title: Detection of HIV-1 p24 at Attomole Level by Ultrasensitive ELISA with Thio-NAD Cycling.
abstract: To reduce the window period between HIV-1 infection and the ability to diagnose it, a fourth-generation immunoassay including the detection of HIV-1 p24 antigen has been developed. However, because the commercially available systems for this assay use special, high-cost instruments to measure, for example, chemiluminescence, it is performed only by diagnostics companies and hub hospitals. To overcome this limitation, we applied an ultrasensitive ELISA coupled with a thio-NAD cycling, which is based on a usual enzyme immunoassay without special instruments, to detect HIV-1 p24. The p24 detection limit by our ultrasensitive ELISA was 0.0065 IU/assay (i.e., ca. 10(-18) moles/assay). Because HIV-1 p24 antigen is thought to be present in the virion in much greater numbers than viral RNA copies, the value of 10(-18) moles of the p24/assay corresponds to ca. 10(3) copies of the HIV-1 RNA/assay. That is, our ultrasensitive ELISA is chasing the detection limit (10(2) copies/assay) obtained by PCR-based nucleic acid testing (NAT) with a margin of only one different order. Further, the detection limit by our ultrasensitive ELISA is less than that mandated for a CE-marked HIV antigen/antibody assay. An additional recovery test using blood supported the reliability of our ultrasensitive ELISA.
authors.affiliation: Kagawa School of Pharmaceutical Sciences, Tokushima Bunri University, Sanuki, Japan.
authors.forename: Akira
authors.initials: A
authors.lastname: Nakatsuma
authors.affiliation: Kagawa School of Pharmaceutical Sciences, Tokushima Bunri University, Sanuki, Japan.
authors.forename: Mugiho
authors.initials: M
authors.lastname: Kaneda
authors.affiliation: Kagawa School of Pharmaceutical Sciences, Tokushima Bunri University, Sanuki, Japan.
authors.forename: Hiromi
authors.initials: H
authors.lastname: Kodama
authors.affiliation: Kagawa School of Pharmaceutical Sciences, Tokushima Bunri University, Sanuki, Japan; TAUNS Laboratories, Inc., Izunokuni, Japan.
authors.forename: Mika
authors.initials: M
authors.lastname: Morikawa
authors.affiliation: BL Co., Ltd., Numazu, Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Watabe
authors.affiliation: TAUNS Laboratories, Inc., Izunokuni, Japan.
authors.forename: Kazunari
authors.initials: K
authors.lastname: Nakaishi
authors.affiliation: Faculty of Science, Hokkaido University, Sapporo, Japan.
authors.forename: Masakane
authors.initials: M
authors.lastname: Yamashita
authors.affiliation: Faculty of Pharmaceutical Sciences, Health Sciences University of Hokkaido, Ishikari-Tobetsu, Japan.
authors.forename: Teruki
authors.initials: T
authors.lastname: Yoshimura
authors.affiliation: Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo, Japan.
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Miura
authors.affiliation: Kagawa School of Pharmaceutical Sciences, Tokushima Bunri University, Sanuki, Japan.
authors.forename: Masaki
authors.initials: M
authors.lastname: Ninomiya
authors.affiliation: Kagawa School of Pharmaceutical Sciences, Tokushima Bunri University, Sanuki, Japan.
authors.forename: Etsuro
authors.initials: E
authors.lastname: Ito
mesh_terms: D042241:Early Diagnosis; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D016655:HIV Core Protein p24 / Q000097:blood* / Q000276:immunology; D015658:HIV Infections / Q000097:blood / Q000175:diagnosis* / Q000276:immunology; D015497:HIV-1 / Q000276:immunology*; D006801:Humans; D057230:Limit of Detection; D009243:NAD / Q000031:analogs & derivatives* / Q000378:metabolism
chemical_list: D016655:HIV Core Protein p24; D009243:NAD; C003451:thionicotinamide adenine dinucleotide