KB
K.E. Blokhuis
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Hardware Modification Free Active 3D-Drift Correction in Single-Molecule Localization Microscopy (SMLM)
Model-Based Control of a Super Resolution Microscope
Single-molecule localization microscopy (SMLM) enables imaging at nanometer-scale resolution but is highly sensitive to sample drift. Here, I present a live 3D drift correction approach that uses only fiducial markers and does not require any hardware modifications. The method uses fluorescent light from fiducial markers, extracted directly from the main imaging camera during acquisition. Using the computationally efficient Phasor approach to estimate the 3D-position of the beads \cite{phasor}, the control bandwidth is mostly limited by the maximum frame rate of the camera during acquisition (e.g. rates of >22 Hz at 25 fps). In addition, a system identification framework is proposed to identify drift dynamics, enabling the implementation of an optimal model-based control strategy. Experiments reached closed-loop stability with a precision of 0.6 nm in lateral direction and 2.4 nm in axial direction, showing the potential of the hardware-free drift correction approach.
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Single-molecule localization microscopy (SMLM) enables imaging at nanometer-scale resolution but is highly sensitive to sample drift. Here, I present a live 3D drift correction approach that uses only fiducial markers and does not require any hardware modifications. The method uses fluorescent light from fiducial markers, extracted directly from the main imaging camera during acquisition. Using the computationally efficient Phasor approach to estimate the 3D-position of the beads \cite{phasor}, the control bandwidth is mostly limited by the maximum frame rate of the camera during acquisition (e.g. rates of >22 Hz at 25 fps). In addition, a system identification framework is proposed to identify drift dynamics, enabling the implementation of an optimal model-based control strategy. Experiments reached closed-loop stability with a precision of 0.6 nm in lateral direction and 2.4 nm in axial direction, showing the potential of the hardware-free drift correction approach.