Escherichia coli is an indicator of fecal contamination in recreational water and some strains are responsible for serious food poisoning in humans. Therefore, having a rapid, sensitive, selective, and simple E. coli detection is important. In this study, DNA nanopyramids anchor is utilized for immunological detection of E. coli lipopolysaccharides, lysate, and whole bacteria. By measuring the E. coli amount electrochemically, detection limits of 0.20 ng mL(-1)E. coli lipopolysaccharides and 1.20 CFU mL(-1) equivalent of lysed E. coli bacteria are obtained. Our method confers selectivity to detect E. coli in the presence of various other macromolecules. Therefore, our system can be developed to test the E. coli content in real water samples.