用于I型病毒性出血性败血症病毒的抗原捕获酶联免疫吸附测定

Antigen-capture ELISA for viral haemorrhagic septicaemia virus serotype I.

作者信息

Mourton C, Bearzotti M, Piechaczyk M, Paolucci F, Pau B, Bastide J M, de Kinkelin P

机构信息

Unité de Recherche en Immunologie, Faculté de Pharmacie, Montpellier, France.

出版信息

J Virol Methods. 1990 Sep;29(3):325-33. doi: 10.1016/0166-0934(90)90059-o.

DOI:10.1016/0166-0934(90)90059-o

PMID:2266147

Abstract

An antigen-capture ELISA for viral haemorrhagic septicaemia virus serotype I (VHSV I) was developed. The assay employs two monoclonal antibodies (mAb) directed against distinct epitopes of the viral envelope glycoprotein (Gp). The antigen bound by the capture mAb (A17) was detected by addition of a second mAb (L7) conjugated to horseradish peroxidase, followed by addition of the enzyme substrate. The technique is highly sensitive, enabling detection of the virus at a protein concentration as low as 1 ng/ml of total proteins (1.5 fmol of envelope Gp per ml) in purified preparations. VHSV I was also detected in culture supernatants (5 x 10(5) PFU/ml) and in extracts of kidney and spleen of rainbow trout infected experimentally (5 x 10(5) PFU/ml). The assay was highly specific: infectious haematopoietic necrosis virus, infectious pancreatic necrosis virus, spring viraemia of carp virus, pike fry rhabdovirus, eel rhabdovirus and perch rhabdovirus could not be detected by the antigen-capture ELISA for VHSV I.

摘要

开发了一种用于检测I型病毒性出血性败血症病毒（VHSV I）的抗原捕获ELISA。该检测方法使用了两种针对病毒包膜糖蛋白（Gp）不同表位的单克隆抗体（mAb）。通过加入与辣根过氧化物酶偶联的第二种单克隆抗体（L7），然后加入酶底物，来检测捕获单克隆抗体（A17）所结合的抗原。该技术高度灵敏，能够在纯化制剂中检测到低至1 ng/ml总蛋白（每毫升1.5 fmol包膜Gp）浓度的病毒。在培养上清液（5×10⁵ PFU/ml）以及实验感染的虹鳟肾和脾提取物（5×10⁵ PFU/ml）中也检测到了VHSV I。该检测方法具有高度特异性：传染性造血坏死病毒、传染性胰腺坏死病毒、鲤春病毒血症病毒、梭鲈弹状病毒、鳗鲡弹状病毒和鲈弹状病毒不能通过用于VHSV I的抗原捕获ELISA检测到。