This strategy requires careful validation to ensure the tagged protein maintains physiological expression patterns and binding characteristics.

A critical consideration is the adjustment of standard CLIP protocols, as stringent lysis and wash conditions may be incompatible with CELF6 immunoprecipitation . Researchers should incorporate appropriate controls including: (1) immunoprecipitation from uncrosslinked samples, (2) immunoprecipitation from wild-type tissue lacking the target protein, and (3) input samples to normalize for starting transcript abundance, thus avoiding bias toward highly expressed genes . For enrichment analysis, statistical comparison with these controls allows rigorous identification of true CELF6 targets. Post-CLIP-Seq bioinformatic analysis should focus on motif discovery within binding regions, with particular attention to UGU-rich sequences previously identified as preferential binding sites for CELF family proteins . Additionally, researchers should examine the positional distribution of binding sites, as CELF6 has shown enrichment near polyadenylation signals, suggesting functional relevance in mRNA processing .

What are the best practices for optimizing Western blot protocols using HRP-conjugated CELF6 antibodies?

Optimizing Western blot protocols for CELF6 detection requires careful consideration of several parameters to achieve specific and sensitive results. When using HRP-conjugated CELF6 antibodies, protein extraction methods significantly impact detection quality. CELF6 is primarily detected at approximately 78 kDa for the YFP/HA-tagged version or about 50-55 kDa for the endogenous protein . Researchers should employ protease inhibitors during extraction to prevent degradation, particularly important since CELF6 undergoes proteasomal degradation in a cell cycle-dependent manner .

Blocking conditions require careful optimization; 5% nonfat milk has been successfully used in published protocols , though BSA-based blocking may be preferable when phospho-specific detection is important. Primary antibody concentration should be titrated for each experimental system, typically starting at 1:1000 dilution and adjusting based on signal-to-noise ratio. For enhanced sensitivity without background, incorporation of longer, lower-temperature incubations (overnight at 4°C) often yields superior results. When studying cell cycle-regulated expression of CELF6, synchronization methods significantly impact detection outcomes; both double-thymidine block and selective CDK1 inhibitor (RO-3306) approaches have successfully revealed CELF6's dynamic expression patterns . For quantitative analysis, normalization to appropriate loading controls is essential, with consideration for controls that remain stable across experimental conditions, particularly important when examining cell cycle-dependent expression.

What methodological approaches can detect interactions between CELF6 and other proteins in regulatory complexes?

Investigating CELF6's protein interactions requires sophisticated methodological approaches that preserve physiologically relevant complexes. Co-immunoprecipitation (Co-IP) has successfully demonstrated interactions between CELF6 and regulatory proteins such as β-TrCP, which mediates its ubiquitination and degradation . When designing Co-IP experiments, researchers should consider using epitope-tagged versions of CELF6 (e.g., GFP-CELF6) to overcome limitations with direct antibody precipitation while ensuring the tags don't interfere with complex formation.

For detecting dynamic interactions that occur during specific cellular processes, such as cell cycle progression, synchronized cell populations are essential. Cell synchronization can be achieved through double-thymidine block for G1/S boundary studies or selective CDK1 inhibitors like RO-3306 for G2/M arrest . When investigating ubiquitination-dependent interactions, incorporation of proteasome inhibitors (e.g., MG132) prevents degradation of ubiquitinated CELF6, significantly enhancing detection sensitivity . Advanced approaches like proximity ligation assay (PLA) can provide spatial information about CELF6 interactions within intact cells, while mass spectrometry following immunoprecipitation can identify novel interaction partners. For validating functional consequences of these interactions, researchers should combine protein interaction studies with transcriptional analyses of target genes like p21, which has demonstrated significant sensitivity to CELF6 expression levels across multiple cell types .

How do CELF6 expression patterns across different tissue types affect antibody selection and experimental design?

CELF6 displays distinct tissue-specific and subcellular expression patterns that critically impact antibody selection and experimental design. In the central nervous system, CELF6 exhibits cell type-specific expression with both cytoplasmic and nuclear localization . This dual localization necessitates careful consideration when designing subcellular fractionation protocols and selecting antibodies that recognize epitopes accessible in both compartments.

When studying CELF6 in cancer cell lines such as HCT116 colorectal cancer cells or HepG2 hepatocellular carcinoma cells, researchers should account for cell cycle-dependent expression fluctuations . Antibody selection should consider these temporal dynamics, with preference for clones that maintain consistent epitope recognition regardless of potential post-translational modifications occurring throughout the cell cycle. For immunohistochemistry applications, different fixation methods may affect epitope accessibility; while the commercial antibody ab173282 is validated for paraffin-embedded tissues , optimization may be required for frozen sections or alternative fixation protocols.

Cross-reactivity with other CELF family members presents another significant consideration, as CELF3-6 share functional similarities in their impact on RNA regulation . Researchers should carefully validate antibody specificity using appropriate controls, including CELF6 knockout tissues when available . When designing multiplexed experiments to study CELF6 alongside other markers, potential spectral overlap between fluorophores should be minimized, and sequential detection protocols may be necessary when studying CELF6 in relation to other CELF family members with similar molecular weights.

How can researchers troubleshoot common issues with HRP-conjugated CELF6 antibodies in immunohistochemistry?

Immunohistochemical detection of CELF6 presents several technical challenges that require systematic troubleshooting approaches. One common issue is high background signal, which may result from non-specific binding. This can be addressed by optimizing blocking conditions (using different concentrations of BSA or normal serum from the secondary antibody species) and implementing more stringent washing steps with detergents like Tween-20 or Triton X-100 at appropriate concentrations .

Weak or absent CELF6 signal often stems from inadequate antigen retrieval, particularly critical for formalin-fixed, paraffin-embedded tissues where protein cross-linking masks epitopes. Researchers should compare heat-induced epitope retrieval methods using citrate buffer (pH 6.0) versus EDTA buffer (pH 9.0) to determine optimal conditions for their specific tissue type . For HRP-conjugated antibodies specifically, endogenous peroxidase activity can generate false positive signals, necessitating an effective quenching step (typically 0.3% H₂O₂ in methanol) prior to antibody application.

When examining CELF6 in neural tissues, consideration of its cell type-specific expression pattern is crucial; apparent "negative" results may actually reflect biological distribution rather than technical failure . For dual immunofluorescence studies, researchers should verify that HRP substrates aren't quenching fluorescent signals through photo-oxidation. Finally, when quantifying CELF6 expression, standardization of image acquisition parameters and analysis methods is essential for obtaining reproducible results, particularly important when examining subtle changes in expression levels across experimental conditions or disease states.

How do researchers interpret CELF6 expression data in the context of neurodevelopmental studies?

Interpreting CELF6 expression data in neurodevelopmental studies requires comprehensive understanding of its regulatory functions and target specificity. CELF6 predominantly binds to 3'UTRs of mRNAs coding for synaptic proteins, suggesting a critical role in synaptic development and function . When analyzing expression patterns across developmental timepoints, researchers should consider CELF6's generally repressive effect on target mRNAs, as increased CELF6 expression may correlate with decreased levels of key synaptic proteins .

Data interpretation should account for cell type-specific expression patterns of CELF6 within the CNS, as global tissue measurements may mask significant changes within specific neuronal populations . When examining CELF6 in relation to potential target genes, researchers should verify whether observed expression changes align with CELF6's established mechanism of decreasing RNA abundance in a UGU motif-dependent manner . For comprehensive understanding, researchers should integrate protein-level data (from Western blots or immunohistochemistry) with transcript-level analyses (qPCR or RNA-seq) to distinguish between transcriptional and post-transcriptional regulatory effects.

Comparative analyses with other CELF family members (particularly CELF3-5) provide valuable context, as these proteins demonstrate similar repressive functions but with potentially different magnitudes of effect . Finally, when examining CELF6 knockdown or knockout models, researchers should anticipate derepression of target genes, with particular attention to key targets such as FOS and FGF13, which have shown altered protein expression levels and localization in CELF6-deficient models .

What analytical approaches best quantify CELF6 binding affinity and specificity in different experimental contexts?

Quantifying CELF6 binding characteristics requires sophisticated analytical approaches tailored to specific experimental questions. For examining sequence specificity, massively parallel reporter assays have proven effective in testing hundreds of CELF6-bound sequence elements simultaneously . This approach allows systematic mutation of potential binding motifs (particularly UGU-rich sequences) to establish sequence-function relationships. Analysis should incorporate appropriate statistical methods to identify significant interactions between CELF6 expression and sequence elements, with p-value thresholds clearly defined (typically p<0.05) .

For quantifying binding affinity differences across experimental conditions, researchers should employ methods that capture both on-rates and off-rates. Analysis of CLIP-seq data requires sophisticated bioinformatic pipelines that account for target RNA abundance in input samples to avoid bias toward highly expressed genes . Statistical identification of true targets should incorporate controls from wild-type tissue samples to identify RNAs that interact non-specifically with capture reagents .

How can CELF6 antibodies be utilized to study protein degradation pathways in different experimental systems?

CELF6 antibodies provide valuable tools for investigating protein degradation mechanisms, as CELF6 undergoes regulated degradation via the ubiquitin-proteasome pathway . When designing experiments to study CELF6 degradation, researchers should incorporate proteasome inhibitors such as MG132, which has been demonstrated to stabilize CELF6 levels, while lysosomal inhibitors like bafilomycin A1 (BAF) or hydroxychloroquine (HCQ) show no significant effect .

For investigating ubiquitination patterns, co-immunoprecipitation protocols using CELF6 antibodies followed by immunoblotting with ubiquitin antibodies have successfully demonstrated CELF6's polyubiquitination . When examining the role of specific E3 ubiquitin ligases in CELF6 degradation, researchers should combine overexpression and silencing approaches, as demonstrated with β-TrCP, which mediates CELF6 degradation . Time-course experiments with cycloheximide chase assays allow quantification of CELF6 protein half-life under different experimental conditions, providing insights into degradation kinetics.

For studying cell cycle-dependent degradation patterns, synchronized cell populations are essential, with double-thymidine block or selective CDK1 inhibitor RO-3306 approaches successfully revealing CELF6's dynamic expression patterns . When analyzing CELF6 degradation in relation to its mRNA regulatory functions, researchers should examine the correlation between protein stability and target mRNA levels, as changes in CELF6 degradation may directly impact its regulatory targets like p21 . Finally, when comparing degradation patterns across cell types (e.g., HCT116 vs. HepG2), standardized protocols are essential for valid comparative analyses .

What are the considerations for using CELF6 antibodies in studies involving RNA-protein complex analysis?

Analysis of RNA-protein complexes involving CELF6 requires specialized methodological considerations to maintain complex integrity while achieving specific detection. When performing RNA immunoprecipitation (RIP) or CLIP experiments, standard stringent lysis and wash conditions may disrupt CELF6-RNA interactions . Researchers should optimize buffer compositions to balance between sufficient stringency to reduce non-specific binding and gentleness to preserve physiologically relevant complexes.

For detecting CELF6 within ribonucleoprotein complexes, antibody epitope accessibility may be affected by protein-protein or protein-RNA interactions. Multiple antibodies recognizing different epitopes may be necessary to comprehensively capture CELF6 in various complex configurations. When examining co-localization of CELF6 with RNA targets in intact cells, proximity ligation assays combined with in situ hybridization provide powerful visualization of specific interactions within their cellular context.

Quantitative analysis of CELF6-RNA binding should account for potential competition or cooperation with other RNA-binding proteins, as CELF6 binding sites may show enrichment for motifs recognized by different RBP families . For functional validation of CELF6-RNA interactions, reporter assays incorporating wild-type and mutated binding sequences have effectively demonstrated CELF6's repressive effect on target sequences, which is abrogated by mutation of UGU-rich motifs . Finally, when comparing CELF6 with other CELF family members (CELF3-5) in RNA complex formation, researchers should consider their similar but non-identical binding preferences and potentially different magnitudes of regulatory effect .

How can researchers address non-specific binding issues with CELF6 antibodies in complex tissue samples?

Non-specific binding presents a significant challenge when using CELF6 antibodies in complex neural tissues. Optimization of blocking conditions is critical; while 5% nonfat milk has been successfully used in published protocols , alternative blockers like BSA, casein, or commercial blocking reagents may provide superior results in specific applications. Pre-absorption of antibodies with recombinant CELF6 protein can help identify specific versus non-specific signals, particularly valuable when working with newly developed antibodies or untested tissue types.

For Western blot applications, gradient gels that provide better resolution in the 50-80 kDa range can help distinguish CELF6 from similarly sized proteins that might contribute to non-specific bands . When working with tissues expressing multiple CELF family members, validation using CELF6 knockout samples is ideal for confirming antibody specificity . For immunohistochemistry applications, inclusion of peptide competition controls, where the antibody is pre-incubated with the immunizing peptide, helps distinguish specific staining from background.

Advanced fluorescence techniques like spectral unmixing can help separate true CELF6 signal from tissue autofluorescence, particularly problematic in brain samples. For multiplexed detection, careful optimization of sequential staining protocols may be necessary to prevent cross-reactivity between multiple antibodies. Finally, when quantitative analysis is crucial, implementation of calibration standards with known quantities of recombinant CELF6 protein allows conversion of signal intensity to absolute protein amounts, facilitating more precise comparisons across experimental conditions.

What experimental designs best capture the functional consequences of CELF6-mediated RNA regulation?

Experimental designs capturing CELF6's functional impact on RNA regulation should incorporate multiple complementary approaches. Massively parallel reporter assays have proven particularly powerful, enabling simultaneous testing of hundreds of CELF6-bound sequence elements with and without mutating putative binding motifs . This approach allows systematic assessment of sequence-function relationships across numerous target UTRs.

For investigating the impact on specific target mRNAs, research designs should combine gain-of-function (CELF6 overexpression) and loss-of-function (knockout or knockdown) approaches in relevant cell types . Measurement of both mRNA and protein levels of targets is essential, as CELF6 has been shown to primarily decrease RNA abundance, which subsequently affects protein expression . For capturing dynamic regulation, time-course experiments following CELF6 manipulation provide insights into primary versus secondary effects.

To distinguish between CELF6's impacts on different RNA processing steps, experimental designs should incorporate specific assays for RNA stability (actinomycin D chase), translation efficiency (polysome profiling), and alternative splicing (exon-junction spanning PCR). For validating in vitro findings, complementary in vivo studies using CELF6 knockout mouse models have successfully demonstrated derepression of target genes, confirming CELF6's repressive function in physiological contexts . Finally, when studying tissue-specific or developmental regulation, conditional knockout or knockdown systems allow temporal and spatial control of CELF6 expression, critical for understanding its context-dependent functions.

How do post-translational modifications impact CELF6 antibody recognition and experimental outcomes?

Post-translational modifications (PTMs) significantly influence CELF6 antibody recognition, necessitating careful consideration in experimental design. CELF6 undergoes ubiquitination as part of its regulated degradation via the proteasome pathway . This modification can potentially mask epitopes, particularly those in regions containing lysine residues targeted for ubiquitination. When studying ubiquitinated CELF6, denaturation conditions in Western blotting protocols may need optimization to fully expose relevant epitopes.

CELF6's cell cycle-regulated expression pattern suggests potential phosphorylation events may also occur, as is common for proteins with dynamic expression during cell cycle progression . Phospho-specific antibodies may be valuable for investigating these regulatory events, though standard CELF6 antibodies may show varying affinity for phosphorylated versus non-phosphorylated forms. When examining samples treated with phosphatase inhibitors versus controls, researchers might observe subtle mobility shifts or intensity differences reflecting these modifications.

For comprehensive PTM analysis, complementary approaches combining immunoprecipitation with mass spectrometry provide the most detailed characterization. When studying CELF6 in different subcellular compartments, consideration of compartment-specific modifications is essential, as nuclear versus cytoplasmic CELF6 may carry distinct PTM signatures affecting antibody recognition . Finally, when comparing CELF6 across experimental conditions that might alter its PTM status (such as cell cycle synchronization or stress responses), multiple antibodies recognizing different epitopes provide the most complete detection profile, minimizing the risk of false negative results due to epitope masking.

What are the latest methodological advances in studying CELF6's role in neurodevelopmental disorders?

Recent methodological advances have significantly enhanced our understanding of CELF6's role in neurodevelopmental processes. Single-cell transcriptomics combined with CELF6 target identification has revealed cell type-specific regulatory networks, providing unprecedented resolution of CELF6's function in distinct neuronal populations . This approach helps explain how mutations affecting a broadly expressed RNA-binding protein can result in cell type-specific phenotypes.

Advanced spatial transcriptomics techniques now allow visualization of CELF6 targets in their native tissue context, revealing how CELF6-mediated regulation contributes to proper neuronal positioning and connectivity during development. For functional validation, CRISPR-Cas9 gene editing approaches enable precise modification of CELF6 binding sites within endogenous target UTRs, allowing researchers to assess the contribution of specific regulatory interactions to neuronal development and function.

Induced pluripotent stem cell (iPSC) models derived from patients with neurodevelopmental disorders provide valuable platforms for studying CELF6's role in human neurons. When combined with isogenic controls created through CRISPR correction of disease-associated variants, these models offer insights into causality rather than mere correlation. In vivo, cell type-specific and temporally controlled CELF6 manipulation using Cre-loxP systems allows dissection of developmental versus maintenance roles in neural circuits . Finally, integration of CELF6 binding data with genome-wide association studies of neurodevelopmental disorders has identified potential convergence between CELF6 regulatory networks and disease-associated genetic variants, providing new avenues for understanding pathological mechanisms.

How can researchers integrate CELF6 protein-level data with transcriptomic analyses to understand regulatory networks?

Integrating CELF6 protein data with transcriptomics requires sophisticated analytical frameworks that account for CELF6's multi-faceted regulatory roles. CLIP-seq data revealing CELF6 binding sites should be analyzed in conjunction with RNA-seq from matched samples, enabling correlation between binding events and transcript abundance . This integration should incorporate differential expression analysis comparing wild-type and CELF6-deficient conditions to distinguish direct versus indirect regulatory effects.

For detecting condition-specific regulation, integration should extend beyond simple correlation to include analysis of binding site features that might confer regulatory specificity, such as proximity to polyadenylation signals or co-occurrence with binding sites for other RBPs . When examining cell cycle-dependent regulation, synchronized cell populations allow temporal correlation between CELF6 protein levels and target transcript dynamics, revealing potential regulatory windows .

Network analysis approaches that incorporate both CELF6 binding data and transcript abundance changes can identify regulatory hubs and potential feedback mechanisms. For functional validation of predicted networks, perturbation experiments targeting key nodes should be designed, with measurement of both CELF6 binding and downstream transcript changes. Finally, integration should consider the repressive nature of CELF6 function, expecting negative correlations between CELF6 binding intensity and target transcript abundance . This comprehensive integration provides a systems-level understanding of how CELF6 coordinates complex gene expression programs in neuronal development and function.

What are the experimental considerations for studying CELF6 interactions with other RNA-binding proteins?

Studying CELF6's interactions with other RNA-binding proteins requires methodological approaches that distinguish direct protein-protein interactions from co-binding to the same RNA molecule. Sequential immunoprecipitation (IP followed by re-IP with antibodies against different RBPs) can identify protein complexes containing both CELF6 and other factors. When examining potential competition or cooperation with other RBPs, CLIP-seq data analysis should evaluate the proximity and arrangement of binding sites for different factors .

Functional studies should incorporate combinatorial manipulation of CELF6 and potential interacting RBPs, measuring the impact on shared target transcripts. Antagonistic relationships, as observed between CELF1 and MBNL1, might extend to CELF6 and warrant specific investigation . When designing in vitro binding experiments, researchers should consider that cell-specific factors may influence interaction dynamics, as the complement of RBPs differs significantly between cell types .

For high-throughput screening of potential interactions, proximity-dependent labeling approaches (BioID or APEX) coupled with mass spectrometry provide comprehensive identification of proteins in close proximity to CELF6 in living cells. When validating interactions, researchers should employ multiple complementary techniques (co-IP, proximity ligation assay, FRET) to distinguish true interactions from artifacts. Finally, computational analysis integrating binding site data for multiple RBPs can predict potential cooperative or competitive interactions based on motif arrangements and relative binding affinities, generating testable hypotheses for experimental validation.

How should researchers design experiments to capture cell type-specific functions of CELF6 in complex tissues?

Capturing cell type-specific CELF6 functions in complex tissues requires specialized experimental designs that maintain cellular context while enabling precise analysis. Single-cell approaches represent powerful strategies, with single-cell RNA-seq combined with CELF6 target identification revealing cell type-specific regulatory networks . For intact tissue analysis, laser capture microdissection followed by molecular profiling allows isolation of specific cell populations expressing CELF6 for downstream analysis.

Genetic approaches employing cell type-specific Cre drivers for conditional CELF6 manipulation (overexpression or knockout) provide powerful tools for dissecting functions in defined neuronal populations . When analyzing protein expression, multiplexed immunofluorescence combining CELF6 detection with cell type-specific markers enables quantitative analysis of expression levels across different cell populations within the same tissue section.