This review presents a method for detecting and identifying single molecules in solution using fluorescence correlation spectroscopy (FCS), combined with devices for trapping single molecules in an electric field. The method is applied to studies of molecular evolution, enabling fast screening of large mutant spectra where targets are labeled by specific fluorescent ligands. It allows monitoring concentrations down to 10^-15 M without amplification, making it highly sensitive for molecular diagnostics. The method is particularly useful for detecting rare structures and activities, diagnosing diseases at early stages, and advancing evolutionary biotechnology. The technique involves detecting the quanta of fluorescence emitted by a probe that specifically binds to a target molecule diffusing through an illuminated cavity. The corresponding signal is discriminated from background noise by auto- or cross-correlating the fluorescent light intensity fluctuations. FCS has been developed and refined with recent advances in laser and microscopy technology, enabling detection of single molecules with high sensitivity. The method is combined with screening and separation techniques used in applied molecular evolution, allowing for the early detection and selection of mutants during molecular evolution. The results suggest that this method could revolutionize three fields: large-scale screening techniques for rare structures, molecular diagnostics with high sensitivity, and evolutionary biotechnology for biased adaptation to new functions. The principle of FCS relies on the detection of single molecules in a small volume, where the signal-to-noise ratio is high enough to detect single quantum bursts of fluorescent light. The method is particularly effective for detecting single molecules in solutions with concentrations as low as 10^-15 M. The technique has been applied to various biological systems, including DNA and RNA sequencing, and has shown promise in identifying single virus particles and detecting rare molecular interactions. The method also involves electrical trapping techniques to separate and isolate single molecules, which can be used in conjunction with FCS for screening large numbers of molecules. These techniques allow for the identification and amplification of specific molecular structures, which is crucial for molecular evolution studies. The review highlights the potential of FCS in various applications, including molecular diagnostics, evolutionary biotechnology, and DNA and RNA sequencing. The method has been developed through collaboration between researchers at the Max-Planck-Institut für Biophysikalische Chemie and Karolinska Institutet, and has been supported by various research grants and funding sources.This review presents a method for detecting and identifying single molecules in solution using fluorescence correlation spectroscopy (FCS), combined with devices for trapping single molecules in an electric field. The method is applied to studies of molecular evolution, enabling fast screening of large mutant spectra where targets are labeled by specific fluorescent ligands. It allows monitoring concentrations down to 10^-15 M without amplification, making it highly sensitive for molecular diagnostics. The method is particularly useful for detecting rare structures and activities, diagnosing diseases at early stages, and advancing evolutionary biotechnology. The technique involves detecting the quanta of fluorescence emitted by a probe that specifically binds to a target molecule diffusing through an illuminated cavity. The corresponding signal is discriminated from background noise by auto- or cross-correlating the fluorescent light intensity fluctuations. FCS has been developed and refined with recent advances in laser and microscopy technology, enabling detection of single molecules with high sensitivity. The method is combined with screening and separation techniques used in applied molecular evolution, allowing for the early detection and selection of mutants during molecular evolution. The results suggest that this method could revolutionize three fields: large-scale screening techniques for rare structures, molecular diagnostics with high sensitivity, and evolutionary biotechnology for biased adaptation to new functions. The principle of FCS relies on the detection of single molecules in a small volume, where the signal-to-noise ratio is high enough to detect single quantum bursts of fluorescent light. The method is particularly effective for detecting single molecules in solutions with concentrations as low as 10^-15 M. The technique has been applied to various biological systems, including DNA and RNA sequencing, and has shown promise in identifying single virus particles and detecting rare molecular interactions. The method also involves electrical trapping techniques to separate and isolate single molecules, which can be used in conjunction with FCS for screening large numbers of molecules. These techniques allow for the identification and amplification of specific molecular structures, which is crucial for molecular evolution studies. The review highlights the potential of FCS in various applications, including molecular diagnostics, evolutionary biotechnology, and DNA and RNA sequencing. The method has been developed through collaboration between researchers at the Max-Planck-Institut für Biophysikalische Chemie and Karolinska Institutet, and has been supported by various research grants and funding sources.