Extended Data Fig. 1: ABC synthesis using different stoichiometries of BP-Tz and Ab-TCO.
From: Antibody–bottlebrush prodrug conjugates for targeted cancer therapy
a. Non-reducing SDS-PAGE analysis of model IgG1-based ABCs as a function of synthesis stoichiometry. BAR = average brush–antibody ratio. b. Non-reducing SDS-PAGE analysis of Trastuzumab-based ABCs as a function of synthesis stoichiometry. c. Flow cytometry histograms showing Cy5.5-HER2 uptake into HER2 + BT-474 cells as a function of synthesis stoichiometry (that is, the constructs are present as a mixture of BAR values). The x-axis represents the Cy5.5 fluorescence intensity. Since the number of Cy5.5 dyes per ABC increases with BAR, histograms are normalized to the total Cy5.5 loading for each construct to enable comparison (normalized to 40 µg/mL Cy5.5-BP, 1 h). d. Flow cytometry histograms for the same constructs shown in panel c normalized by antibody dose (normalized to 10 µg/mL mAb, 1 h). The x-axis represents the Cy5.5 fluorescence intensity. e. and f. Flow cytometry histograms for BT-474 cell uptake of isolated Cy5.5-HER2 ABCs with different BAR values (25 µg/mL, 1 h) (that is, ABCs with each BAR were separated from the synthesis mixture). Results are presented as mean ± SEM (n = 3 biological replicates). Statistical analysis was done using a 2-tailed t-test. For these statistical tests, **** denotes P < 0.001.
