(A) Experimental setup for ZMW-based single-molecule SNAPf experiments. (B) Full-chip average fluorescence values for SNAPf translation detected by ZMW. When the reaction is treated with a translation inhibitor, GFP is substituted for SNAPf, or plasmid is omitted, fluorescence decreases significantly. (C) The fluorescent signal from an individual ZMW well. Colocalization of red Cy5 signal (Cy5-Z50S) to green DY549 signal (SNAPf) is observed, and the dyes photobleach after 200 and 100 sec, respectively.