Canine adenovirus type-1 antibody ELISA detection kit and application thereof

A detection kit, canine adenovirus technology, applied in measurement devices, instruments, scientific instruments, etc., can solve the problems in the laboratory development stage (Jiang Lili et al., 2008; Ge Yanhua et al., 2010; Walker et al., etc., to improve solubility The effect of expression efficiency, high sensitivity and strong repeatability

Active Publication Date: 2019-12-13
INST OF SPECIAL ANIMAL & PLANT SCI OF CAAS
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Problems solved by technology

At present, most of the ELISA detection methods for CAdV-1 antibody are in the stage of laboratory development (Jiang Lili et al., 2008; Ge Yanhua et al., 2010; Walker et al., 2016), but commercialized CAdV-1 antibody ELISA detection reagents have not yet been released on the market box

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  • Canine adenovirus type-1 antibody ELISA detection kit and application thereof
  • Canine adenovirus type-1 antibody ELISA detection kit and application thereof
  • Canine adenovirus type-1 antibody ELISA detection kit and application thereof

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Embodiment 1

[0038] Preparation and screening of embodiment 1 coated antigen

[0039] 1. Test method

[0040] 1.1 Purification of CAdV-1 antigen by CsCl density gradient centrifugation

[0041] CAdV-1F1301 strain infected MDCK cells. After 24 hours, the culture medium was collected, frozen and thawed three times, centrifuged at 2000×g at 4°C for 10 minutes, and the culture medium was concentrated 20 times using a Millipore ultrafiltration tube (molecular weight cut-off 50KDa). Refer to the literature (Zou et al. ., 2018), CAdV-1 was purified using CsCl density gradient ultracentrifugation. Spike the concentrated viral supernatant at 1.25, 1.30 and 1.35 g / cm 3 In the CsCl gradient solution, centrifuge at 150000×g for 4h at 4°C, and collect 1.30-1.35g / cm 3 The virus band on the interface was dialyzed twice in a buffer containing 10mM Tris-Cl, 150mM NaCl and 5% glycerol (pH7.6), and the concentration of the purified CAdV-1 antigen was determined using a standardized BCA kit , stored at -8...

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Abstract

The invention discloses a canine adenovirus type-1 (CAdV-1) antibody ELISA detection kit and application thereof. According to the detection kit, a pCold II prokaryotic expression system is utilized to prepare a PB (penton base) and Knob protein with good reactogenicity, the PB and the Knob protein are subjected to a large quantity of induced expressions, two types of protein after renaturation are used as envelop antigens respectively and compared with a purified CAdV-1 totivirus, and finally the Knob protein is determined as the optimal ELISA envelop antigen. On this basis, the ELISA detection kit with high efficiency, good sensitivity and good specificity for a CAdV-1 serum antibody is provided; results of simultaneous sample detection through the detection kit and an SN method show that the sensitivity of the ELISA detection kit is 97.14%, the specificity is 90.00%, and the SN coincidence rate is 93.57%; and the detection kit has the advantages of high sensitivity, good specificity, strong repeatability and the like.

Description

technical field [0001] The invention relates to a canine adenovirus type 1 antibody detection kit, in particular to a canine adenovirus type 1 antibody ELISA detection kit and a preparation method thereof. The invention belongs to the field of canine adenovirus type 1 antibody ELISA detection kit. Background technique [0002] Canine adenovirus type 1 (CAdV-1) is a double-stranded DNA virus. CAdV-1 mainly causes hepatitis, nephritis (dogs) and encephalitis (bears and foxes). , otter) caused lethal infection (Jeong-Won et al., 2014; Lepczyk, 2015). CAdV-1 infection has been reported all over the world and has attracted widespread attention. The monitoring of CAdV-1 virus requires a detection method with strong specificity, high sensitivity and accuracy. The traditional detection methods of CAdV-1, such as virus isolation and identification, SN, HA-HI, etc., are time-consuming and labor-intensive, resulting in large error; although PCR and qPCR have high detection sensitiv...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569G01N33/558
CPCG01N33/56983G01N33/558Y02A50/30
Inventor 朱言柱闫喜军廉士珍
Owner INST OF SPECIAL ANIMAL & PLANT SCI OF CAAS
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