Fig. 3.
Example of measurement of PSTED using the apparent total area measurement of a subresolution naturally occurring cellular structure and the linearisation method. (A) Paired confocal and STED images of microtubules in DAOY cells labelled with Alexa 594 at two STED depletion powers (775 nm pulsed laser). Note the apparent size and area decrease as a function of depletion power. Scale bar represents 2 μm. (B) R as a function of STED depletion power (from Eq. (3)) as measured by total microtubule area for DAOY cells labelled with Alexa 594. (C). R as a function of STED depletion power and bead size (FWHM) for subresolution (approx. 20 nm) size Alexa 594 labelled beads. PSTED was calculated from the reciprocal of the slope of the fitted line (R2 = 0.99) in B and C. Error bars in C represent 1 standard deviation.