Escherichia coli (E. coli) outbreaks are a serious health issue in the United States today. These outbreaks are caused by ingesting the contaminated food with pathogenic E. coli strains. One possible way to prevent outbreaks is by detecting the harmful E. coli via a biosensor with a DNA aptamer before consuming the contaminated food. Using the Systematic Evolution of Ligands by EXponential enrichment (SELEX) technique, the DNA aptamer sequence with a high affinity and specificity to the target E. coli strain K12 can be determined. However, the current SELEX procedure is time-consuming. Also, the selection progress cannot be monitored in real time with the conventional SELEX method without fluorescently labeling or radiolabeling. In this project, we present the development of a faster and safer SELEX method combined with surface plasmon resonance (SPR-Cell-SELEX) to identify DNA aptamers that specifically targets E. coli K12. The SELEX with surface plasmon resonance (SPR) can resolve these disadvantages of the current SELEX technique by measuring the molecular binding kinetics in real-time without any labeling.