Fig. 2. Light-controlled DNA photocrosslinking.

a, Schematic for light-directed barcode attachment on glass slides. Biotinylated single-stranded DNA oligos are immobilized onto glass surfaces with biotin–streptavidin binding. Fluorescent barcode strands containing a CNVK moiety in the complementary domain are hybridized to these immobilized oligos. Target pixels corresponding to ROIs in the field of view are UV-illuminated in a parallelized fashion using a DMD to photocrosslink the barcodes in a photomask pattern. Uncrosslinked strands are removed by stringent washes, which reveals the encoded barcode pattern in fluorescence. b, Custom patterning (right) achieved by using a cat photo (left) to create a binary photomask and photocrosslinking the fluorescent CNVK-containing barcode strands onto a functionalized glass slide. c, Iterative photocrosslinking using three photomasks (left) that define three ROIs to attach three orthogonal barcode strands onto a DNA-coated glass slide, forming a Penrose triangle (right).