title: Development of lectin-linked immunomagnetic separation for the detection of hepatitis a virus.
abstract: The accuracy and sensitivity of PCR-based methods for detection of hepatitis A virus (HAV) are dependent on the methods used to separate and concentrate the HAV from the infected cells. The pH and ionic strength affect the binding affinity of the virus to cells. In this study, we initially investigated the effects of pH (4.0-10.0) and metal ions (Fe²⁺, Co²⁺, Cu²⁺, Mg²⁺, K⁺, and Ca²⁺) on the binding of HAV to oyster digestive cells. The lowest relative binding (RB) of HAV to the cells was found at pH 4.0 and in FeSO₄ solution (64.6% and 68.1%, respectively). To develop an alternative to antibody-dependent immunomagnetic separation prior to detection of HAV using RT-PCR, the binding of HAV to five lectins, peanut agglutinin (PNA), Dolichos biflorus agglutinin (DBA), Helix pomatia agglutinin (HPA), Ulex europaeus agglutinin (UEA-1) and soybean agglutinin (SBA), was evaluated using ELISAs. SBA showed significantly higher RB to HAV than the other lectins tested. In addition, HAV could be concentrated within 30 min using SBA-linked magnetic bead separation (SMS) prior to the RT-PCR assay. Our findings demonstrate the feasibility of using SMS combined with RT-PCR to detect HAV at dilutions ranging from 10⁻¹-10⁻⁴ of a HAV stock (titer: 10⁴ TCID₅₀/mL).
authors.affiliation: Department of Aqualife Medicine, Chonnam National University, Yeosu, Jeonnam 550-749, Korea. rhtkdan7@naver.com.
authors.forename: Sang-Mu
authors.initials: SM
authors.lastname: Ko
authors.affiliation: Korea Basic Science Institute, Daejeon 305-806, Korea. joseph@kbsi.re.kr.
authors.forename: Joseph
authors.initials: J
authors.lastname: Kwon
authors.affiliation: Department of Food Science and Technology and Functional Food Research Center, Chonnam National University, Gwangju 500-757, Korea. bipinvaidya@hotmail.com.
authors.forename: Bipin
authors.initials: B
authors.lastname: Vaidya
authors.affiliation: Korea Basic Science Institute, Daejeon 305-806, Korea. jschoi@kbsi.re.kr.
authors.forename: Jong Soon
authors.initials: JS
authors.lastname: Choi
authors.affiliation: Department of Food Science and Technology and Functional Food Research Center, Chonnam National University, Gwangju 500-757, Korea. lhmein@naver.com.
authors.forename: Hee-Min
authors.initials: HM
authors.lastname: Lee
authors.affiliation: Department of Aqualife Medicine, Chonnam National University, Yeosu, Jeonnam 550-749, Korea. ohmj@jnu.ac.kr.
authors.forename: Myung-Joo
authors.initials: MJ
authors.lastname: Oh
authors.affiliation: Department of Forest Products and Technology, Chonnam National University, Gwangju 500-757, Korea. baehj@jnu.ac.kr.
authors.forename: Hyeun-Jong
authors.initials: HJ
authors.lastname: Bae
authors.affiliation: Department of Food Science and Technology and Functional Food Research Center, Chonnam National University, Gwangju 500-757, Korea. sycho5633@gmail.com.
authors.forename: Se-Young
authors.initials: SY
authors.lastname: Cho
authors.affiliation: Department of Food Science and Technology and Functional Food Research Center, Chonnam National University, Gwangju 500-757, Korea. oks665@naver.com.
authors.forename: Kyung-Seo
authors.initials: KS
authors.lastname: Oh
authors.affiliation: Department of Food Science and Technology and Functional Food Research Center, Chonnam National University, Gwangju 500-757, Korea. dwkim@jnu.ac.kr.
authors.forename: Duwoon
authors.initials: D
authors.lastname: Kim
mesh_terms: D006506:Hepatitis A / Q000175:diagnosis*; D030041:Hepatitis A virus / Q000302:isolation & purification*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D018189:Immunomagnetic Separation / Q000379:methods*; D037102:Lectins / Q000378:metabolism*; D008670:Metals / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013048:Specimen Handling / Q000379:methods*; D013997:Time Factors; D053585:Virus Attachment
chemical_list: D037102:Lectins; D008670:Metals