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Nanometer antibody capable of specifically recognizing enterobacter sakazakii and application of nanometer antibody

A technology of Enterobacter sakazakii and nano-antibody, which is applied in the field of molecular biology, can solve the problems of long production cycle, large number of animals, and high cost, and achieve the effects of reducing animal sacrifice, fast expression speed, and good stability

Active Publication Date: 2022-03-01
NORTHWEST A & F UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The current immunological detection methods for Enterobacter sakazakii are mainly based on polyclonal antibodies and monoclonal antibodies. Traditional antibody preparation techniques often require a large number of animals, with long preparation cycles, low yields, and high costs.
However, there have been no reports related to nanobodies against Enterobacter sakazakii

Method used

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  • Nanometer antibody capable of specifically recognizing enterobacter sakazakii and application of nanometer antibody
  • Nanometer antibody capable of specifically recognizing enterobacter sakazakii and application of nanometer antibody
  • Nanometer antibody capable of specifically recognizing enterobacter sakazakii and application of nanometer antibody

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Experimental program
Comparison scheme
Effect test

Embodiment

[0051] 1. Construction of anti-Enterobacter sakazakii phage display nanobody library

[0052] 1.1 Immunity of camels:

[0053] Select a healthy camel that has not been immunized with any antigen, and add 1 mL concentration of 10 8 CFU / mL of Enterobacter sakazakii inactivated whole bacteria, 0.1mL of 2mg / mL lipopolysaccharide, 0.1mL of 2mg / mL flagellin and 0.1mL of 2mg / mL sonicated whole protein were mixed evenly and mixed with 1.3mL of Fu Formula complete adjuvant was mixed, and the obtained emulsified mixture was used to immunize the Bactrian camel, and then the antigen was emulsified with Freund's incomplete adjuvant for the subsequent immunization, and the immunization was boosted once every two weeks, a total of 5 immunizations. One week after each immunization, the camel blood was collected to detect the antibody titer in the serum.

[0054] 1.2 Extraction of blood total RNA:

[0055] After the fifth immunization, the peripheral blood of the camel was collected, and th...

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Abstract

The invention provides a nano antibody capable of specifically recognizing enterobacter sakazakii and application of the nano antibody, and belongs to the field of molecular biology and the technical field of immunoassay. The nano antibody is provided with three complementary determining regions (CDR1, CDR2 and CDR3) which are shown as SEQ ID No.1 to SEQ ID No.3 in sequence. The nano antibody is high in stability, high in expression speed, high in yield, good in specificity and free of cross reaction with staphylococcus aureus; the detection sensitivity of a phage-mediated double nano antibody sandwich enzyme-linked immunosorbent assay method established by using the nano antibody is 5.08 * 10 < 4 > CFU / mL; the problems that an existing detection method is poor in specificity and high in cost can be solved, and a phage-mediated double-nano-antibody sandwich enzyme-linked immunosorbent assay method can be more widely applied.

Description

technical field [0001] The invention belongs to the field of molecular biology and the technical field of immune analysis, and specifically relates to a nanobody specifically recognizing Enterobacter sakazakii and an application thereof. Background technique [0002] Enterobacter sakazakii is an important food-borne pathogen. For infants and young children, it is easy to cause sepsis, meningitis, etc., and can lead to sequelae of the nervous system and even rapid death; for adults, infection with Enterobacter sakazakii It can cause sepsis, bacteremia, and local infection, and its infection can lead to high morbidity and mortality. According to statistics, the probability of death due to Enterobacter sakazakii infection is as high as 50%. Milk powder is the main channel for Enterobacter sakazakii infection, which mostly occurs in infants and young children. Due to the cumbersome production process of most foods such as milk powder, microbial contamination may occur in every ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/12C12N15/13C12N15/70C12N1/21C12Q1/689G01N33/569G01N33/68C12R1/19C12R1/01
CPCC07K16/1228C12N15/70C12Q1/689G01N33/56916G01N33/6854G01N2333/24C07K2317/565C07K2317/567C07K2317/569C07K2317/94Y02A50/30
Inventor 王妍入张瑶季艳伟郭兵王建龙
Owner NORTHWEST A & F UNIV
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