Extended Data Fig. 4: DEL selection against HeLa cells with different FR expression levels.
From: Selection of DNA-encoded chemical libraries against endogenous membrane proteins on live cells
a) Besides FA, another FR ligand, methotrexate (MTX) was conjugated with a DNA strand and added to the 4,800-member library as shown in Fig. 5. This 4,802-member DEL was selected against the DNA-tagged FR on HeLa cells with different FR levels; a tag with 7-nt complementarity was used in the selections. b) FP analysis to measure the binding affinity of the MTX–DNA conjugate to the target protein FR, and a Kd of ~26.6 µM was obtained. n = 3 biologically independent FP samples were measured. Data are presented as mean values ± SD (standard deviation) based on biologically independent replicates. c) HeLa cells were cultured in FA-deficient medium and harvested after different passages. Six batches of the cells with different FR expression levels were fluorescently labelled with BP-1/CP-1 and analysed with flow cytometry for each batch. The average number of DNA molecules on each cell was measured; based on a size of 3,000 cubic µm for HeLa cells, the FR concentration for each cell batch was calculated (200 µL selection volume; see Section 9 for calculation method). The 4,802-member DEL was subjected to the same selection procedure as described in Fig. 5,6 against these cell batches, respectively; the selection results were processed also in the same way and summarized in the table. EF: enrichment factor. For each selection, a control without FR tagging was also conducted. d) Column graph summarizing the selection results shown in c).
