title: Multiplex screening for blood-borne viral, bacterial, and protozoan parasites using an OpenArray platform.
abstract: The use of nucleic acid tests for detection of pathogens has improved the safety of blood products. However, ongoing pathogen emergence demonstrates a need for development of devices testing for multiple pathogens simultaneously. One approach combines two proven technologies: Taqman chemistry for target identification and quantification and the OpenArray nanofluidic real-time PCR platform for spatial multiplexing of assays. A panel of Taqman assays was developed to detect nine blood-borne pathogens (BBPs): four viral, two bacterial, and three protozoan parasites. The custom BBP OpenArray plate with 18 assays was tested for specificity and analytical sensitivity for nucleic acid from each purified pathogen and with pathogen-spiked human blood and plasma samples. For most targets, the limits of detection (10 to 10,000 copies/mL) were comparable with existing real-time platforms. The testing of the BBP OpenArray with pathogen-spiked coded human plasma or blood samples and negative control specimens demonstrated no false-positive results among the samples tested and correctly identified pathogens with the lowest concentration detected ranging from 10 cells/mL (Trypanosoma cruzi) to 10,000 cells/mL (Escherichia coli). These results represent a proof of concept that indicated the BBP OpenArray platform in combination with Taqman chemistry may provide a multiplex real-time PCR pathogen detection method that points the way for a next-generation platform for infectious disease testing in blood.
authors.affiliation: Life Technologies Corp., South San Francisco, California.
authors.forename: Elena
authors.initials: E
authors.lastname: Grigorenko
authors.affiliation: Division of Emerging and Transfusion Transmitted Diseases, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland.
authors.forename: Carolyn
authors.initials: C
authors.lastname: Fisher
authors.affiliation: Life Technologies Corp., South San Francisco, California.
authors.forename: Sunali
authors.initials: S
authors.lastname: Patel
authors.affiliation: Division of Emerging and Transfusion Transmitted Diseases, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland.
authors.forename: Caren
authors.initials: C
authors.lastname: Chancey
authors.affiliation: Division of Emerging and Transfusion Transmitted Diseases, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland.
authors.forename: Maria
authors.initials: M
authors.lastname: Rios
authors.affiliation: Division of Emerging and Transfusion Transmitted Diseases, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland.
authors.forename: Hira L
authors.initials: HL
authors.lastname: Nakhasi
authors.affiliation: Division of Emerging and Transfusion Transmitted Diseases, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland. Electronic address: robert.duncan@fda.hhs.gov.
authors.forename: Robert C
authors.initials: RC
authors.lastname: Duncan
mesh_terms: D001424:Bacterial Infections / Q000097:blood* / Q000175:diagnosis / Q000235:genetics; D017848:Blood-Borne Pathogens; D004269:DNA, Bacterial / Q000032:analysis; D016054:DNA, Protozoan / Q000032:analysis; D004279:DNA, Viral / Q000032:analysis; D006801:Humans; D057230:Limit of Detection; D025202:Molecular Diagnostic Techniques / Q000379:methods; D016133:Polymerase Chain Reaction / Q000379:methods*; D011528:Protozoan Infections / Q000097:blood* / Q000175:diagnosis / Q000235:genetics; D019914:Taq Polymerase; D014777:Virus Diseases / Q000097:blood* / Q000175:diagnosis / Q000235:genetics
chemical_list: D004269:DNA, Bacterial; D016054:DNA, Protozoan; D004279:DNA, Viral; D019914:Taq Polymerase