Aptamers evolved from live cells as effective molecular probes for cancer study. Researchers developed a cell-based aptamer selection strategy to identify molecular signatures on the surface of targeted cells by exploiting differences at the molecular level between any two cell types. A group of aptamers were generated for the specific recognition of leukemia cells, with equilibrium dissociation constants (Kd) in the nanomolar-to-picomolar range. The cell-based selection process is simple, fast, and reproducible, and can be performed without prior knowledge of target molecules. The selected aptamers can specifically recognize target leukemia cells mixed with normal human bone marrow aspirates and can also identify cancer cells closely related to the target cell line in real clinical specimens. This approach holds great promise for developing specific molecular probes for cancer diagnosis and biomarker discovery. The study highlights the challenges in understanding human diseases at the molecular level due to the lack of effective probes. Cancers and other diseases originate from genetic mutations, leading to changes in cell behavior and morphology. Traditional cancer diagnosis relies on tumor morphology, which is not suitable for early diagnosis or evaluating molecular changes. Molecular characteristics, especially proteomic features, are more suitable for cancer classification. However, identifying molecular signatures of cancer remains a challenge. Molecular-level differences exist between any two cell types, such as normal vs. tumor cells. These differences are significant for disease understanding and diagnosis. However, identifying these differences is challenging with current technologies. The study presents a practical strategy to compare cancer cells with normal cells and identify molecular differences. A new class of molecular probes, aptamers, were isolated for recognizing molecular differences on cell surfaces. These aptamers can specifically detect target cancer cells in the presence of other cells, enabling effective disease studies and early diagnosis. Aptamers are single-stranded DNA or RNA that can bind to their targets, ranging from small molecules to proteins. They are selected through the SELEX process, which involves iterative rounds of binding and selection. Aptamers have many applications in bioanalysis, biomedicine, and biotechnology due to their high specificity, low molecular weight, and ease of production. The study aimed to develop effective aptamers for leukemia studies. A cell-based SELEX method was used to select aptamers specific to target cells. A counterselection strategy was employed to enrich DNA sequences that only interact with target cells. The selected aptamers can recognize molecular differences on the cell surface, providing valuable tools for identifying new biomarkers. The development of specific probes for molecular signatures on cancer cell surfaces offers new opportunities for personalized medicine. The study demonstrated that the selected aptamers can specifically recognize target cells in real biological samples, showing high specificity and affinity. The aptamers can be used to isolate disease-specific protein targets, facilitating the discovery of clinically important biomarkers. The binding sites of the aptamers were found to be likely membrane proteins. The cell-based SELEX strategy provides a simple, fast, andAptamers evolved from live cells as effective molecular probes for cancer study. Researchers developed a cell-based aptamer selection strategy to identify molecular signatures on the surface of targeted cells by exploiting differences at the molecular level between any two cell types. A group of aptamers were generated for the specific recognition of leukemia cells, with equilibrium dissociation constants (Kd) in the nanomolar-to-picomolar range. The cell-based selection process is simple, fast, and reproducible, and can be performed without prior knowledge of target molecules. The selected aptamers can specifically recognize target leukemia cells mixed with normal human bone marrow aspirates and can also identify cancer cells closely related to the target cell line in real clinical specimens. This approach holds great promise for developing specific molecular probes for cancer diagnosis and biomarker discovery. The study highlights the challenges in understanding human diseases at the molecular level due to the lack of effective probes. Cancers and other diseases originate from genetic mutations, leading to changes in cell behavior and morphology. Traditional cancer diagnosis relies on tumor morphology, which is not suitable for early diagnosis or evaluating molecular changes. Molecular characteristics, especially proteomic features, are more suitable for cancer classification. However, identifying molecular signatures of cancer remains a challenge. Molecular-level differences exist between any two cell types, such as normal vs. tumor cells. These differences are significant for disease understanding and diagnosis. However, identifying these differences is challenging with current technologies. The study presents a practical strategy to compare cancer cells with normal cells and identify molecular differences. A new class of molecular probes, aptamers, were isolated for recognizing molecular differences on cell surfaces. These aptamers can specifically detect target cancer cells in the presence of other cells, enabling effective disease studies and early diagnosis. Aptamers are single-stranded DNA or RNA that can bind to their targets, ranging from small molecules to proteins. They are selected through the SELEX process, which involves iterative rounds of binding and selection. Aptamers have many applications in bioanalysis, biomedicine, and biotechnology due to their high specificity, low molecular weight, and ease of production. The study aimed to develop effective aptamers for leukemia studies. A cell-based SELEX method was used to select aptamers specific to target cells. A counterselection strategy was employed to enrich DNA sequences that only interact with target cells. The selected aptamers can recognize molecular differences on the cell surface, providing valuable tools for identifying new biomarkers. The development of specific probes for molecular signatures on cancer cell surfaces offers new opportunities for personalized medicine. The study demonstrated that the selected aptamers can specifically recognize target cells in real biological samples, showing high specificity and affinity. The aptamers can be used to isolate disease-specific protein targets, facilitating the discovery of clinically important biomarkers. The binding sites of the aptamers were found to be likely membrane proteins. The cell-based SELEX strategy provides a simple, fast, and