Since their invention in 1986 by Arthur Ashkin and colleagues, optical tweezers have become an essential tool in several fields of physics, spectroscopy, biology, nanotechnology, and thermodynamics. In this Tutorial, we provide a primer on how to calibrate optical tweezers and how to use them for advanced applications. After a brief general introduction on optical tweezers, we focus on describing and comparing the various available calibration techniques. Then, we discuss some cutting-edge applications of optical tweezers in a liquid medium, namely to study single-molecule and single-cell mechanics, microrheology, colloidal interactions, statistical physics, and transport phenomena. Finally, we consider optical tweezers in vacuum, where the absence of a viscous medium offers vastly different dynamics and presents new challenges. We conclude with some perspectives for the field and the future application of optical tweezers. This Tutorial provides both a step-by-step guide ideal for non-specialists entering the field and a comprehensive manual of advanced techniques useful for expert practitioners. All the examples are complemented by the sample data and software necessary to reproduce them.
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