AbstractAdvancements in lab‐on‐a‐chip technologies have revolutionized the single‐cell analysis field. However, an accessible platform for in‐depth screening and specific retrieval of single cells, which moreover enables studying diverse cell types and performing various downstream analyses, is still lacking. As a solution, FLUIDOT is introduced, a versatile microfluidic platform incorporating customizable microwells, optical tweezers and an interchangeable cell‐retrieval system. Thanks to its smart microfluidic design, FLUIDOT is straightforward to fabricate and operate, rendering the technology widely accessible. The performance of FLUIDOT is validated and its versatility is subsequently demonstrated in two applications. First, drug tolerance in yeast cells is studied, resulting in the discovery of two treatment‐tolerant populations. Second, B cells from convalescent COVID‐19 patients are screened, leading to the discovery of highly affine, in vitro neutralizing monoclonal antibodies against SARS‐CoV‐2. Owing to its performance, flexibility, and accessibility, it is foreseen that FLUIDOT will enable phenotypic and genotypic analysis of diverse cell samples and thus elucidate unexplored biological questions.