(203e) Loop Mediated Isothermal Amplification and Detection of Escherichia Coli Using Lateral Flow Strips

The need for rapid point-of-care diagnostics for infectious diseases has been steadily increasing globally, especially in developing countries. E. Coli is responsible for a large number of infections including water and food borne infections, urinary tract infection (UTI) and life threatening conditions such as neonatal meningitis and neonatal sepsis. Detection of E. Coli is generally done by bacterial cultures which usually takes 48-72 hours and requires skilled operators and expensive equipment and laboratories. Recently, loop mediated isothermal amplification (LAMP) technique has been developed which amplifies the DNA at around 63-65 °C with high specificity. The LAMP procedure utilizes 4 or 6 primers and can be performed on a regular water bath or a heating block maintained at the specified temperature. We developed a Nucleic Acid Lateral Flow (NALF) device to detect LAMP amplification products in an easy to read format. The detection of the LAMP E. Coli product is performed on a lateral flow strip which primarily consists of a nitrocellulose membrane with proteins/antibodies immobilized at specified locations. Labeled primers, namely biotin and FITC labeled outer primers are used in the amplification which takes about 30-60 minutes and the detection is performed using a sandwich type immunoassay. Results show a clear visible difference between the positive and negative samples and similar sensitivity and specificity when compared to PCR, making this assay a reliable, rapid and inexpensive diagnostic tool. An additional advantage could be the utility of this detection method in multiplexed assays where more than one type of E. Coli (such as toxigenic or hemorrhagic) or different bacteria (ie Klebsiella sp, Staphylococcus sp, etc) can be detected in as single assay and on a single strip in a point of care manner.