Transfect HEK cells with BCR genes (gantenerumab heavy & light chains, CD79A, CD79B), and Lyn. Tage each component with FP. Check for membrane localization
Proper protein Assembly in HEK cells
Transfect HEK cells with BCR genes, Lyn, Syk WT & Protein Kinase domain mutation. Culture both Syk types wit hand without AB and ELISA for phosphorylated Syk and/or Lyn
Recruitment of Syk and/or Lyn to the BCR
Transfect HEK cells with BCR genes, Lyn tagged with BFP, Syk tagged with RFP. Culture with and without Beta Amyloid. Check for membrane localization.
TEV Protease Cleavage
Transfect HEK cells with BCR genes, Lyn, and Syk. Fuse TEV cleavage sites with different fluorescent proteins to the heavy chain, CD79A, and CD79B. Fuse TEV protease to Lyn or Syk. Culture with and without Beta Amyloid and check for membrane delocalization.
Full system operation
Transfect HEK cells with BCR genes, Lyn, and Syk. Fuse TEV cleavage sites with TF to all/most promising BCR proteins, Fuse TEV protease to Lynor Syk. Transfect TF promoter expressing RFP. Culture with and without Beta Amyloid and check for flourescence. Characterize signal strength based off of flourescence.