FOLR2 Antibody

What are the key considerations for selecting FOLR2 antibodies in cancer research?

Antibody selection requires evaluating:

• Epitope specificity: Monoclonal antibodies (e.g., MAB5697 targeting Gln22-His228) offer precise binding compared to polyclonal options .

• Cross-reactivity: Validate species specificity (e.g., human vs. mouse/rat) using orthogonal methods like Western blot or IHC .

• Application compatibility: Choose antibodies optimized for:

  • Flow cytometry: Optimal for cell surface detection (e.g., FOLR2+ CLL cells in co-culture systems) .

  • Immunohistochemistry (IHC): Use clones like EPR25731-70 for tissue sections .

  • Western blot: Polyclonal AF5697 detects 38 kDa FOLR2 in placental lysates under reducing conditions .

How do I validate FOLR2 antibody specificity for novel cancer models?

Implement a multi-step validation strategy:

• Positive controls: Use established FOLR2-expressing cells (e.g., neutrophils, placental tissue) to confirm staining patterns .

• Negative controls: Include FOLR2 knockout (KO) samples (e.g., CRISPR-edited NLCs) to rule out non-specific binding .

• Orthogonal verification: Cross-reference antibody data with:

  • qRT-PCR: Quantify FOLR2 mRNA levels in experimental vs. control groups .

  • Protein knockdown: Use siRNA/shRNA to correlate FOLR2 expression reductions with antibody signal loss .

How do I optimize FOLR2 detection in complex tumor microenvironments?

For studies involving trogocytosis or cellular interactions (e.g., nurse-like cell (NLC)-CLL crosstalk):

• Co-culture systems: Use time-course experiments to track FOLR2 transfer dynamics:

  • Day 0-3: Baseline FOLR2 expression on NLCs and CLL cells.

  • Day 6-10: Monitor FOLR2 acquisition by CLL cells post-activation (e.g., CD40L/IL-15) .

• Flow cytometry gating: Define stringent gates using:

  • FMO controls: Exclude false positives for FOLR2+ populations.

  • Enrichment scoring: Calculate FOLR2 enrichment scores (ES_FOLR2) to quantify trogocytic transfer .

What methodologies address conflicting FOLR2 expression data across studies?

Resolve discrepancies through:

• Antibody cross-validation: Compare results using multiple FOLR2 antibodies (e.g., MAB5697 vs. AF5697) to identify epitope-dependent variations .

• Context-dependent analysis:

  • Tissue-specific expression: FOLR2 may be upregulated in placenta/myeloid cells but downregulated in normal epithelium .

  • Cancer subtype variability: NSCLC vs. CLL may exhibit distinct FOLR2 roles (proliferation vs. survival) .

• Protocols for batch effects: Standardize:

  • Fixation methods: Formalin-fixed paraffin-embedded (FFPE) vs. fresh frozen samples.

  • Antigen retrieval: ER2 (pH8) for IHC vs. alternative buffers for Western blot .

How do I integrate FOLR2 data with functional outcomes in NSCLC models?

Combine molecular and phenotypic assays:

• Proliferation/apoptosis: Measure FOLR2 knockdown effects using:

  • Cell cycle analysis: Propidium iodide (PI) staining to detect G1/S phase arrest .

  • Apoptosis assays: Annexin-V/7-AAD staining to quantify caspase-3 activation .

• Folate metabolism: Assess intracellular folate levels post-FOLR2 silencing to link receptor function to metabolic dependencies.

Why might FOLR2 antibodies fail in Western blot?

Common issues and solutions:

How do I distinguish FOLR2 from FOLR1 in dual-expressing cells?

Employ:

• Epitope-specific antibodies: Select FOLR2-exclusive clones (e.g., MAB5697) avoiding FOLR1 cross-reactivity .

• Dual-color flow cytometry: Use anti-FOLR2 (e.g., PE-conjugated) and anti-FOLR1 (e.g., APC-conjugated) with stringent compensation.

• Gene expression profiling: Validate mRNA levels using FOLR2-specific primers in qRT-PCR .

What role does FOLR2 play in therapeutic resistance?

Investigate:

• Folate metabolism: FOLR2-mediated folate uptake may sustain cancer cell proliferation under metabolic stress .

• Targeted therapies:

  • Antibody-drug conjugates (ADCs): Exploit FOLR2 expression for selective toxin delivery.

  • CRISPR screens: Identify FOLR2 synthetic lethality partners in NSCLC or CLL models .