Medical Microbiology Laboratory, Department of Microbiology, Centre of Excellence in Life Sciences, Bharathidasan University, Tiruchirappalli 620 024, India; Amity Institute of Virology & Immunology (AIVI), Amity University Uttar Pradesh Campus, Sector-125, Noida 201313, India.
Amity Institute of Virology & Immunology (AIVI), Amity University Uttar Pradesh Campus, Sector-125, Noida 201313, India.
Int J Biol Macromol. 2016 Oct;91:465-70. doi: 10.1016/j.ijbiomac.2016.05.109. Epub 2016 May 31.
Leptospiral outer membrane protein LipL21 and its truncated N-terminal immunogenic region (I-LipL21) were evaluated for diagnosis of leptospirosis. The complete coding sequence of LipL21 nucleotide sequence was subjected to BCPred and VaxiJen analysis for determination of B cell specific immunogenic epitopes. Epitope1 ACS STD TGQ KDA TTV GDG (1.8837), Epitope2 WGG PPE QRN DGK TPR DTN (0.9483), Epitope3 VKG VGV YEC KAT GSG SDP (1.4077) and Epitope4 NEW ECQ CVI YAK FPG GKD (0.4462) were predicted. LipL21 and N-terminal fragment having B-cell specific epitopes with higher VaxiJen score >0.9 as truncated I-LipL21 were cloned independently in pET15b and expressed in Escherichia coli. IgM ELISA and dot blot assay was performed for sera samples collected from Delhi-NCR for leptospiral whole cell lysate (WCL), recombinant LipL21 and I-LipL21. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were found to be 92.5%, 92.8%, 83.3%, and 97% respectively for recombinant I-LipL21 by IgM-ELISA. 11-14.8% increased sensitivity was observed over LipL21 and WCL. The I-LipL21 dot blot assay showed a further increased sensitivity of 3.8% over the IgM-ELISA. Therefore I-LipL21 may be the ideal candidate protein for diagnosis of leptospirosis.
钩端螺旋体外膜蛋白 LipL21 及其截短的 N 端免疫原性区域（I-LipL21）被评估用于诊断钩端螺旋体病。LipL21 核苷酸序列的完整编码序列经过 BCPred 和 VaxiJen 分析，以确定 B 细胞特异性免疫原性表位。预测出 4 个表位：表位 1 ACS STD TGQ KDA TTV GDG（1.8837）、表位 2 WGG PPE QRN DGK TPR DTN（0.9483）、表位 3 VKG VGV YEC KAT GSG SDP（1.4077）和表位 4 NEW ECQ CVI YAK FPG GKD（0.4462）。LipL21 和具有更高 VaxiJen 评分>0.9 的 N 端片段被克隆到 pET15b 中，并在大肠杆菌中表达。针对来自德里-NCR 的血清样本进行了 IgM ELISA 和斑点印迹分析，用于检测钩端螺旋体全细胞裂解物（WCL）、重组 LipL21 和 I-LipL21。IgM-ELISA 对重组 I-LipL21 的灵敏度、特异性、阳性预测值（PPV）和阴性预测值（NPV）分别为 92.5%、92.8%、83.3%和 97%。与 LipL21 和 WCL 相比，灵敏度提高了 11-14.8%。I-LipL21 斑点印迹分析显示，灵敏度比 IgM-ELISA 进一步提高了 3.8%。因此，I-LipL21 可能是诊断钩端螺旋体病的理想候选蛋白。
