Since its introduction in the late 1990s, the classic FISH method has involved overnight incubation and intensive washing steps to achieve accurate DNA probe hybridization results. To reduce turnaround time (TAT) and eliminate some of the technical limitations, alternative one-day FISH methods have gradually become established over the past decades. By reducing the hybridization time to 60–120 minutes, this shortened procedure enabled a FISH reaction to be performed and evaluated within a few hours. In addition, lower denaturation temperatures led to improved tissue and cell morphology in patient samples. The term “same-day FISH” refers to the goal of documenting cell/tissue morphology and molecular cytogenetic (FISH) findings on the same day as sample collection. This requirement is increasing with the growing number of specific clinical needs, for example, in hematological malignancies defined by genetic aberrations. Similarly, the clinical benefit of rapid prenatal testing for common copy number variations (CNVs) of amniotic or chorionic villus samples is recognized. The market for DNA probes, pretreatment, hybridization, and washing solutions suitable for one-day protocols is gradually expanding. Overall, the one-day FISH diagnostic procedure has found its place in the diagnostic arsenal and has partially replaced the conventional (overnight) FISH procedure, at least in selected urgent cases.