Multiplex Screening for Virus, Bacteria, and Parasite Blood Borne Pathogens with the OpenArray Platform

Wednesday, October 26, 2011 — Poster Session III
10:00 a.m. – Noon	Natcher Conference Center	FDA/CBER	TECH-2

Authors

• C. Fisher
• E. Grigorenko
• S. Patel
• K. Munnelly
• H. Nakhasi
• R. Duncan

Abstract

Multiplex screening for virus, bacteria and parasite blood borne pathogens is needed for detection of newly emerging pathogens and for screening of US blood donors. A limiting factor for pathogen detection by multiplex assays is the number of agents that can be detected simultaneously. We report advanced multiplexing with an OpenArray nanofluidic system. Each OpenArray has 48 blocks of nanoliter-scale through-holes on a microscope slide sized metal wafer. Each block contained the same 36 primer pairs and probes to simultaneously perform 2688 PCR reactions. Primer and probe sets for viruses (HIV-1, HCV, HBV, West Nile Virus), parasites (Trypanosoma cruzi, Leishmania, Plasmodium falciparum), and Gram negative bacteria (such as Escherichia coli) were designed with TaqMan fluorescent probes and custom manufactured in an OpenArray. The performance of the OpenArray was evaluated with purified pathogen nucleic acid, with nucleic acid from human blood and human plasma spiked with pathogens, and with blinded samples. The pathogens were correctly identified and there were no false positives in the blinded samples. The limits of detection ranged from 10 to 500 pathogens per ml. Our results confirmed that multiplex screening with the OpenArray platform was a successful method for pathogen detection in blood.

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