目的:介绍25-羟基维生素D的临床背景,并对复星长征公司研制生产的25-羟基维生素D测定试剂盒(克隆酶供体免疫测定法)进行初步方法学评价,为临床应用提供依据。方法:依据《临床化学体外诊断试剂(盒)通用技术要求》对25-羟基维生素D测定试...目的:介绍25-羟基维生素D的临床背景,并对复星长征公司研制生产的25-羟基维生素D测定试剂盒(克隆酶供体免疫测定法)进行初步方法学评价,为临床应用提供依据。方法:依据《临床化学体外诊断试剂(盒)通用技术要求》对25-羟基维生素D测定试剂盒(克隆酶供体免疫测定法)进行分析性能评估分析与验证,并评价其与其他厂家试剂的相关性。结果:试剂启用后可稳定2周;与英国Immunodiagnostic Systems Limited(IDS Ltd)公司生产的试剂测试结果相关性良好,其相关系数为0.9680。结论:复星长征公司研制生产的用克隆酶供体免疫测定法检测25-羟基维生素D试剂盒各项分析性能完全能够达到指标要求,其检测结果可为临床诊断提供科学依据。展开更多
Objective:Evaluation of a polyclonal and a monoclonal Helicobacter pylori stool antigen test(HpSAT)for the detection of Helicobacter pylori(HP)infection in children.Methods:43 children underwent a 13C-urea breath test...Objective:Evaluation of a polyclonal and a monoclonal Helicobacter pylori stool antigen test(HpSAT)for the detection of Helicobacter pylori(HP)infection in children.Methods:43 children underwent a 13C-urea breath test(13C UBT)and a HpSAT for the detection of HP.A child was considered HP positive if the 13C UBT was positive.Two HpSATs were tested:the “Premier Platinum HpSA”test(PP HpSAT)(polyclonal enzyme immunoassay)and the “ImmunoCard STAT! HpSA”test(ICS HpSAT)(one-step immunochromatographic assay).Results:The mean age of the children was 8.9 y(range 3.5-17.5 y).Of the 43 children,18(41.9%)were HP positive and 25(58.1%)were HP negative.The PP HpSAT showed a sensitivity of 94.4%and a specificity of 100.0%.With the ICS HpSAT,equivocal results occurred in 5/42(11.9%)of the tests due to a problem with the visual interpretation of the change in colour of the test-line.The ICS HpSAT had a sensitivity of 100.0%and a specificity of 76.0%when the test was considered positive in case of any change of colour of the test-line,which is the correct practice according to the manufacturer.The ICS HpSAT had a sensitivity of 100.0%and a specificity of 96.0%when the test was considered positive only in case of a “significant change of colour”of the test-line.Conclusion:Compared to the 13C UBT,the PP HpSAT shows a comparably good sensitivity and specificity,the ICS HpSAT has a comparably good sensitivity but lower specificity due to a high percentage of equivocal results when the test is used according to the manufacturer’s instructions,and the ICS HpSAT has a comparably good sensitivity and specificity when a weakly positive test is considered negative for the diagnosis of HP infection in children.The ICS HpSAT is easy to perform with results available within 10 min,and is therefore of particular interest in ambulatory medicine.展开更多
目的:制备大肠杆菌β-半乳糖苷酶(β-galactosidae)酶供体(ED)和酶受体(EA)片段,获得具有全酶活性的β-半乳糖苷酶。方法:以pSV-β-galactosidase control vector为模板设计引物,用PCR方法获得ED和EA片段DNA序列,插入克隆载体pGEM-T-eas...目的:制备大肠杆菌β-半乳糖苷酶(β-galactosidae)酶供体(ED)和酶受体(EA)片段,获得具有全酶活性的β-半乳糖苷酶。方法:以pSV-β-galactosidase control vector为模板设计引物,用PCR方法获得ED和EA片段DNA序列,插入克隆载体pGEM-T-easy中,获得重组质粒,经酶切、PCR及测序鉴定正确后,将酶切目的片段分别插入原核表达载体pET20b+,并转化大肠杆菌BL21,通过IPTG诱导表达出ED、EA融合蛋白,以亲和层析纯化蛋白,通过ED与EA蛋白形成β-半乳糖苷酶、全酶活性试验来判断ED、EA的功能。结果:获得与pSV-β-galactosidase control vector一致的ED、EA碱基序列,构建了重组表达质粒pET20b-ED及pET20b-EA,并分别转化入大肠杆菌DE3,以IPTG诱导行SDS-PAGE电泳,在相对分子质量为14000及116000处分别可见ED蛋白和EA蛋白条带,应用镍凝胶亲和层析纯化获得目的蛋白。结论:成功地制备了ED、EA蛋白,形成全酶活性的β-半乳糖苷酶。展开更多
文摘目的:介绍25-羟基维生素D的临床背景,并对复星长征公司研制生产的25-羟基维生素D测定试剂盒(克隆酶供体免疫测定法)进行初步方法学评价,为临床应用提供依据。方法:依据《临床化学体外诊断试剂(盒)通用技术要求》对25-羟基维生素D测定试剂盒(克隆酶供体免疫测定法)进行分析性能评估分析与验证,并评价其与其他厂家试剂的相关性。结果:试剂启用后可稳定2周;与英国Immunodiagnostic Systems Limited(IDS Ltd)公司生产的试剂测试结果相关性良好,其相关系数为0.9680。结论:复星长征公司研制生产的用克隆酶供体免疫测定法检测25-羟基维生素D试剂盒各项分析性能完全能够达到指标要求,其检测结果可为临床诊断提供科学依据。
文摘Objective:Evaluation of a polyclonal and a monoclonal Helicobacter pylori stool antigen test(HpSAT)for the detection of Helicobacter pylori(HP)infection in children.Methods:43 children underwent a 13C-urea breath test(13C UBT)and a HpSAT for the detection of HP.A child was considered HP positive if the 13C UBT was positive.Two HpSATs were tested:the “Premier Platinum HpSA”test(PP HpSAT)(polyclonal enzyme immunoassay)and the “ImmunoCard STAT! HpSA”test(ICS HpSAT)(one-step immunochromatographic assay).Results:The mean age of the children was 8.9 y(range 3.5-17.5 y).Of the 43 children,18(41.9%)were HP positive and 25(58.1%)were HP negative.The PP HpSAT showed a sensitivity of 94.4%and a specificity of 100.0%.With the ICS HpSAT,equivocal results occurred in 5/42(11.9%)of the tests due to a problem with the visual interpretation of the change in colour of the test-line.The ICS HpSAT had a sensitivity of 100.0%and a specificity of 76.0%when the test was considered positive in case of any change of colour of the test-line,which is the correct practice according to the manufacturer.The ICS HpSAT had a sensitivity of 100.0%and a specificity of 96.0%when the test was considered positive only in case of a “significant change of colour”of the test-line.Conclusion:Compared to the 13C UBT,the PP HpSAT shows a comparably good sensitivity and specificity,the ICS HpSAT has a comparably good sensitivity but lower specificity due to a high percentage of equivocal results when the test is used according to the manufacturer’s instructions,and the ICS HpSAT has a comparably good sensitivity and specificity when a weakly positive test is considered negative for the diagnosis of HP infection in children.The ICS HpSAT is easy to perform with results available within 10 min,and is therefore of particular interest in ambulatory medicine.
文摘目的:制备大肠杆菌β-半乳糖苷酶(β-galactosidae)酶供体(ED)和酶受体(EA)片段,获得具有全酶活性的β-半乳糖苷酶。方法:以pSV-β-galactosidase control vector为模板设计引物,用PCR方法获得ED和EA片段DNA序列,插入克隆载体pGEM-T-easy中,获得重组质粒,经酶切、PCR及测序鉴定正确后,将酶切目的片段分别插入原核表达载体pET20b+,并转化大肠杆菌BL21,通过IPTG诱导表达出ED、EA融合蛋白,以亲和层析纯化蛋白,通过ED与EA蛋白形成β-半乳糖苷酶、全酶活性试验来判断ED、EA的功能。结果:获得与pSV-β-galactosidase control vector一致的ED、EA碱基序列,构建了重组表达质粒pET20b-ED及pET20b-EA,并分别转化入大肠杆菌DE3,以IPTG诱导行SDS-PAGE电泳,在相对分子质量为14000及116000处分别可见ED蛋白和EA蛋白条带,应用镍凝胶亲和层析纯化获得目的蛋白。结论:成功地制备了ED、EA蛋白,形成全酶活性的β-半乳糖苷酶。
