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Human parainfluenza virus quantum dot immunochromatography typing detection card, preparation method and applications

A technology of human parainfluenza virus and immune chromatography, which is applied in the field of medical testing, can solve the problems of easy false positive, complicated operation steps and high cost

Active Publication Date: 2016-01-27
湖北诺美华抗体药物技术有限公司
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AI Technical Summary

Problems solved by technology

Neither immunofluorescence method nor immunoenzyme method can perform one-step detection, and both have the disadvantages of complicated operation steps, professional operation, long detection time (more than 2 hours), and high cost.
The PCR method has the advantages of rapidity, sensitivity and specificity, and is an important method for studying HPIV infection at present. However, due to the high requirements for experimental equipment and operation of PCR, and the possibility of false positives, it cannot be used as a commonly used clinical diagnosis method in my country.

Method used

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  • Human parainfluenza virus quantum dot immunochromatography typing detection card, preparation method and applications
  • Human parainfluenza virus quantum dot immunochromatography typing detection card, preparation method and applications
  • Human parainfluenza virus quantum dot immunochromatography typing detection card, preparation method and applications

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Experimental program
Comparison scheme
Effect test

preparation example Construction

[0084] 1. Preparation of conjugated pads

[0085] (1) Preparation, purification and renaturation of recombinant HN1-His, HN2-His, HN3-His fusion proteins

[0086] Bioinformatics analysis was performed on the HN proteins of human parainfluenza viruses I, II, and III to obtain the peptides with the most abundant antigenic epitopes in their extracellular domains, and to find their corresponding gene coding sequences. According to the preference of codons, after codon optimization, restriction sites were introduced into the 5' and 3' ends of the sequence, and the whole gene sequences were chemically synthesized, which were denoted as hn1, hn2, and hn3. See the sequence listing for their gene sequences. The three gene sequences were respectively cloned into the expression vector pET-28a(+) according to conventional methods, and then the recombinant fusion proteins HN1-His, HN2-His and HN3-His were expressed. The three recombinant proteins all exist in the form of inclusion bodies...

Embodiment 1

[0132] Embodiment 1 (preparation embodiment)

[0133] Conjugate pad preparation

[0134] (1) Preparation, purification and renaturation of recombinant HN1-His, HN2-His, HN3-His fusion proteins

[0135] (1) Cloning of related genes

[0136] Bioinformatics analysis was performed on the HN proteins of human parainfluenza virus types I, II, and III (the accession numbers in the NCBI protein database are AAC23946.1, BAA00739.1, and ACF28540.1, respectively), and the antigens in their extracellular domains were respectively obtained Find the corresponding DNA coding sequence for the peptide with the most abundant epitope, and then optimize its codon according to the codon preference of Escherichia coli, and introduce a restriction site NdeI at the 5' end and a termination signal at the 3' end After TAA and enzyme cutting site XhoI, chemically synthesize the whole gene sequence respectively (the whole sequence synthesis is handed over to GenScript Biotechnology Co., Ltd., and the a...

Embodiment 2

[0166] Embodiment 2 (preparation embodiment)

[0167] Preparation of sample pads

[0168]Prepare sample pad treatment solutions with different formulations, observe the release effect of quantum dot-labeled antibodies, and optimize through multiple orthogonal experiments to obtain the optimal sample pad treatment solution formulation (that is, described in the present invention). Take a piece of glass cellulose membrane, soak it in the sample pad treatment solution for at least 3 hours, then place it in a biological safety cabinet at 37°C, ventilate and dry it, and cut it into a size of 4cm*2.5cm / strip to prepare the sample pad , Store in airtight and dry place at 25°C. Tests have proved that the use of the sample pad greatly improves the release rate of the quantum dot-labeled antibody on the binding pad and achieves a better application effect.

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Abstract

The invention provides a human parainfluenza virus quantum dot immunochromatography typing detection card, a preparation method and applications. The detection card comprises a base plate, a sample pad, a water absorption pad, a combination pad and a detection layer. The combination pad is coated with a mixture of rabit-anti I-type, II-type and III-type human parainfluenza virus HN protein polyclonal antibodies labeled with quantum dots respectively. The detection layer is composed of a solid phase nitrocellulose membrane with three detection lines and a quality control line. The three detection lines are coated with rabit-anti I-type, II-type and III-type human parainfluenza virus polyclonal antibodies respectively. The quality control line is coated with anti-rabit IgG. The detection layer is pasted on the base plate. The combination pad and the water absorption pad are arranged above two ends of the detection layer respectively, overlap with part of the detection layer and are pasted with the detection layer and the base plate respectively. The sample pad is arranged on the combination pad, overlaps with part of the combination pad and is pasted with the combination pad and the base plate. The provided detection card has advantages of simple operation, rapid detection, quantification, high sensitivity and the like.

Description

technical field [0001] The invention relates to the technical field of medical detection, in particular to a human parainfluenza virus quantum dot immune chromatography typing detection card and its preparation method and application. Background technique [0002] Human parainfluenza virus (HPIV) is an important pathogen of acute respiratory infection in children. It is mainly transmitted through droplets, and can also be transmitted through contact with mucous membranes of eyes, mouth or nose. The incidence rate is highest in infants and young children. HPIV infection has a global distribution and is a common pathogen of community-acquired respiratory tract infection. The incidence of HPIV infection in children with acute respiratory tract infection is as high as 30-40%, which is the cause of severe lower respiratory tract infection in children, second only to respiratory syndrome. pathogens of cellular viruses. The pathogen was first discovered in the late 1950s, formerly...

Claims

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Application Information

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IPC IPC(8): G01N33/569G01N33/531G01N33/533
Inventor 胡征杨波董俊
Owner 湖北诺美华抗体药物技术有限公司
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