シトリニンに対するモノクローナル抗体の作製

概要

シトリニン（CIT）は，腎毒性を有するマイコトキシンで，麦類やチーズやベニコウジカビ色素などを汚染している．日本ではベニコウジ色素中CIT濃度200 µg/kg以下との規格が定めされている．そこで，高感度かつ簡便なCITの測定法の開発を目的として抗CITモノクローナル抗体の作製を行った．CIT-KLHでBALB/cマウスとCB6F1マウスを各12匹をそれぞれ3回免疫した結果，それぞれのマウスで1匹ずつ2匹のマウスでCITに対する抗体価の上昇を確認した．抗体価の上昇の認められたマウスの脾臓細胞を常法で細胞融合を行った．HAT選択後，クローニングを行い安定な抗CITモノクローナル抗体産生ハイブリドーマCIT.6とCIT.7を得た．CIT.6抗体を用いた至適化した競合的間接ELISAで0.3 µg/mLまでのCITを検出可能であった．本法を用いて，食品中などのCITの高感度かつ簡易検査法の確立が期待される．
Citrinin (CIT) is a nephrotoxic mycotoxin that contaminates barley, cheese, and Monascus color. In Japan, there is the regulation that the CIT concentration in Monascus color is less than 200 µg/kg. Therefore, we prepared anti-CIT monoclonal antibody for the purpose of developing a highly sensitive and simple analyzed method for CIT in these. Twelve BALB/c mice and 12 CB6F1 mice were immunized with CIT-KLH three times each, and as a result, an increase in the antibody titer against CIT was confirmed in each of the one mouse. The mouse spleen cells were fused by standard methods. After HAT selection and cloning, we obtained stable anti-CIT monoclonal antibody-producing hybridomas CIT.6 and CIT.7. Optimized competitive indirect ELISA using CIT.6 antibody was able to detect CIT up to 0.3 µg/mL. It is expected that a highly sensitive and simple analyzed method for CIT in foods will be established.