Figure 2

Exemplary focus sweep data for determining the z-position of SPPs within a microfluidic transport system. Curves (a–c) show the TBG, as a measure for particle sharpness, in dependence on the set z-position of the SPPs with respect to the focal plane. The data was acquired by sweeping the sample through the focal plane of the used microscope (starting with a position below the focal plane) in 25 nm steps for Dynabeads M-270/micromer-M 3 µm, and in 50 nm steps for micromer-M 4 µm, recording reference images at every step. Curves (a) and (b) show the general progression of TBG for different z-position regimes (below, within, and above the focal plane) and demonstrate that the chosen approach for evaluating the particle sharpness is suitable for different types of SPP. The regions of highest sensitivity (steepest slope) for z-position detection with approximately linear progression are highlighted in red. For micromer-M 4 µm SPP (c), only the linear regime of the highest sensitivity focus sweep data is displayed which was taken for further particle transport experiments as the z-position calibration. Exemplary background-subtracted images of the respective SPP at the indicated z-position are shown in the insets of each plot.