A Rapid and Simple Integrated Extraction Amplification and Detection Device for Y. pestis
Abstract
In order to fulfill the need for a simple self-contained device for the detection of Yersinia pestis target DNA, we combined unique approaches that exploited solid phase methods for DNA extraction, isothermal amplification and visual detection. In Phase I of this project, optimization of DNA extraction and purification was achieved using a unique capture system, Xtra BindTM. A homogeneous method of isothermal strand displacement amplification was used in the first part of this program to amplify a 572 base pair PCR product insert of the caf1 gene. A detection method using lateral flow immunochemistry was adapted to the detection of the amplified products. By September 1999, we demonstrated the feasibility of all the elementary steps within the device. In the Phase II of this project, new primer sets were designed for Y pestis genomic DNA and the best set optimized for low limit of detection. Extensive changes were made to the extraction, amplification and detection chemistries in order to achieve our limit of detection in human plasma samples. In addition, changes in the design of the device were made in order to render it functional. A level of 1 X 10 to the 3 starting copies of target was achieved within the device.
Document Details
- Document Type
- Technical Report
- Publication Date
- Oct 01, 2000
- Accession Number
- ADB260563
Entities
People
- Hugh Maguire
- Jeff Ives
- John C. Gerdes
- Roy Mondesire
- Sam Woronoff