In this work, three-dimensional manipulation of multiple defects and structures is performed in the framework of holographic optical trapping approach using a spatial light modulator. A holographic optical tweezers system is constructed using a liquid crystal spatial light modulator to generate multiple optical traps. We optimize the tweezers setup to perform polarization-sensitive holographic optical trapping and then explore properties of optical trapping in thermotropic liquid crystals and compare them to the case of isotropic fluids. One of the major challenges complicating the quantitative measurements in these fluids is the anisotropic nature of the liquid crystal medium, which makes the tight focusing of the laser beam difficult and considerably weakens optical trapping forces. Using liquid crystals with low birefringence allows us to mitigate these artefacts. Optical trapping forces and the trap stiffness are first calibrated for different laser powers using viscous drag forces. This is then used to probe inter-particle and defect-particle interaction forces as well as to characterize tension of line defects in the bulk of liquid crystals.© (2009) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.