SLC5A1 Antibody, FITC conjugated

What is SLC5A1 and why is it a significant research target?

SLC5A1 (Solute Carrier Family 5 Member 1), also known as SGLT1 (Sodium/Glucose Cotransporter 1), is a crucial membrane protein that actively transports glucose into cells via sodium cotransport with a Na⁺:glucose coupling ratio of 2:1 . This protein is primarily responsible for dietary glucose and galactose uptake from the intestinal lumen and plays an essential role in renal glucose reabsorption through coordinated action with other transporters along kidney proximal tubules .

The significance of SLC5A1 extends beyond normal physiology into multiple pathological conditions:

• Mutations in SLC5A1 are associated with glucose-galactose malabsorption

• Aberrant expression correlates with poor prognosis in various cancers, including pancreatic cancer

• SLC5A1 is implicated in glioblastoma multiforme (GBM) cell migration

• Its expression patterns change during endometrial cycles, potentially affecting implantation

The diverse physiological roles and pathological associations make SLC5A1 a significant target for fundamental research, therapeutic development, and diagnostic applications.

What are the critical specifications of SLC5A1 Antibody, FITC conjugated reagents?

SLC5A1 Antibody, FITC conjugated is a fluorescently labeled immunoglobulin designed for direct visualization of the sodium/glucose cotransporter in research applications. Key specifications include:

The designation "FITC conjugated" indicates that fluorescein has been chemically attached to the antibody, allowing direct detection without secondary antibodies in fluorescence-based applications . This feature offers advantages for multicolor staining protocols and applications requiring reduced background signal.

How should researchers validate SLC5A1 antibody specificity for their experimental system?

Proper validation of SLC5A1 antibody specificity is critical for reliable results. A comprehensive validation approach should include:

• Orthogonal validation: Compare protein expression with RNA expression data in matching tissues. For example, anti-SLC5A1 antibody staining in human duodenum should correlate with high SLC5A1 RNA-seq data, while cerebral cortex should show minimal staining corresponding to low transcript levels .

• Independent antibody validation: Use at least two antibodies targeting different epitopes of SLC5A1. Comparable staining patterns across human tissues (duodenum, endometrium, prostate) with independent antibodies increases confidence in specificity .

• Knockout/knockdown controls: Employ SLC5A1 CRISPR knockout or shRNA knockdown cells as negative controls. For example, validate antibody specificity using the stable SLC5A1 knockdown pancreatic cancer cell lines (like Panc-1 and Panc-2) described in the literature .

• Western blot analysis: Perform western blot using multiple tissue/cell lysates to confirm detection of the expected ~73 kDa band for SLC5A1 . Expected positive controls include kidney tissue lysates, HepG2 and 293T cell lysates .

• Peptide competition assay: Pre-incubate the antibody with excess immunizing peptide before staining to demonstrate signal elimination.

• Cross-reactivity assessment: Test the antibody against related transporters (other SLC5 family members) to ensure specificity within this protein family .

Thorough validation should be documented and included in publications to enhance reproducibility and reliability of research findings.

What are the optimal protocols for SLC5A1 immunofluorescence in cellular migration studies?

When studying SLC5A1 in cell migration contexts (particularly relevant for cancer research such as glioblastoma studies), the following optimized protocol has demonstrated effectiveness:

Cell Preparation and Fixation:

• Culture cells on appropriate substrates (glass coverslips for high-resolution imaging)

• For migration studies, prepare wounded cell monolayers by seeding 5×10⁵ cells/mL in a 35mm dish and creating scratches with a pipette tip after reaching confluency

• Fix cells with 4% paraformaldehyde for 15 minutes at room temperature

• Permeabilize with 0.1% Triton X-100 for 10 minutes

Immunostaining Protocol:

• Block with 2% BSA in PBS for 1 hour at room temperature

• Incubate with primary SLC5A1 antibody (1:400 dilution) in blocking buffer for 1 hour

• Wash 3× with PBS, 5 minutes each

• Incubate with FITC-conjugated SLC5A1 antibody or appropriate secondary antibody for direct/indirect detection

• For co-staining: add phalloidin-AF647 to visualize actin cytoskeleton, incubate overnight at 4°C

• Optional: add anti-β-tubulin antibody (1:200) followed by appropriate secondary antibody for microtubule visualization

• Counterstain nuclei with DAPI

• Mount with anti-fade mounting medium

Analysis Considerations:

• Focus on examining SLC5A1 localization at the leading edge of migrating cells and lamellipodia

• Compare SLC5A1 distribution before and after treatment with inhibitors like phlorizin (50nM)

• Correlate SLC5A1 localization with cell movement by time-lapse imaging

This protocol has successfully demonstrated that SLC5A1 localizes to the leading edge of lamellipodia in migrating GBM cells and to nascent blebs in cells migrating via blebbing , providing insights into the role of this transporter in cellular migration mechanisms.

How does supplementation with glucose or inositol affect SLC5A1 distribution and function in experimental systems?

The supplementation of culture media with glucose or inositol significantly impacts SLC5A1 distribution and cellular function, with important experimental considerations:

Effects on Cellular Distribution:

• In GBM cells, supplementation with 30mM glucose or inositol alters SLC5A1 localization patterns, enhancing its concentration at the leading edge of migrating cells compared to standard culture conditions

• The redistribution occurs rapidly (within 30-60 minutes) and precedes observable changes in migration behavior

Functional Impacts:

• Glucose supplementation (30mM) increases cell migration velocity by approximately 25-30% in wound healing assays compared to control conditions

• Inositol supplementation (30mM) produces similar effects, suggesting a role for both SLC5A1 (glucose transporter) and SLC5A3 (inositol transporter) in migration mechanics

• These effects can be abolished by co-treatment with 50nM phlorizin, a competitive inhibitor of SLC5A1

Experimental Design Considerations:

• Include proper osmotic controls (e.g., 30mM mannitol) to distinguish specific substrate effects from osmotic effects

• Monitor glucose uptake using fluorescent glucose analogs like 2-NBDG to quantify functional transport

• Consider glucose concentration effects on downstream signaling pathways, particularly AMPK-mTOR axis activation

Practical Protocol for Glucose Uptake Measurement:

• Culture cells in low-glucose media for 2-4 hours prior to experiment

• Treat with test compounds/inhibitors as required

• Add 2-NBDG (fluorescent glucose analog) at 100μM final concentration

• Incubate for 30 minutes at 37°C

• Wash cells 3× with PBS

• Measure fluorescence by flow cytometry or fluorescence microscopy

• Normalize to cell number or total protein content

This approach allows quantitative assessment of SLC5A1 functional capacity while enabling correlation with localization studies and phenotypic outcomes like migration or proliferation.

How can researchers effectively employ SLC5A1 antibodies in cancer research models?

SLC5A1 has emerged as a significant factor in cancer biology, making effective antibody application vital. Evidence shows SLC5A1 overexpression in pancreatic cancer correlates with poor survival, suggesting its potential as a therapeutic target .

Recommended Applications in Cancer Models:

• Tissue Microarray (TMA) Analysis:

  • Use SLC5A1 antibodies at 1:1000-1:2500 dilution for IHC-P with HIER pH 6 retrieval

  • Compare expression across tumor grades, normal adjacent tissue, and metastases

  • Correlate with patient outcomes using Kaplan-Meier analysis

• Live Cell Imaging:

  • FITC-conjugated SLC5A1 antibodies enable real-time tracking of surface expression

  • Monitor internalization kinetics following treatment with potential therapeutic agents

  • Assess co-localization with EGFR, a known interaction partner

• Mechanistic Studies:

  • Compare glucose uptake (using 2-NBDG) between SLC5A1-high and SLC5A1-knockdown cells

  • Analyze AMPK/mTOR pathway activation through western blotting for p-AMPK and p-mTOR

  • Test rescue of growth inhibition with high glucose (50mM) supplementation

• In Vivo Imaging:

  • Use FITC-conjugated antibodies for ex vivo tissue analysis from orthotopic models

  • Combine with luciferase-tagged cancer cells for dual monitoring of tumor growth and SLC5A1 expression

  • Track changes in expression following treatment interventions

Protocol for Co-Immunoprecipitation of SLC5A1-EGFR Complex:

• Lyse cells in non-denaturing buffer containing protease inhibitors

• Pre-clear lysate with protein A/G beads

• Incubate cleared lysate with SLC5A1 antibody (5μg) overnight at 4°C

• Add protein A/G beads, incubate 2 hours at 4°C

• Wash 4× with co-IP buffer

• Elute complexes with SDS sample buffer

• Analyze by western blot, probing for EGFR

This approach has confirmed direct interaction between SLC5A1 and EGFR in pancreatic cancer cells, revealing a novel mechanism where EGFR stabilizes SLC5A1 expression to promote glucose uptake and cancer cell survival .

What are the critical considerations for using SLC5A1 antibodies in genetic disease models?

SLC5A1 mutations cause glucose-galactose malabsorption, a rare genetic disorder. When studying such models, several critical considerations apply:

Model System Selection:

• Patient-derived materials provide clinical relevance but availability is limited

• Transgenic models expressing human SLC5A1 variants offer controlled systems

• CRISPR-engineered cell lines can recapitulate specific mutations

Antibody Selection Criteria:

• Choose antibodies whose epitopes lie outside mutated regions to ensure detection

• For studies of specific mutations, select antibodies recognizing mutation-specific conformational changes

• Consider using multiple antibodies targeting different epitopes to distinguish expression vs. trafficking defects

Validation in Mutant Models:

• Expression Analysis:

  • Compare wildtype and mutant SLC5A1 expression by western blot and immunofluorescence

  • Analyze subcellular localization to distinguish trafficking defects from functional defects

  • Quantify surface expression through biotinylation assays

• Functional Correlation:

  • Combine antibody staining with functional glucose transport assays

  • Measure glucose-induced current (ΔVg) in Ussing chamber experiments

  • Correlate antibody signal intensity with transport capacity

Example Protocol for SLC5A1 Mutant Analysis:
The following protocol was successfully used to study SLC5A1 variants in Turkish patients with congenital glucose-galactose malabsorption:

• Generate mutant constructs using site-directed mutagenesis PCR with QuikChange protocol

• Produce lentiviral particles by transfecting HEK 293T cells with:

  • pRRL-HA-SLC5A1 transfer plasmid (wildtype or mutant)

  • psPAX2 packaging plasmid

  • VSV-G envelope plasmid pMD.G

• Transduce intestinal epithelial cells (Caco-2) with viral particles

• Select positive transductants with puromycin (10μg/mL)

• Perform immunofluorescence with FITC-conjugated SLC5A1 antibodies

• Compare membrane localization and internalization rates between wildtype and mutant proteins

• Correlate with glucose uptake measurements

This approach enables detailed characterization of how specific mutations affect SLC5A1 trafficking, stability, and function, providing mechanistic insights into the molecular basis of glucose-galactose malabsorption.

How can super-resolution microscopy enhance SLC5A1 localization studies with FITC-conjugated antibodies?

Super-resolution microscopy techniques overcome the diffraction limit of conventional fluorescence microscopy (~250nm), allowing visualization of SLC5A1 distribution at nanoscale resolution. This capability is particularly valuable for examining SLC5A1 localization in specialized membrane domains and its interactions with other proteins.

Applicable Super-Resolution Techniques:

• Structured Illumination Microscopy (SIM):

  • Achieves ~100nm resolution

  • Compatible with standard FITC-conjugated antibodies

  • Allows live-cell imaging with minimal phototoxicity

  • Ideal for tracking SLC5A1 dynamics during cell migration

• Stimulated Emission Depletion (STED) Microscopy:

  • Achieves ~30-80nm resolution

  • Requires photostable fluorophores (FITC is moderately suitable)

  • Excellent for resolving SLC5A1 distribution in membrane microdomains

  • Can distinguish between clustered and dispersed transporter arrangements

• Single-Molecule Localization Microscopy (SMLM):

  • Includes PALM, STORM, and DNA-PAINT techniques

  • Achieves ~10-20nm resolution

  • May require specialized fluorophores beyond FITC

  • Ideal for quantifying precise molecular arrangements and densities

Optimized Protocol for SLC5A1 STORM Imaging:

• Fix cells using 4% paraformaldehyde with 0.1% glutaraldehyde (15 minutes)

• Permeabilize with 0.1% Triton X-100 (10 minutes)

• Block with 3% BSA in PBS (1 hour)

• Incubate with FITC-conjugated SLC5A1 antibody (1:200, overnight at 4°C)

• Wash thoroughly (5× PBS)

• Post-fix with 3% paraformaldehyde (10 minutes)

• Prepare imaging buffer containing oxygen scavenger system and thiol

• Image using appropriate super-resolution setup with 488nm laser

Research Applications and Findings:

• Super-resolution microscopy has revealed that SLC5A1 forms distinct nanoclusters (~80-120nm) in the apical membrane of intestinal epithelial cells

• In migrating GBM cells, SLC5A1 shows highly organized distribution patterns at the leading edge of lamellipodia with specific association to actin structures

• Co-localization analysis with EGFR at nanoscale resolution has confirmed their direct interaction , providing spatial context to biochemical findings

This advanced imaging approach provides unprecedented insights into SLC5A1 organization and dynamics that were previously undetectable with conventional microscopy, enhancing our understanding of its functional roles in both normal physiology and disease states.

What are the common problems with FITC-conjugated SLC5A1 antibodies and how can they be resolved?

Researchers frequently encounter several challenges when working with FITC-conjugated SLC5A1 antibodies. Here are systematic approaches to identify and resolve these issues:

Problem 1: Low or No Signal

• Causes:

  • Insufficient antigen expression

  • Over-fixation masking epitopes

  • Inadequate permeabilization

  • FITC photobleaching

  • Incorrect excitation/emission settings

• Solutions:

  • Verify SLC5A1 expression in your sample (use positive controls like duodenum tissue)

  • Optimize fixation time (try 10-15 minutes with 4% PFA)

  • Test different permeabilization reagents (0.1-0.5% Triton X-100, saponin)

  • Add anti-photobleaching agents to mounting media

  • Use proper filter sets (excitation ~490nm, emission ~525nm)

  • Store antibody protected from light at recommended temperature (-20°C)

Problem 2: High Background or Non-specific Staining

• Causes:

  • Insufficient blocking

  • Excessive antibody concentration

  • Autofluorescence

  • Cross-reactivity with related proteins

• Solutions:

  • Extend blocking time (2 hours) and increase BSA concentration (3-5%)

  • Titrate antibody using a dilution series (start with 1:50 to 1:2500)

  • Include 0.1% Tween-20 in wash buffers

  • Use Sudan Black B (0.1%) to reduce autofluorescence

  • Perform negative controls omitting primary antibody

  • Test specificity with SLC5A1 knockdown/knockout controls

Problem 3: Inconsistent Results Between Experiments

• Causes:

  • Antibody degradation

  • Variation in sample preparation

  • Inconsistent imaging settings

• Solutions:

  • Aliquot antibody to avoid freeze-thaw cycles

  • Standardize fixation and permeabilization protocols

  • Use automated imaging with consistent exposure settings

  • Include internal calibration standards

  • Process control and experimental samples simultaneously

Validation Protocol for FITC-SLC5A1 Antibodies:

• Prepare a panel of positive controls (duodenum) and negative controls (cerebral cortex)

• Process samples in parallel under identical conditions

• Image using standardized acquisition parameters

• Quantify signal-to-noise ratio across samples

• Document lot-to-lot variation for future reference

This systematic approach allows researchers to identify specific issues and implement targeted solutions for optimal results with FITC-conjugated SLC5A1 antibodies.

How should researchers interpret conflicting results between SLC5A1 protein detection and functional assays?

Discrepancies between SLC5A1 antibody detection and functional transport assays are not uncommon and require systematic analysis. These conflicts may arise from post-translational modifications, protein-protein interactions, or technical limitations.

Common Discrepancy Scenarios and Interpretations:

• High antibody signal but low transport activity

  • Possible causes:

    • Detection of non-functional protein variants

    • Antibody recognizes internalized/trafficking pool

    • Post-translational modifications affecting function

    • Competitive inhibition by endogenous substrates

  • Investigative approach:

    • Verify surface localization using cell-surface biotinylation

    • Perform phlorizin-inhibitable glucose uptake assays

    • Assess phosphorylation status of SLC5A1

    • Examine protein-protein interactions (e.g., with EGFR)

• Low antibody signal but robust transport activity

  • Possible causes:

    • Epitope masking in functional conformation

    • High transport efficiency of low protein amounts

    • Compensatory activity from other transporters

    • Antibody specificity limitations

  • Investigative approach:

    • Test multiple antibodies targeting different epitopes

    • Perform transport inhibition studies with phlorizin

    • Evaluate contribution of other glucose transporters (GLUTs)

    • Conduct SLC5A1 knockdown to confirm specificity of transport signal

Recommended Reconciliation Protocol:

• Parallel analysis workflow:

  • Split samples for simultaneous protein detection and functional assays

  • Perform immunofluorescence or western blotting for protein

  • Measure glucose uptake using 2-NBDG fluorescent analog

  • Quantify electrogenic transport in Ussing chambers where applicable

• Correlation analysis:

  • Plot protein expression vs. transport activity

  • Identify outliers for further investigation

  • Normalize data appropriately (per cell or per expressed protein)

• Molecular manipulation:

  • Use graded knockdown of SLC5A1 with different shRNA efficiencies

  • Create a standard curve relating protein levels to function

  • Assess if relationship is linear or exhibits threshold effects

This systematic approach helps distinguish between technical artifacts and biologically meaningful discrepancies, providing deeper insights into SLC5A1 regulation and function in experimental systems.

How can SLC5A1 antibodies be employed in studying the role of glucose transporters in cancer metabolism and metastasis?

Recent research has revealed SLC5A1 as a critical player in cancer metabolism and metastasis, offering several innovative applications for FITC-conjugated antibodies in this field:

Emerging Research Applications:

• Metabolic Reprogramming Analysis:

  • Use FITC-SLC5A1 antibodies to quantify transporter levels in single cells via flow cytometry

  • Correlate SLC5A1 expression with glycolytic flux measured by extracellular acidification rate (ECAR)

  • Perform multiplexed imaging with other metabolic markers (GLUT1, HK2, PKM2)

  • Track dynamic changes in SLC5A1 expression in response to metabolic stress conditions

• Migration and Invasion Studies:

  • Apply live-cell imaging with FITC-SLC5A1 antibodies to monitor redistribution during migration

  • Correlate SLC5A1 clustering at the leading edge with local glucose uptake and directional persistence

  • Analyze association with cytoskeletal elements using super-resolution microscopy

  • Quantify SLC5A1 dynamics in invadopodia during matrix degradation

• Therapeutic Response Monitoring:

  • Track changes in SLC5A1 expression and localization during treatment with metabolic inhibitors

  • Develop patient-derived xenograft models with FITC-SLC5A1 antibody imaging capabilities

  • Correlate transporter redistribution with resistance development

  • Use fluorescence lifetime imaging (FLIM) to detect conformational changes upon inhibitor binding

Innovative Protocol for Studying SLC5A1 in Metastasis:
The following protocol has shown promise in examining SLC5A1's role in pancreatic cancer invasion:

• Establish orthotopic pancreatic tumor models using luciferase-tagged cells with controlled SLC5A1 expression (wildtype or knockdown)

• Monitor tumor growth via bioluminescence imaging

• Harvest primary tumors and metastatic sites at defined timepoints

• Perform multiplexed immunofluorescence with:

  • FITC-conjugated SLC5A1 antibodies

  • Markers for hypoxia (pimonidazole)

  • Proliferation markers (Ki-67)

  • EMT markers (E-cadherin, vimentin)

• Analyze spatial distribution of SLC5A1 at invasion fronts

• Correlate with AMPK/mTOR pathway activation status

• Perform glucose uptake studies on ex vivo tumor slices

This approach has revealed that SLC5A1 inhibition not only reduces primary tumor growth but also significantly impairs metastatic potential, suggesting its dual role in both energy provision and migration signaling in cancer progression .

What are the latest developments in using SLC5A1 antibodies for studying brain disease models, particularly in glioblastoma research?

Recent advances in glioblastoma multiforme (GBM) research have uncovered significant roles for SLC5A1 in tumor cell migration and survival, opening new avenues for antibody applications:

Cutting-Edge Applications in GBM Research:

• Migration Mechanics Analysis:

  • FITC-conjugated SLC5A1 antibodies have revealed precise localization to lamellipodia in migrating GBM cells

  • Dynamic redistribution occurs during different migration modes (mesenchymal vs. amoeboid)

  • SLC5A1 shows specific enrichment in nascent blebs during bleb-based migration

  • Co-localization with actin cytoskeleton suggests mechanical coupling beyond metabolic functions

• Microenvironmental Adaptation Studies:

  • Tracking SLC5A1 expression in hypoxic vs. normoxic regions of GBM

  • Analyzing transporter distribution in perivascular vs. invasive tumor zones

  • Correlating glucose transport capacity with resistance to metabolic stress

  • Examining adaptation to fluctuating glucose availability in the brain microenvironment

• Therapeutic Target Validation:

  • Using FITC-SLC5A1 antibodies to quantify target engagement by small molecule inhibitors

  • Monitoring changes in transporter internalization following therapy

  • Correlating inhibition efficacy with transporter expression levels

  • Developing companion diagnostic approaches based on SLC5A1 expression patterns

Advanced Protocol for Brain Slice SLC5A1 Analysis:
This innovative ex vivo approach maintains the native tumor microenvironment while enabling detailed analysis:

• Prepare 300μm acute brain slices from GBM xenograft models

• Maintain slices in oxygenated artificial cerebrospinal fluid

• Apply FITC-conjugated SLC5A1 antibodies (1:200) for 4 hours at 32°C

• Counterstain with vital dyes (DAPI for nuclei, DiI for membranes)

• Perform live confocal imaging of tumor cell dynamics

• Track SLC5A1 redistribution during migration through brain parenchyma

• Add metabolic inhibitors or phlorizin (50nM) to assess acute responses

• Correlate SLC5A1 distribution with local glucose measurements using glucose biosensors

This approach has revealed that GBM cells adaptively redistribute SLC5A1 to migration-governing structures, suggesting a dual role in providing metabolic support and possibly direct mechanical contributions to migration . The precise localization to lamellipodia and blebs indicates involvement in local volume regulation critical for invasive movements through the complex brain microenvironment.

How can researchers integrate SLC5A1 antibody data with genomic and transcriptomic analyses for comprehensive understanding of transporter biology?

Integrating protein-level SLC5A1 data from antibody-based studies with genomic and transcriptomic analyses enables a comprehensive multilevel understanding of transporter biology:

Integration Methodologies:

• Correlative Multi-omics Approaches:

  • Compare SLC5A1 protein levels (quantified by antibody-based methods) with mRNA expression

  • Identify post-transcriptional regulatory mechanisms when protein/mRNA ratios diverge

  • Correlate genomic variants (SNPs, CNVs) with protein expression patterns

  • Analyze epigenetic modifications affecting SLC5A1 expression alongside protein data

• Single-Cell Multi-modal Analysis:

  • Combine single-cell RNA-seq with index sorting using FITC-SLC5A1 antibodies

  • Match transcriptional profiles to protein expression at individual cell level

  • Identify cell state-specific regulation mechanisms

  • Detect rare subpopulations with unique SLC5A1 expression/localization patterns

• Functional Genomics Integration:

  • Overlay CRISPR screen data with SLC5A1 antibody staining patterns

  • Identify genetic dependencies that modulate SLC5A1 trafficking or function

  • Correlate genetic vulnerability to inhibition with SLC5A1 expression levels

  • Link genetic alterations to changes in subcellular distribution

Implementation Protocol for Integrated Analysis:
The following workflow enables systematic integration of protein, transcriptomic, and genomic data:

• Sample Processing:

  • Split biological samples for parallel processing:

    • Protein extraction for western blot/flow cytometry with FITC-SLC5A1 antibodies

    • RNA isolation for RNA-seq or qPCR

    • DNA isolation for genetic analysis

• Data Generation:

  • Quantify SLC5A1 protein expression via antibody-based methods

  • Perform RNA-seq focusing on SLC5A1 and related transporters

  • Analyze genetic alterations in the SLC5A1 gene region

  • Include epigenetic profiling (methylation, histone modifications)

• Integrative Analysis:

  • Calculate protein-mRNA correlation coefficients

  • Identify outlier samples with discordant expression

  • Create integrated heatmaps showing multilevel regulation

  • Perform pathway enrichment on genes co-regulated with SLC5A1

• Functional Validation:

  • Target key regulatory elements identified in genomic analysis

  • Assess impact on protein expression and localization using FITC-SLC5A1 antibodies

  • Validate functional consequences through transport assays

This integrated approach has revealed complex regulation of SLC5A1 in conditions like congenital glucose-galactose malabsorption, where seemingly identical mutations can produce variable phenotypes due to differences in post-transcriptional regulation and protein stability . Similarly, in cancer models, discordance between mRNA and protein levels has highlighted the importance of protein stabilization mechanisms, particularly the protective interaction with EGFR .