Figure 1: Scheme of the particle-tracking set-up and the image analysis routine.
From: Camera-based three-dimensional real-time particle tracking at kHz rates and Ångström accuracy
Spherical particles within a fluidic cell are mounted on the sample stage of a home-built microscope. The latter comprises a piezo-actuated nanopositiong stage allowing for sample displacements in all three dimensions with Ångström precision. A pair of magnets above the fluidic cell is used to exert forces on DNA-tethered magnetic beads. Tracking of the particle positions is carried out either in high-resolution mode supporting tracking of up to four particles in parallel at high spatial resolution and kHz rates or in multibead mode supporting tracking of tens of particles at moderate spatial resolution and rates up to 500 Hz. For high-resolution tracking, the sample is illuminated using a fibre-coupled mercury lamp. Bead images are acquired simultaneously from distant ROIs using a CMOS camera. Image processing (see red arrows) is carried out in parallel within the CPU regarding x, y and z position calculations as well as the GPU regarding radial intensity profile calculations. The red squares in the high-resolution sample image indicate the 160 pixels × 160 pixels ROIs that are streamed from the camera into CPU and GPU. For multibead tracking, the sample is illuminated with a 625-nm LED. Full frame images are streamed into the GPU where also all image processing is carried out (see grey arrows). The red square in the multibead sample image corresponds to the size of the 72 pixels × 72 pixels ROIs that are used for tracking. The size of the shown sample image regions is 308 pixels × 224 pixels.
