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Sandwiched ELISA detection method for cryptosporidium parvum and applications thereof

A technology for Cryptosporidium microsporidium and a detection method, which is applied to measurement devices, instruments, scientific instruments and other directions, can solve the problems of low sensitivity, time-consuming and labor-intensive cryptosporidium, and achieve the effects of short test time, few consumables, and low cost.

Inactive Publication Date: 2017-05-31
HENAN AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] Aiming at the shortcomings of low sensitivity, time-consuming and labor-intensive detection of Cryptosporidium in feces, the present invention provides a sandwich ELISA detection method of Cryptosporidium parvum and its application

Method used

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  • Sandwiched ELISA detection method for cryptosporidium parvum and applications thereof
  • Sandwiched ELISA detection method for cryptosporidium parvum and applications thereof
  • Sandwiched ELISA detection method for cryptosporidium parvum and applications thereof

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Embodiment Construction

[0043] The coating solution is obtained by adding 1.59g of anhydrous sodium carbonate, 2.93g of sodium bicarbonate, and distilled water to 1000mL;

[0044] Washing solution, that is, PBST, consists of 8.0g sodium chloride, 0.2g potassium chloride, 0.2g potassium dihydrogen phosphate, Na 2 HPO 4 12H 2 O 2.9g, Tween-20 0.5mL, add distilled water 1000mL;

[0045] The blocking solution is obtained by adding 1g of bovine serum albumin and 100mL of coating solution;

[0046] The stop solution is obtained by adding 11.1mL of concentrated sulfuric acid to 88.9mL of distilled water;

[0047] PBS consists of NaCl8.0g, KCl0.2g, KH2 PO 4 12H 2 O 2.9g, obtained by adding distilled water to 1000mL;

[0048] TMB chromogenic solution:

[0049] Buffer A: Weigh 66.5063g of potassium citrate, dissolve it in 800mL of four-distilled water and adjust the pH value to 4.0 with concentrated hydrochloric acid, add 314μL of H 2 o 2 , dilute to 1L with water, store at 4°C;

[0050] Buffer B: We...

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Abstract

The invention discloses a sandwiched ELISA detection method for cryptosporidium parvum. The sandwiched ELISA detection method specifically comprises the following steps: (1) preparing a detective antigen and an immune antigen; (2) immunizing a healthy adult rabbit with the immune antigen, and preparing rabbit-anti-cryptosporidium parvum serum antibody IgG; (3) immunizing a healthy laying hen with the immune antigen, and preparing rabbit-anti-cryptosporidium parvum yolk antibody IgY; and (4) sandwiched ELISA detection steps by taking IgG as a capturing antibody and IgY as a detection antibody: enveloping IgG, blocking, adding antigen, adding IgY, adding enzyme-labelled antibody, adding chromophore-labeled substrate and stopping reaction. Compared with a nest type PCR method, the sandwiched ELISA detection method only needs a blocking buffer, a substrate solution, a PBS and the like, the prices of which are low, and only a microplate reader is needed in testing, so that the sandwiched ELISA detection method has the advantages of being fewer in consumable items, low in cost, simple and convenient to operate, short in test time, more applicable to detection of a great amount of clinical samples, and the like.

Description

technical field [0001] The invention belongs to the technical field of detection of Cryptosporidium parvum, and in particular relates to a sandwich ELISA detection method of Cryptosporidium parvum and an application thereof. Background technique [0002] Cryptosporidium (Cryptosporidium) is an important zoonotic intestinal protozoa, its host range is very wide, it can infect mammals including humans, birds, reptiles, amphibians and fish etc. Animals, mainly parasitic in the epithelial cells of the host's gastrointestinal tract. Cryptosporidium is mostly transmitted through contaminated water and food, and its infection can lead to zoonotic parasitic diseases with diarrhea as the main clinical symptom. At present, 31 species are considered to be effective, among them, Cryptosporidium parvum (C. Diarrhea in many vertebrates including humans. Immunocompetent hosts usually present with self-limited short-term acute diarrhea after infection with Cryptosporidium; while immunoco...

Claims

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Application Information

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IPC IPC(8): G01N33/569
CPCG01N33/56905
Inventor 菅复春张璐周昕薛张龙现宁长申王荣军
Owner HENAN AGRICULTURAL UNIVERSITY
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