Up to 100-fold speedup and multiplexing in optimized DNA-PAINT

DNA-PAINT’s imaging speed has recently been significantly enhanced by optimized sequence design and buffer conditions. However, this implementation has not reached an ultimate speed-limit and importantly is only applicable to imaging single targets. To further improve acquisition speed, we here introduce concatenated, periodic DNA sequence motifs, yielding up to 100-fold faster sampling compared to traditional DNA-PAINT. We extend this approach to six orthogonal sequence motifs, now enabling speed-optimized multiplexed imaging.