微量滴度酶标板法测定棉蚜(Aphis gossypii Glover)酯酶特性与有机磷抗性的关系

A 96-Well Microplate Reader for Measuring Characterization of Esterase and Insecticide Resistance in the Cotton Aphids (Aphis gossypii Glover)

通过生物测定表明高密棉蚜对有机磷的抗性高于北京棉蚜 ,用紫外分光光度计比色法 (A法 )及微量滴度酶标板法 (B法 )测定高密棉蚜及北京棉蚜的 α-乙酸萘酯酯酶活力和α-乙酸萘酯酯酶动力学。北京棉蚜和高密棉蚜的 α- NA酯酶活力分别为 2 .2 3、4 .4 8(A法 )和1.13、3.30 (B法 ) μmol· mg-1pro.· min-1,高密、北京棉蚜的酶活之比为 2 .0 0 (A法 )、2 .92 (B法 ) ;北京棉蚜、高密棉蚜的 K m值分别为 :6 .0 6× 10 -5、7.51× 10 -5(A法 )和 7.6 6× 10 -5、8.87× 10 -5(B法 ) mol· L-1,Vmax 值为 2 .53、 5.82 (A 法 )和 1.2 8、 3.6 1(B法 )μmol· mg-1· min-1。比较紫外分光光度计比色法及微量滴度酶标板法的测定结果 ,表明微量滴度酶标板法的测定结果是可靠的。

The bioassay showed that Gaomi cotton aphid had higher resistance to organophosphate insecticides than Beijing cotton aphid. Using two approaches, a conventional spectrophotometer (method A) and a 96 well microplate reader method (method B),to detect general α naphthyl acetate(α NA) esterase activity and kinetics of α NA esterase of Gaomi and Beijing cotton aphid. The α NA esterase activity of Beijing and Gaomi cotton aphid were 2.23 and 4.48 (method A),1.28 and 3.30 μmol·mg -1 protein·min -1 (method B),respectively; The α NA esterase activity ratio of G/B were 2.00 and 2.56; K m of α NA esterase in Beijing and Gaomi cotton aphid were 6.06×10 -5 and 7.51×10 -5 (method A),7.66×10 -5 and 8.87×10 -5 (method B) mol·L -1 , respectively; V max were 2.53 and 5.82 (method A),1.28 and 3.61(method B) μmol·mg -1 protein·min -1 , respectively. Data obtained from the 96 well microplate reader were compared to those obtained, under similar conditions, in a conventional spectrophotometer. The approach of using the 96 well microplate reader should be generally applicable to monitor pest resistance to insecticide.