FIGURE 1.

Biochemical characterization of Miro1 specific Nbs. (A) Amino acid sequence alignment of the complementary determining region (CDR) 3 of seven unique Miro1-Nbs positively identified by phage ELISA (complete amino acid sequences of all Miro1-Nbs are shown in Supplementary Table S2). (B) Recombinant expression and purification of Nbs using immobilized metal affinity chromatography (IMAC) and size exclusion chromatography (SEC). (C) For biolayer interferometry based affinity measurements, Miro1-Nbs were biotinylated and immobilized on streptavidin sensors. Kinetic measurements were performed by using four concentrations of purified Miro1 As an example, the sensogram of Miro1 at concentrations 7.8, 15.6, 31.2, and 62.5 nM on immobilized M41-Nb (illustrated with increasingly darker shades from low to high concentration) is shown and global 1:1 fits are illustrated as dashed line (upper panel).