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.