Model-based estimation of 3-D stiffness parameters in photonic-force microscopy.

We propose a system to characterize the 3-D diffusion properties of the probing bead trapped by a photonic-force microscope. We follow a model-based approach, where the model of the dynamics of the bead is given by the Langevin equation. Our procedure combines software and analog hardware to measure the corresponding stiffness matrix. We are able to estimate all its elements in real time, including off-diagonal terms. To achieve our goal, we have built a simple analog computer that performs a continuous preprocessing of the data, which can be subsequently digitized at a much lower rate than is otherwise required. We also provide an effective numerical algorithm for compensating the correlation bias introduced by a quadrant photodiode detector in the microscope. We validate our approach using simulated data and show that our bias-compensation scheme effectively improves the accuracy of the system. Moreover, we perform experiments with the real system and demonstrate real-time capabilities. Finally, we suggest a simple adjunction that would allow one to determine the mass matrix as well.

Intracellular nanomanipulation by a photonic-force microscope with real-time acquisition of a 3D stiffness matrix.

A traditional photonic-force microscope (PFM) results in huge sets of data, which requires tedious numerical analysis. In this paper, we propose instead an analog signal processor to attain real-time capabilities while retaining the richness of the traditional PFM data. Our system is devoted to intracellular measurements and is fully interactive through the use of a haptic joystick. Using our specialized analog hardware along with a dedicated algorithm, we can extract the full 3D stiffness matrix of the optical trap in real time, including the off-diagonal cross-terms. Our system is also capable of simultaneously recording data for subsequent offline analysis. This allows us to check that a good correlation exists between the classical analysis of stiffness and our real-time measurements. We monitor the PFM beads using an optical microscope. The force-feedback mechanism of the haptic joystick helps us in interactively guiding the bead inside living cells and collecting information from its (possibly anisotropic) environment. The instantaneous stiffness measurements are also displayed in real time on a graphical user interface. The whole system has been built and is operational; here we present early results that confirm the consistency of the real-time measurements with offline computations.