Customize FLS6 Antibody

Description

Molecular Identity and Function

Fas Ligand (FasL/TNFSF6) is a type II transmembrane protein belonging to the TNF superfamily. It binds to its receptor Fas (TNFRSF6/CD95) to initiate apoptosis, playing critical roles in immune regulation, T-cell homeostasis, and peripheral tolerance . Key features include:

Structure: 281 amino acids (human), with a conserved extracellular domain (81% homology to mouse FasL) .
Expression: Activated cytotoxic T cells, NK cells, and immune-privileged tissues .

Mechanistic Insights

Neutralization: Anti-FasL antibodies block FasL-induced cytotoxicity (e.g., IC₅₀ of 1–5 ng/mL for MAB126 in Jurkat cell assays) .
Therapeutic Potential: FasL pathway modulation is explored in autoimmune diseases (e.g., rheumatoid arthritis) and cancer immunotherapy . For example, Fc-optimized antibodies like FLYSYN show promise in targeting FLT3 in AML .

Clinical and Preclinical Data

Autoimmunity: FasL antibodies reduce pathogenic T-cell activity in murine models of lupus and rheumatoid arthritis .
Cancer: In gastric cancer, CAR-T cells targeting folate receptor α (FOLR1) synergize with FasL blockade to enhance tumor clearance .
Biomarker Correlation: Elevated FasL levels correlate with disease severity in interstitial lung disease (ILD) .

Validation Challenges

Antibody specificity remains a concern:

Only ~50–75% of commercial antibodies pass knockout-validated specificity tests .
Polyclonal antibodies (e.g., bs-0216R) often show broader reactivity but lower batch-to-batch consistency compared to monoclonals .

Future Directions

Engineering: Affinity maturation (e.g., site-saturation mutagenesis) improves antibody potency, as seen in antimalarial antibody CIS43_Var10 .
Multispecific Designs: Bispecific antibodies targeting FasL and checkpoint inhibitors (e.g., PD-1) are under preclinical evaluation .

Product Specs

Buffer

Preservative: 0.03% ProClin 300; Constituents: 50% Glycerol, 0.01M Phosphate Buffered Saline (PBS), pH 7.4

Form

Liquid

Lead Time

14-16 week lead time (made-to-order)

Synonyms

FLS6 antibody; At5g43935 antibody; MRH10Probable flavonol synthase 6 antibody; EC 1.14.20.6 antibody

Target Names

FLS6

Uniprot No.

F4K7D5

Q&A

FAQs for FLS6 Antibody Research

Advanced Research Questions

How to optimize FLS6 antibody effector functions for therapeutic applications?
- Approach: Engineer the Fc domain using mutations like Ser239Asp/Ile332Glu (DLE) to enhance antibody-dependent cellular cytotoxicity (ADCC) or Met428Leu/Asn434Ser (LS) to extend serum half-life .
- Validation:
  - Compare ADCC activity in in vitro reporter assays with/without mutations .
  - Measure pharmacokinetics in human FcRn transgenic mice for LS mutations .
- Key Data:
  
  Mutation Effect
  DLE 70-fold ↑ FcγRIIa affinity
  LS 3–4-fold ↑ serum half-life
How to resolve epitope masking in FLS6 antibody-based immunohistochemistry?
- Solution:
  - Pre-treat tissue sections with proteinase K or citrate buffer for antigen retrieval .
  - Use tyramide signal amplification (TSA) for low-abundance targets .
- Troubleshooting: If staining remains inconsistent, test alternative fixation methods (e.g., acetone vs. paraformaldehyde) to preserve epitope integrity .
What strategies mitigate cross-reactivity between FLS6 and homologous antigens?
- Epitope Mapping: Generate escape mutant viruses or use alanine scanning to identify critical binding residues .
- Structural Refinement: Perform X-ray crystallography or cryo-EM to guide site-directed mutagenesis for improved specificity .

Mutation	Effect
DLE	70-fold ↑ FcγRIIa affinity
LS	3–4-fold ↑ serum half-life

Methodological Considerations

How to design a robust FLS6 antibody neutralization assay?
- Protocol:
  1. Use pseudovirus particles expressing the target antigen in a luciferase-based system .
  2. Titrate FLS6 antibody (0.1–10 μg/mL) and measure neutralization efficacy via luminescence .
- Data Interpretation: Correlate neutralization titers with SPR-derived binding affinity (e.g., KD < 1 nM indicates high potency) .
How to analyze FLS6 antibody-dependent phagocytosis in macrophages?
- Assay:
  - Opsonize target cells with FLS6 antibody (10 μg/mL, 1 h).
  - Co-culture with primary macrophages (MOI 5:1) and quantify phagocytosis via imaging flow cytometry .
- Advanced Tip: Incorporate FcγRIIa-specific inhibitors (e.g., IV.3 antibody) to confirm receptor dependency .

Data Conflict Resolution

How to address discrepancies in FLS6 antibody binding across species?
- Root Cause Analysis:
  - Compare epitope conservation using Clustal Omega alignment of target antigen sequences .
  - Test antibody binding to recombinant orthologs (e.g., human vs. murine) via ELISA .
- Case Study: N6 neuraminidase antibodies showed reduced binding to H5N6 subtypes due to lateral ridge mutations .
Why does FLS6 antibody show variable protection in animal challenge models?
- Factors:
  - Dosing: Prophylactic vs. therapeutic administration (e.g., 10 mg/kg pre-infection vs. 5 mg/kg post-infection) .
  - Host immune status: Deplete CD8+ T cells to isolate antibody-mediated effects .
- Validation: Repeat studies in FcγR-humanized mice to standardize effector mechanisms .

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FLS6 Antibody