黃立平 豬圓環(huán)病毒型檢測技術課件

1、豬圓環(huán)病毒2型檢測技術報告人：黃立平 博士哈爾濱獸醫(yī)研究所2014-4-29獸醫(yī)生物技術及獸醫(yī)免疫的進展與應用豬病論壇PCV2簡介-PCV2的形態(tài)及其基因組無囊膜，20面體對稱，17nmntPCV2簡介-Cap蛋白的結構PCV2簡介-PCV2的致病性 （PMWS） （PDNS） （PRDC） （RF）PCV2相關疾病的防控免疫效力：特異性抗體/檢測方法抗原含量：檢測方法/毒價測定/抗原捕獲ELISA疫苗免疫PCV2抗體檢測方法免疫過氧化物酶單層細胞試驗（IPMA）間接免疫熒光（IFA）膠體金免疫層析檢測試紙條（GICA）ELISA 間接ELISA：PCV2 (荷蘭BIOCHEK, 美國synb

2、iotics，西班牙Ingenasa)；PCV2-Cap；PCV2-dCap(武漢科前,瑞普) 競爭ELISA：基于中和性單抗IFAIPMAELISAGICA敏感性高、特異性好敏感性高、特異性好高通量和自動化操作簡便，快速，使用于基層成本高、反應板需冷凍保存反應板需冷凍保存無法現(xiàn)場診斷無法定量檢測PCV2-Cap單克隆抗體的制備和鑒定PCV2a/JF1PCV2競爭ELISA抗體檢測試劑盒的原理1 包被PCV2陽性血清2 捕獲 病毒3 與被檢血清抗體結合4 與酶標單體反應及顯色 Harbin Veterinary Research Institute, CAAS, ChinaThe cut-of

3、f value of the C-ELISA PI 45.6% was considered positive. PI35.13% was considered negative. Harbin Veterinary Research Institute, CAAS, ChinaConinsidence of C-ELISA and SNA for 132 sera from immunnization and chanllenge pigsDetection methodsSerum neutralizing testTesting positive numbersTesting negat

4、ive numbersTotalC- ELISATesting positive numbers79685Testing negative numbers14647Total8052132Using serum neutralization assay as a reference standard, the sensitivity of C-ELISA was 98.8% (79/80) and specificity was 88.5% (46/52) .試劑盒試劑盒名稱試劑盒 APCV2抗體酶聯(lián)免疫檢測試劑盒試劑盒 BPCV2-dCap-ELISA抗體檢測試劑盒試劑盒 CPCV2-ELI

5、SA抗體檢測試劑盒試劑盒 DPCV2競爭ELISA抗體檢測試劑盒試劑盒 EIngezim Circo IgG 1 PCV K.1 ELISA kitPCV2標準陽性和陰性血清：IPMA法標定不同效價的標準PCV2陽性血清50份，PCV2陰性血清30份 PCV2 C-ELISA 抗體檢測試劑盒和商品化試劑盒的比較試驗Comparison for sensitivity, specificity and coincidence rate of five kinds of kitsDetection itemsThe PCV2 antibody detection kitsKit AKit BKit

6、 CC-ELISAKit ESensitivity76.0% (38/50)74.0% (37/50)82.0% (41/50)92.0% (46/50)96.0% (48/50)Specificity93.3% (28/30)83.3% (25/30)83.3% (25/30)90.0% (27/30)96.7% (29/30)Consistence rate82.5% (66/80)77.5% (62/80)82.5% (66/80)91.3% (73/80)96.3% (77/80) Harbin Veterinary Research Institute, CAAS, ChinaSer

7、ological profile of anti-PCV2 neutralizing antibodies from immunization and challenge tests by C-ELISA Challenge with PCV21st immunizationBoost Harbin Veterinary Research Institute, CAAS, ChinaSerological profile of anti-PCV2 neutralizing antibodies from a commercial pig farm by C-ELISAPCV2競爭ELISA保存期試驗PCV2競爭ELISA抗體檢測試劑盒PCV2相關疾病的防控免疫效力：特異性抗體/檢測方法抗原檢測：PCR/毒價測定/capture-ELISA疫苗免疫PCV2抗原捕獲ELISA檢測試劑盒的原理1 包被PCV2陽性血清2 捕獲 病毒3 與HRP-單抗結合4 顯色EEEEEEPCV2抗原捕獲ELISA和PCV2毒價測定結果相關性分析Cut-off valuePCV2抗原捕獲ELISA的最小檢出量是400TCID50/ml。PCV2抗原捕獲ELISA檢測試劑盒特異性PCV2抗原捕獲ELISA試