Conventional in vitro or ex vivo bioanalytical quantitative sample preparation methods for the determination of compounds in biological tissues are often coupled with challenges in obtaining an assay representative of the system of interest. The rising interest in in vivo microsampling bioanalytical methods is due to the unique advantages they offer over their in vitro counterparts. In vivo solid-phase microextraction (SPME), a diffusion-based microsampling tool, has been successfully applied in recent studies to various biological systems. This review presents recent trends in tissue bioanalysis using in vivo SPME as a sample preparation tool. Efforts were made to discuss the various bioapplications of the method while highlighting possible strategies for improved sensitivity where needed. In vivo SPME devices currently employed for the various applications have also been described. In addition, we highlight selectivity of a new class of biocompatible coatings that can potentially improve the coverage of metabolites for untargeted metabolomics.