63 plan apo 1.4 na oil immersion objective

The diffusion of individual fluorescent nanoparticles in Matrigel was quantified using multiple particle tracking (MPT) as previously described [25 ]. Fluorescent nanoparticles (2 μg/mL) were diluted in cold Matrigel (4 °C) and then added to Lab-Tek glass-bottom chamber. The chamber was placed in at 37 °C incubator for a minimum of 15 min before imaging to allow Matrigel to form into a gel matrix. The movement of individual nanoparticles in Matrigel was imaged at a frame rate of 20 frames/s for a total of 400 frames (20 s) for 40 and 100 nm nanoparticles and at a frame rate of 10 frames/s for a total of 400 frames (40 s) for 20 nm nanoparticles. Images were captured using a Zeiss LSM5 Duo slit scanning confocal microscope (Zeiss, Thornwood, NY) with a 63× Plan-Apo/1.4 NA oil-immersion objective. Movies were analyzed using a custom written MATLAB automated tracking code to extract x, y-coordinates of nanoparticles over time, as previously described [24 (link), 25 ]. Each particle type was imaged at least three separate times with at least 100 particles tracked per sample every time. The geometric mean of the mean squared displacement (MSD) was calculated per particle type and the average was calculated as a function of time scale.