Mitsui Y, Takamura N, Fujimaki Y, Yamaguchi T, Kitagawa T, Aoki Y
Institute of Tropical Medicine, Nagasaki University, Japan.
Trop Med Int Health. 1996 Aug;1(4):528-34. doi: 10.1046/j.1365-3156.1996.d01-77.x.
A sensitive and reproducible competitive enzyme-linked immunosorbent assay (ELISA) for the determination of the concentration of diethylcarbamazine (DEC) in biological fluids was developed. Since DEC has no functional group to conjugate with bovine serum albumin (BSA), N-(2-aminoethyl)-N-ethyl-4-methyl-1-piperazinecarboxamide (DEC-NH2) was first synthesized. This compound was then converted to carboxyl DEC (DEC-COOH) and conjugated to BSA and to poly-L-lysine for use as immunogen and solid-phase marker, respectively. The competitive ELISA was conducted by simultaneously incubating DEC with mouse anti-DEC antiserum over DEC-poly-L-lysine solid phase. Subsequently, the binding of anti-DEC antibody was detected by using sheep anti-mouse IgG peroxidase conjugate as a tracer. The reliability, determined by the coefficient of variation for inter and intra-assay, was satisfactory. The cross-reactivities of anti-DEC antibodies with DEC metabolites, related compounds and ivermectin were negligible. Using this assay, DEC levels were easily determined in serum of Mongolian jirds (Meriones unguiculatus) up to 4 hours following a single dose of DEC citrate base (100 mg/kg of body weight) via intraperitoneal route.
建立了一种灵敏且可重复的竞争性酶联免疫吸附测定法（ELISA），用于测定生物体液中二乙碳酰嗪（DEC）的浓度。由于DEC没有可与牛血清白蛋白（BSA）偶联的官能团，因此首先合成了N-（2-氨基乙基）-N-乙基-4-甲基-1-哌嗪甲酰胺（DEC-NH2）。然后将该化合物转化为羧基DEC（DEC-COOH），并分别与BSA和聚-L-赖氨酸偶联，用作免疫原和固相标记物。竞争性ELISA通过将DEC与小鼠抗DEC抗血清在DEC-聚-L-赖氨酸固相上同时孵育来进行。随后，使用羊抗小鼠IgG过氧化物酶偶联物作为示踪剂检测抗DEC抗体的结合。通过批内和批间变异系数确定的可靠性令人满意。抗DEC抗体与DEC代谢物、相关化合物和伊维菌素的交叉反应可忽略不计。使用该测定法，通过腹腔注射单剂量柠檬酸DEC碱（100mg/kg体重）后，在长达4小时的时间内可轻松测定长爪沙鼠（Meriones unguiculatus）血清中的DEC水平。
