development of sensitive detection method for the pathogenic bacteria in food using a labo-disc

2019 Fiscal Year Final Research Report

• Project/Area Number: 17K07825
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Food science
• Research Institution: Soka University
• Principal Investigator: Kubo Izumi 創価大学, 理工学部, 教授 (40214986)
• Project Period (FY): 2017-04-01 – 2020-03-31
• Keywords: 抗体固定化ビーズ / 抗サルモネラ抗体 / 卵黄 / PCR / ラボディスク

Outline of Final Research Achievements

Salmonella enterica is one of the dangerous pathogens. Rapid detection methods are required to prevent its infection. In our laboratory, a microfluidic disc that enabled isolation of cells has been developed. It is possible to isolate S.enterica cells, amplify the specific invA gene by PCR, and detect it by fluorescence probe in the microchambers on the disc. In this study, we demonstrate the collection method of S.enterica from egg yolk that can remove egg yolk components, and the rapid detection of collected S.enterica by PCR on the microfluidic disc. The S.enterica cells were collected from egg yolk by using antibody-coated immunomagnetic beads. S.enterica collected using beads was detected by PCR on the disc of each microchamber. We could detect 50000 cells/ml or higher concentration of S. enterica within 6 hour. Effectiveness of the antibody modified plastic beads was investigated to detect S. enterica from egg yolk, too. With the plastic beads, 5000 cells/ml was detectable.

Free Research Field

分析化学

Academic Significance and Societal Importance of the Research Achievements

食中毒菌の検出を培養による増菌を行わず、抗体固定化ビーズによる濃縮を利用し、このビーズをそのままマイクロチャンバーに導入して菌固有遺伝子を増幅する方法を確立することで、大幅な迅速化を図る点が本研究の特色である。培養法では実際に多種多様な菌が混在する食品試料では、増菌しやすい菌に培地中の栄養を奪われて、食中毒の原因菌が十分に増菌せず、検出できていない場合もあったと考えられる。本研究では抗体固定化ビーズでサルモネラ菌をビーズ上に濃縮でき、それによって、培養法より短時間での測定が可能なPCR法よりもさらに高感度に検出できる可能性が示された。