BRAF (Phospho-Ser446) Antibody is a rabbit polyclonal antibody designed to recognize endogenous BRAF protein only when phosphorylated at Ser446 . This phosphorylation event is associated with BRAF's activation and regulation within the MAPK/ERK signaling pathway, which governs cell proliferation, differentiation, and survival .
Key Features:
Immunogen: Synthetic peptide spanning residues 411–460 of human BRAF, including the phosphorylated Ser446 site .
Specificity: Detects BRAF phosphorylation at Ser446 with no cross-reactivity to non-phosphorylated or adjacent phosphorylated residues (e.g., Ser445 or Ser447) .
This antibody is widely used in multiple experimental workflows:
Storage and Handling:
Formulation: Liquid in PBS with 50% glycerol, 0.5% BSA, and 0.02% sodium azide .
Stability: Stable for 12 months at -20°C; avoid freeze-thaw cycles .
Phosphorylation at Ser446 modulates BRAF’s interaction with other kinases (e.g., Raf-1) and regulatory proteins, influencing MAPK pathway output .
In oncogenic Ras-driven models, Ser446 phosphorylation is dynamically regulated during Raf dimerization, affecting downstream ERK activation .
BRAF Inhibitor Resistance: Studies in cutaneous melanoma (CM) reveal that BRAF inhibitor (BRAFi)-resistant cells often exhibit upregulated RTKs (e.g., EGFR) but remain dependent on BRAF signaling . The BRAF (Phospho-Ser446) antibody helps identify compensatory pathways in resistant tumors.
Therapeutic Targeting: While BRAFi-resistant CM cells show EGFR overexpression, cetuximab (an anti-EGFR antibody) induces antibody-dependent cellular cytotoxicity (ADCC) in these cells, suggesting combinatory therapeutic strategies .
Specificity Tests: Antibody specificity is confirmed via peptide inhibition assays and λ-phosphatase treatment, showing loss of signal upon dephosphorylation .
Cross-Reactivity: No cross-reactivity with non-target species (e.g., zebrafish, bovine) reported .
Research Use Only (RUO): Not validated for diagnostic or therapeutic applications .
Sample Handling: Fresh lysates are recommended to avoid protein degradation, as seen in prolonged storage .
Mechanistic Studies: Further exploration of Ser446’s role in Raf dimerization and feedback regulation.
Clinical Correlations: Correlate phospho-Ser446 BRAF levels with patient outcomes in BRAFi-resistant cancers.