Raw image data - DNA-PAINT on 1-micrometer PS particles

Fluorescence microscopy data of single molecule localization microscopy (SMLM) characterization of 1 micrometer polystyrene particles using DNA-paint.Neutravidin-coated 1 μm polystyrene particles (ThermoFisher Scientific F8777) are immobilized on a clean gridded glass coverslip (Ibidi 10816). To this end, the coverslip is first cleaned using 15-minute sonication steps in Triton-X100 and KOH interleaved with sonication in Milli-Q water for rinsing. The coverslip is then dried with nitrogen. A silicone gasket (Grace Bio-Labs SKU 103250) cut to the size of the coverslip is deposited to create a chamber and the assembly is cleaned with oxygen plasma. The cleaned coverslip is incubated with 100 μL of biotinylated BSA (Sigma Aldrich A8549) at 0.1 mg/mL for 30 minutes then rinsed with 1xPBS. Subsequently, the coverslip was incubated for 1 h with 100 μL of 100:1 diluted stock concentration of the particles, then rinsed three times with 200 μL 1xPBS. Thereafter, the sample was incubated with 100 μL of 20 μM biotinylated DNA docking strands for 1 h, washed twice in 1xPBS, and once with the imaging buffer. The docking strand sequence is 5'-biotin-TTATACATCTA-3' (IDT). The imager strand sequence is 3'-atto655-TATGTAGAT-5' (IDT). The imager solution is prepared by diluting the imager strand DNA to 50 pM in imaging buffer consisting of Buffer B with 10 mM MgC2. Both concentrations are adjusted to obtain single events on beads and minimize the background in epifluorescence imaging.

采用单分子定位显微镜（SMLM）对直径为1微米的聚苯乙烯颗粒进行荧光显微镜数据表征，所用方法为DNA绘画技术。ThermoFisher Scientific公司生产的1微米中性载体聚苯乙烯颗粒（F8777）经Neutravidin涂层后，固定于清洁的网格载玻片（Ibidi 10816）上。为此，载玻片首先使用Triton-X100和氢氧化钾交替的15分钟超声处理，并在Milli-Q水中进行超声清洗以进行冲洗。随后，载玻片用氮气吹干。将尺寸与载玻片相匹配的硅橡胶垫圈（Grace Bio-Labs SKU 103250）放置以形成封闭腔室，并使用氧气等离子体进行清洁。清洁后的载玻片与100 μL的0.1 mg/mL生物素化牛血清白蛋白（Sigma Aldrich A8549）在室温下孵育30分钟，然后用1xPBS冲洗。随后，载玻片与100 μL的100:1稀释的颗粒储存浓度溶液孵育1小时，然后用200 μL的1xPBS冲洗三次。之后，样本与100 μL的20 μM生物素化DNA对接链在室温下孵育1小时，用1xPBS冲洗两次，然后用成像缓冲液冲洗一次。对接链序列为5'-biotin-TTATACATCTA-3'（IDT），成像链序列为3'-atto655-TATGTAGAT-5'（IDT）。成像溶液是通过将成像链DNA稀释至成像缓冲液中的50 pM制备而成，该缓冲液包含含有10 mM MgC2的Buffer B。两种浓度均调整为在颗粒上获得单一事件并最小化共聚焦荧光成像中的背景。