Reagent kit for detecting immunoglobulin m (IgM) AlphaLISA for resisting enterovirus71 capsid protein1

A technology of capsid protein and enterovirus, which is applied in the field of immunological detection of enterovirus 71, can solve problems such as difficult application, and achieve the effect of high sensitivity, good specificity, and less serum consumption

Inactive Publication Date: 2012-07-18
中国疾病预防控制中心病毒病预防控制所
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Since there is no effective antiviral treatment for EV71, the current measures to prevent and treat hand, foot and mouth disease mainly rely on the rapid diagnosis of the disease and timely prevention of the epidemic of the virus. The laboratory diagnosis of EV71 mainly relies on the reverse transcription polymerization based on the viral genome Enzyme chain reaction (RT-PCR) and virus isolation test, however, these methods are difficult to be fully applied in most primary clinics and hospitals

Method used

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  • Reagent kit for detecting immunoglobulin m (IgM) AlphaLISA for resisting enterovirus71 capsid protein1
  • Reagent kit for detecting immunoglobulin m (IgM) AlphaLISA for resisting enterovirus71 capsid protein1
  • Reagent kit for detecting immunoglobulin m (IgM) AlphaLISA for resisting enterovirus71 capsid protein1

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Experimental program
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Embodiment

[0015] 1. Construction and identification of EV71 VP1 recombinant protein prokaryotic expression plasmid

[0016] 1 target fragment amplification

[0017] 1.1 PCR cycle amplification

[0018] Using the genome of strain EV71 / Fuyang.Anhui.CHU / 17.08 / 1 (GenBank No.: EU703812) as a template, perform reverse transcription, and amplify the desired fragment EV71 VP1 1 by conventional polymerase chain reaction (PCR) -297 (GenBank Number: HM579945.1), its nucleotide sequence is shown in Seq ID No.2, and the encoded amino acid sequence is shown in Seq ID No.1.

[0019] The upstream and downstream primers were respectively introduced with Nde I and Xho I restriction sites. The primers were designed using Primer Premier 5 (a primer design software) software. The primers were synthesized by Shanghai Sangon Company. The primer sequences are shown in Table 1.

[0020] Table 1 Primer Sequence

[0021]

[0022] The PCR reaction system is shown in Table 2 (total volume 50 μl):

[0023] Ta...

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Abstract

The invention provides a reagent kit for detecting immunoglobulin m (IgM) AlphaLISA for resisting enterovirus71 capsid protein1, which comprises donor microbeads, receptor microbeads and biotinylated enterovirus71 capsid protein1 antibodies. Optimal dose of biotinylated vitamin P (VP)1 is fished out to be 37.5ng through the enzyme-linked immuno sorbent assay (ELISA), and a detecting method is built based on the IgM AlphaLISA for resisting enterovirus71 (EV71) VP1 through optimization of the donor microbeads, the receptor microbeads, blood serum and other test reaction conditions. The reagent kit is used for detecting the enterovirus71, is good in specificity, high in sensitivity and small in blood serum dosage, does not need to be washed, and cannot be influenced by hemolysis.

Description

technical field [0001] The invention relates to the immunological detection of enterovirus 71, in particular to an anti-enterovirus 71 capsid protein 1 IgM AlphaLISA detection kit. Background technique [0002] Enterovirus 71 (EV71) is the main pathogen that infects infants with hand-foot-mouth disease (HFMD). Symptoms following EV71 infection include aseptic meningitis, brainstem encephalitis, and polio-like paralysis. The main feature of the disease is a high mortality rate within a short period of time. Since there is no effective antiviral treatment for EV71, the current measures to prevent and treat hand, foot and mouth disease mainly rely on the rapid diagnosis of the disease and timely prevention of the epidemic of the virus. The laboratory diagnosis of EV71 mainly relies on the reverse transcription polymerization based on the viral genome Enzyme chain reaction (RT-PCR) and virus isolation test, but these methods are difficult to obtain full application in most pri...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569
Inventor 董小平韩俊陈操高晨
Owner 中国疾病预防控制中心病毒病预防控制所
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