SLC7A7 Antibody

What is SLC7A7 and why is it important in biological research?

SLC7A7 is a transporter protein that functions as a heterodimer with SLC3A2 to regulate the transport of cationic amino acids. It operates as an antiporter, exporting cationic amino acids from inside cells in exchange with neutral amino acids plus sodium ions . This protein plays a critical role in:

• Balancing intracellular and extracellular concentrations of specific amino acids

• Serving as building blocks for protein synthesis

• Acting as precursors for various metabolic processes

• Potentially participating in nitric oxide synthesis via the transport of L-arginine

The protein is essential for the correct function of non-polarized cells such as monocytes, making it a significant target for immunological research . Recent studies have also identified SLC7A7 as a potential prognostic biomarker correlated with immune infiltrates in certain cancers, particularly non-small cell lung cancer (NSCLC) .

What are the common synonyms and alternate names for SLC7A7?

When conducting literature searches or ordering antibodies, researchers should be aware of the following alternative names for SLC7A7:

• Y+L amino acid transporter 1

• Monocyte amino acid permease 2 (MOP-2)

• Solute carrier family 7 member 7

• y(+)L-type amino acid transporter 1

• Y+LAT1 or y+LAT-1

Understanding these alternative designations is important when searching literature databases and identifying relevant antibodies for your research.

What criteria should researchers consider when selecting an SLC7A7 antibody?

When selecting an SLC7A7 antibody for research purposes, consider:

• Species reactivity: Verify that the antibody recognizes SLC7A7 in your species of interest. Common commercially available antibodies recognize human and mouse SLC7A7 .

• Applications validated: Ensure the antibody has been validated for your specific application:

  • Western blot (WB)

  • Immunohistochemistry (IHC-P)

  • Immunocytochemistry/Immunofluorescence (ICC/IF)

  • ELISA

• Immunogen information: Review the immunogen used to generate the antibody. For example, some antibodies are generated against recombinant fragments corresponding to human SLC7A7 amino acids 300-400 or 325-382 .

• Clonality: Determine whether a polyclonal or monoclonal antibody is more suitable for your application. Polyclonal antibodies may offer higher sensitivity but potentially lower specificity compared to monoclonals.

• Independent validation: Look for antibodies that have been cited in peer-reviewed publications, indicating successful use in research settings .

How should researchers validate an SLC7A7 antibody before experimental use?

Proper antibody validation is critical to ensure experimental reliability:

• Positive and negative controls:

  • Use tissue or cell lines known to express or lack SLC7A7

  • Consider using NSCLC cell lines as positive controls, as SLC7A7 has been well-studied in this context

• Knockdown/knockout validation:

  • Perform siRNA knockdown or CRISPR knockout of SLC7A7 and confirm reduced antibody signal

  • This approach controls for potential non-specific binding

• Multiple detection methods:

  • Confirm results using at least two different techniques (e.g., WB and IHC)

  • Compare results with mRNA expression data where available

• Titration experiments:

  • Perform antibody dilution series to determine optimal concentration

  • This minimizes background while maintaining specific signal

• Peptide competition assay:

  • Pre-incubate antibody with the immunizing peptide

  • Specific binding should be blocked in the presence of excess peptide

What are the optimal conditions for using SLC7A7 antibodies in Western blotting?

For optimal Western blot results with SLC7A7 antibodies:

• Sample preparation:

  • Use RIPA buffer with protease inhibitors for protein extraction

  • Include phosphatase inhibitors if studying potential phosphorylation states

• Protein loading and separation:

  • Load 20-50 μg of total protein per lane

  • SLC7A7 has a predicted molecular weight of approximately 50-55 kDa

  • Use 10-12% SDS-PAGE gels for optimal resolution

• Transfer conditions:

  • Transfer to PVDF membrane (preferred over nitrocellulose for this protein)

  • Use wet transfer at 100V for 1 hour or 30V overnight at 4°C

• Blocking and antibody incubation:

  • Block with 5% non-fat dry milk in TBST for 1 hour at room temperature

  • Incubate with primary antibody at manufacturer's recommended dilution (typically 1:1000-1:2000) overnight at 4°C

  • For rabbit polyclonal antibodies, incubate with HRP-conjugated anti-rabbit secondary antibody (typically 1:5000-1:10000) for 1 hour at room temperature

• Detection:

  • Use enhanced chemiluminescence (ECL) detection system

  • Exposure time typically ranges from 30 seconds to 5 minutes depending on expression level

What protocols are recommended for SLC7A7 immunohistochemistry in FFPE tissues?

For immunohistochemical detection of SLC7A7 in formalin-fixed paraffin-embedded (FFPE) tissues:

• Tissue processing:

  • Cut 5 μm thickness sections and place on poly-L-lysine coated glass slides

  • Deparaffinize by placing slides in a 60°C oven for 30 minutes

  • Rehydrate through graded ethanol series (100%, 95%, 70%)

• Antigen retrieval:

  • Critical step for SLC7A7 detection

  • Immerse sections in sodium citrate buffer (pH 6.0)

  • Heat in pressure cooker or microwave until boiling, then maintain at sub-boiling temperature for 10-20 minutes

  • Cool to room temperature for 20 minutes

• Endogenous peroxidase blocking:

  • Incubate sections with 3% H₂O₂ in methanol for 15 minutes

  • This quenches endogenous peroxidase activity

• Antibody incubation:

  • Block with 5% normal goat serum for 1 hour at room temperature

  • Incubate with SLC7A7 primary antibody at 1:100 dilution overnight at 4°C in a humidified chamber

  • Wash thoroughly with 0.3% PBST

  • Incubate with secondary goat anti-rabbit antibody for 2 hours at room temperature

• Detection and visualization:

  • Develop with DAB (3,3'-Diaminobenzidine) chromogen kit

  • Counterstain lightly with hematoxylin to reveal cell nuclei

  • Dehydrate, clear, and mount with permanent mounting medium

How is SLC7A7 expression linked to prognosis in cancer, particularly NSCLC?

SLC7A7 has emerged as a significant prognostic marker in cancer research:

Researchers investigating SLC7A7 as a prognostic marker should consider these seemingly paradoxical findings and design experiments that can elucidate the underlying mechanisms.

What is the relationship between SLC7A7 expression and immune cell infiltration in NSCLC?

SLC7A7 has been found to have significant correlations with immune infiltration in NSCLC:

• Correlation with immune cell populations:

  • SLC7A7 expression positively correlates with infiltrating levels of multiple immune cell types in lung adenocarcinoma (LUAD):

    • B cells (r = 0.281, P = 3.27E−10)

    • CD8+ T cells (r = 0.377, P = 6.16E−18)

    • CD4+ T cells (r = 0.393, P = 2.7E−19)

    • Macrophages (r = 0.637, P = 1.31E−56)

    • Neutrophils (r = 0.71, P = 2.12E−75)

    • Dendritic cells (r = 0.717, P = 4.17E−78)

• Cell type-specific expression:

  • Single-cell analyses have shown that SLC7A7 is highly expressed in:

    • Monocytes (both non-classical and classical)

    • Naïve B cells

• Differential correlation patterns:

  • Similar positive correlations were observed in lung squamous cell carcinoma (LUSC)

  • In contrast, SLC7A7 expression was not significantly correlated with tumor purity or immune infiltration in uveal melanoma (UVM)

These findings suggest that SLC7A7 plays an important role in regulating the tumor immune microenvironment specifically in NSCLC, making it a potential target for immunotherapy research.

How does SLC7A7 influence T cell exhaustion and regulatory T cell function in the tumor microenvironment?

Research has revealed complex relationships between SLC7A7 and T cell function:

• T cell exhaustion markers:

  • SLC7A7 expression shows strong positive correlations with exhaustion markers including:

    • TIM-3 (T cell immunoglobulin and mucin domain-containing protein 3)

    • PD-1 (Programmed cell death protein 1)

    • CTLA4 (Cytotoxic T-lymphocyte-associated protein 4)

    • LAG3 (Lymphocyte activation gene 3)

  • The particularly strong correlation with TIM-3 suggests SLC7A7 may play a crucial role in TIM-3-mediated T cell exhaustion

• Regulatory T cell (Treg) association:

  • SLC7A7 expression positively correlates with Treg markers:

    • FOXP3 (Forkhead box P3)

    • CCR8 (C-C motif chemokine receptor 8)

    • STAT5B (Signal transducer and activator of transcription 5B)

    • TGFB1 (Transforming growth factor beta 1)

  • This suggests SLC7A7 may contribute to immunosuppression by enhancing Treg function

• T helper cell regulation:

  • SLC7A7 expression significantly correlates with markers of various T helper subsets:

    • Th1 cells

    • Th2 cells

    • Tfh (T follicular helper) cells

    • Th17 cells

  • These correlations indicate SLC7A7 may broadly influence T cell differentiation and function

Researchers interested in cancer immunotherapy should consider investigating how modulation of SLC7A7 might impact T cell exhaustion and regulatory mechanisms in the tumor microenvironment.

What role does SLC7A7 play in macrophage polarization and function?

SLC7A7's involvement in macrophage biology offers intriguing research opportunities:

Researchers studying macrophage biology in cancer and inflammation should consider targeting SLC7A7 to better understand its role in macrophage polarization and function.

What are common issues when using SLC7A7 antibodies and how can they be resolved?

Researchers may encounter several challenges when working with SLC7A7 antibodies:

• High background in Western blots:

  • Cause: Insufficient blocking or excessive antibody concentration

  • Solution:

    • Increase blocking time (2-3 hours at room temperature)

    • Use 5% BSA instead of milk for blocking

    • Further dilute primary antibody

    • Include 0.05% Tween-20 in all wash and antibody incubation steps

• Weak or no signal in immunohistochemistry:

  • Cause: Inadequate antigen retrieval or epitope masking

  • Solution:

    • Test multiple antigen retrieval methods (citrate buffer pH 6.0, EDTA buffer pH 9.0)

    • Extend antigen retrieval time

    • Consider using amplification systems (e.g., tyramide signal amplification)

    • Test different antibody clones that recognize different epitopes

• Multiple bands in Western blot:

  • Cause: Potential post-translational modifications, degradation products, or non-specific binding

  • Solution:

    • Include protease inhibitor cocktail during sample preparation

    • Compare band patterns with literature or manufacturer data

    • Use positive and negative controls to identify specific bands

    • Perform peptide competition assay to identify specific bands

• Variability between experiments:

  • Cause: Inconsistent protocol execution or antibody degradation

  • Solution:

    • Standardize all protocol steps carefully

    • Aliquot antibodies to avoid freeze-thaw cycles

    • Include internal controls in each experiment

    • Use automated systems where possible to reduce variability

How can researchers optimize detection of SLC7A7 in different subcellular compartments?

SLC7A7 predominantly localizes to the plasma membrane, but can also be found in intracellular compartments. To optimize detection in different subcellular locations:

• For plasma membrane localization:

  • Immunofluorescence optimization:

    • Use non-permeabilizing conditions initially (fix with 4% paraformaldehyde without detergent)

    • Add mild permeabilization (0.1% Triton X-100 for 5 minutes) if needed

    • Co-stain with plasma membrane markers (e.g., Na+/K+ ATPase)

  • Subcellular fractionation for Western blot:

    • Use membrane protein extraction kits specifically designed to isolate plasma membrane proteins

    • Compare with total cell lysates to assess relative distribution

• For intracellular compartment detection:

  • Immunofluorescence approach:

    • Use standard permeabilization (0.2-0.5% Triton X-100)

    • Co-stain with markers for specific compartments (e.g., EEA1 for early endosomes)

    • Consider confocal microscopy for precise localization

  • Biochemical approach:

    • Perform subcellular fractionation to isolate different cellular compartments

    • Use specific markers for each fraction to confirm separation quality

    • Quantify SLC7A7 distribution across fractions

• For trafficking studies:

  • Live cell imaging:

    • Consider using cells expressing SLC7A7 tagged with fluorescent proteins

    • Monitor trafficking in response to various stimuli

  • Pulse-chase experiments:

    • Use surface biotinylation followed by internalization assays

    • Quantify rates of endocytosis and recycling

These approaches can be particularly valuable when studying how SLC7A7 localization changes in response to disease states or experimental manipulations.

What are promising areas for future investigation of SLC7A7 in cancer immunotherapy?

Several promising research directions emerge from current SLC7A7 knowledge:

• Targeting SLC7A7 to modulate T cell exhaustion:

  • Investigate whether inhibiting or enhancing SLC7A7 function affects expression of exhaustion markers

  • Determine if combining SLC7A7 modulation with immune checkpoint inhibitors (anti-PD-1, anti-CTLA4) enhances efficacy

  • Explore mechanisms connecting amino acid transport to T cell exhaustion pathways

• SLC7A7's role in metabolic reprogramming of immune cells:

  • Study how SLC7A7-mediated amino acid transport affects metabolic profiles of tumor-infiltrating immune cells

  • Investigate connections between SLC7A7 activity and immune cell energetics (glycolysis vs. oxidative phosphorylation)

  • Determine if metabolic interventions targeting amino acid utilization can synergize with immunotherapies

• Development of SLC7A7 as a biomarker for immunotherapy response:

  • Conduct retrospective and prospective studies correlating SLC7A7 expression with response to immune checkpoint inhibitors

  • Develop standardized assays for measuring SLC7A7 in clinical samples

  • Investigate whether SLC7A7 expression in specific immune cell populations has stronger predictive value

• Therapeutic targeting of SLC7A7:

  • Develop small molecule inhibitors or activators of SLC7A7

  • Test cell-type specific delivery approaches to target SLC7A7 modulation to specific immune populations

  • Investigate antibody-drug conjugates targeting cells with aberrant SLC7A7 expression

How might single-cell analysis techniques advance our understanding of SLC7A7 function?

Single-cell technologies offer powerful approaches to unraveling SLC7A7 biology:

• Single-cell RNA sequencing applications:

  • Profile SLC7A7 expression across all cell types in the tumor microenvironment

  • Identify co-expression patterns to discover functional networks associated with SLC7A7

  • Track changes in SLC7A7 expression during disease progression or therapeutic intervention

• Single-cell proteomics approaches:

  • Measure SLC7A7 protein levels and post-translational modifications at single-cell resolution

  • Correlate SLC7A7 protein expression with cellular phenotypes and functional states

  • Identify protein interaction networks involving SLC7A7 in specific cell types

• Spatial transcriptomics and proteomics:

  • Map SLC7A7 expression patterns within the spatial context of tumors

  • Identify spatial relationships between SLC7A7-expressing cells and other features of the tumor microenvironment

  • Correlate spatial distribution with functional outcomes and patient prognosis

• Multi-omics integration:

  • Combine single-cell RNA-seq, ATAC-seq, and proteomics to comprehensively understand SLC7A7 regulation

  • Connect genetic variants affecting SLC7A7 to functional consequences

  • Develop predictive models of how SLC7A7 contributes to disease phenotypes

These advanced approaches could help resolve the seemingly paradoxical findings regarding SLC7A7 expression and function in different contexts, potentially leading to more precise therapeutic strategies.