2.3 AFM substrate preparation
For AFM imaging, a 10 mm diameter mica disk (grade V1, Ted Pella) was glued onto a steel disk and placed on the sample stage. The mica surface was freshly cleaved prior to sample deposition. To pre-treat the mica, 20 µl NiCl2 (10 mM) were deposited on the surface,19, 20 incubated for 1 min in a petri dish at near saturation humidity, rinsed with 2 ml distilled water, dried with flowing N2 gas or air, and placed back in the humidity chamber for a few minutes. Two methods were used to bind DNA onto mica. For Method 1 (“quick mix”), 0.6 ul Sample A or B was added to 29.4 ul of attachment buffer. This combination was rapidly pipetted onto mica, incubated for 1 min, rinsed with 1-2 ml water, and dried with N2 or air. For Method 2 (“injection method”), 28.6 ul of attachment buffer was first deposited onto mica and 1.4 ul of Sample A or B was added to the middle of the droplet and slightly mixed, incubated for 5 min, rinsed with water, and dried with N2 or air. MgCl2 was needed in the attachment buffer to improve binding of DNA. For both methods, the samples were stored in a dessicator for > 1 day before examination with AFM.