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弓形虫(Toxoplasma gondii,TOX)是胎儿先天性感染的重要病原体。本文用ELISA法检测脑性瘫痪(cere-bral palsy CP)患儿TOX感染的同时,结合使用聚合酶链反应(polymerase chain reaction,PCR)技术检测,现将结果报告如下。 资料 1.标本来源:186例脑瘫患儿均为本院门诊病儿,男107例,女79例,年龄1～8岁。1161例健康儿童均为来本院的体检者,男650例,女511例,年龄1～7岁。脑瘫的诊断标准参照文献。 2.方法:(1)ELISA法:TOX的ELISA检测试剂由浙江省医学科学院提供。循环抗原(CAg)检测使用双抗体一步法,IgM检测使用抗体捕捉法,IgG检测使用间接ELISA法。被检血清与阴性对照吸光度(OD)的比值≥2.1,重复检测,同样≥2.1,则判为阳性;OD的比值<2.1判为阴性。TOX的CAg、IgM和IgG,其中一项阳性,即认作TOX阳性。(2)PCR法:取肝素钠抗凝外周血200μl,用NH_4Cl低渗法溶解红细胞,剩白细胞加200μg/ml蛋白酶K和0.45％NP4050μl混匀,置55℃消化2小时,98℃水浴5分钟,离心取上清液做PCR。PCR
Toxoplasma gondii (TOX) is an important pathogen of fetal congenital infection. In this paper, the detection of TOX infection in children with cerebellar paralysis (cere-bral palsy CP) by ELISA method, combined with the use of polymerase chain reaction (PCR) detection, the results reported below. Data 1.Samples of origin: 186 cases of children with cerebral palsy are hospital outpatient, 107 males and 79 females, aged 1 to 8 years old. All 1161 healthy children were examined in our hospital. There were 650 men and 511 women aged 1 to 7 years old. The diagnostic criteria of cerebral palsy reference literature. 2. Methods: (1) ELISA method: TOX ELISA test reagents provided by the Zhejiang Academy of Medical Sciences. Antibody capture was used for the detection of circulating antigen (CAg) using the one-step diabodies, indirect ELISA for IgG detection. Serum was seized and negative control absorbance (OD) ratio ≥ 2.1, repeated testing, the same ≥ 2.1, then sentenced to positive; OD ratio of <2.1 sentence was negative. TOX of CAg, IgM and IgG, one of the positive, that is considered TOX positive. (2) PCR: take heparin anticoagulant peripheral blood 200μl, with NH_4Cl hypotonic solution of red blood cells, leukocytes plus 200μg / ml Proteinase K and 0.45% NP4050μl mix, set 55 ℃ digestion for 2 hours, 98 ℃ water bath for 5 minutes , The supernatant was centrifuged to PCR. PCR