The growing demand for membrane protein research, coupled with the structural flexibility and high-throughput screening efficiency of aptamers as ligand candidates, has established aptamers as a robust tool for investigating membrane proteins. In this study, we present a novel in vitro evolution strategy for the development of covalent aptamers, leveraging the principles of fluorine-sulfur exchange (SuFEx) chemistry. This strategy facilitates the screening of aptamers with high specificity for target membrane proteins, ultimately yielding cross-linking aptamers (xl-aptamers). These aptamers exhibit sustained and specific binding affinity toward their target proteins, thereby enabling efficient cell classification. Their performance underscores considerable potential for applications in single-cell and spatial proteomics research.