Identification of an amino acid responsible for the CD3 polymorphism in cynomolgus monkeys (Macaca fascicularis)

Area of Science:

• Immunology
• Molecular Biology
• Primate Research

Background:

• The FN18 monoclonal antibody (mAb) targets CD3 molecules on lymphocytes.
• Polymorphism in the CD3epsilon chain of cynomolgus monkeys (Macaca fascicularis) affects FN18 mAb reactivity.
• Understanding this polymorphism is crucial for immunological studies involving non-human primates.

Purpose of the Study:

• To investigate the cause of differential reactivity of the FN18 mAb against cynomolgus monkey lymphocytes.
• To identify the specific epitope recognized by the FN18 mAb on the CD3epsilon chain.
• To elucidate the role of specific amino acid residues in epitope formation.

Main Methods:

• Expression of CD3epsilon in COS7 cells via plasmid DNA transfection.
• Site-directed mutagenesis of the CD3epsilon gene.
• Analysis of FN18 mAb binding to cells expressing wild-type and mutant CD3epsilon.

Main Results:

• The epitope for FN18 mAb was successfully expressed on COS7 cells using DNA from an FN18-positive monkey.
• Mutagenesis studies revealed that amino acid residue at position 67 is essential for the FN18 epitope.
• This finding explains the variable reactivity of FN18 mAb in cynomolgus monkey populations.

Conclusions:

• CD3epsilon chain polymorphism at residue 67 is responsible for the differential reactivity of the FN18 mAb in cynomolgus monkeys.
• This study provides a molecular basis for understanding T-cell marker variability in non-human primates.
• The findings are important for the accurate use of FN18 mAb in immunological research and diagnostics.