title: Rapid Fluorescent Detection Assay for Human Parainfluenza Viruses.
abstract: Human parainfluenza virus type 1 (hPIV1) does not form clear plaque by the conventional plaque formation assay because of slightly a cytopathic effects in many cell lines infected with hPIV1, thus making in virus titration, isolation and inhibitor evaluation difficult. We have succeeded in fluorescent histochemical visualization of sialidase activities of influenza A and B viruses, Newcastle disease virus and Sendai virus by using a novel fluorescent sialidase substrate, 2-(benzothiazol-2-yl)-4-bromophenyl 5-acetamido-3,5-dideoxy-α-D-glycero-D-galacto-2-nonulopyranosidonic acid (BTP3-Neu5Ac). In this study, we applied the BTP3-Neu5Ac assay for rapid detection of hPIV1 and hPIV type 3. The BTP3-Neu5Ac assay could histochemically visualize dot-blotted hPIVs on a membrane and hPIV-infected cells as local fluorescence under UV irradiation. We succeeded in distinct fluorescent visualization of hPIV1-infected cells in only 3 d using the BTP3-Neu5Ac assay. Due to there being no fixation, hPIV1 was isolated directly from fluorescent stained focus cells by the BTP3-Neu5Ac assay. Establishment of a sensitive, easy, and rapid fluorescent focus detection assay for hPIV, hPIV1 in particular will contribute greatly to progress in hPIV studies.
authors.affiliation: Department of Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka.
authors.forename: Tadanobu
authors.initials: T
authors.lastname: Takahashi
authors.forename: Maiko
authors.initials: M
authors.lastname: Takano
authors.forename: Yuuki
authors.initials: Y
authors.lastname: Kurebayashi
authors.forename: Takashi
authors.initials: T
authors.lastname: Agarikuchi
authors.forename: Chihiro
authors.initials: C
authors.lastname: Suzuki
authors.forename: Keijo
authors.initials: K
authors.lastname: Fukushima
authors.forename: Shunsaku
authors.initials: S
authors.lastname: Takahashi
authors.forename: Tadamune
authors.initials: T
authors.lastname: Otsubo
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Ikeda
authors.forename: Akira
authors.initials: A
authors.lastname: Minami
authors.forename: Takashi
authors.initials: T
authors.lastname: Suzuki
mesh_terms: D001681:Biological Assay / Q000379:methods*; D005453:Fluorescence; D006801:Humans; D019158:N-Acetylneuraminic Acid / Q000378:metabolism; D009439:Neuraminidase / Q000378:metabolism*; D010222:Parainfluenza Virus 1, Human / Q000201:enzymology / Q000302:isolation & purification; D010253:Respirovirus Infections / Q000821:virology*; D013379:Substrate Specificity; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: D014764:Viral Proteins; D009439:Neuraminidase; D019158:N-Acetylneuraminic Acid