Unlock instant, AI-driven research and patent intelligence for your innovation.

Nano antibody 3H12 for resisting pseudomonas aeruginosa exotoxin A

A nanobody, Pseudomonas aeruginosa technology, applied in anti-bacterial immunoglobulin, recombinant DNA technology, introduction of foreign genetic material using vectors, etc. The effect of avoiding phage contamination, less workload, and shorter screening cycle

Active Publication Date: 2021-06-22
深圳市国创纳米抗体技术有限公司
View PDF5 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the steps of RNA extraction and reverse transcription, and RNA is a substance that is easily contaminated and degraded, it is difficult to ensure a high enough screening library capacity, so that subsequent reverse transcription and PCR amplification are easily affected by template specificity. and the impact of quantity, resulting in failure of screening or extension of screening cycle

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Nano antibody 3H12 for resisting pseudomonas aeruginosa exotoxin A
  • Nano antibody 3H12 for resisting pseudomonas aeruginosa exotoxin A
  • Nano antibody 3H12 for resisting pseudomonas aeruginosa exotoxin A

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Example 1 Using Flow Cytometry to Screen Anti-Pseudomonas Exotoxin A Nanobodies

[0040] figure 1A preferred embodiment of the method disclosed in the present invention is given in figure 1 Firstly, B cells with unlabeled antibody were used for flow cytometry detection, and this was used as a negative control; at the same time, Anti-llama pan B cell monoclonalantibody was used as the primary antibody, Rat Anti-Mouse IgM-PE was used as the secondary antibody to label B cells, and FITC was used to label B cells Pseudomonas aeruginosa exotoxin A; establish the FSC-H / FSC-W experimental plate, and finally obtain PE and FITC double positive samples.

[0041] 1.1 Antigen-immunized alpaca: A healthy adult alpaca was selected, and the peripheral blood of the alpaca was collected as a negative control not immunized with antigen, and stored in a -80°C ultra-low temperature refrigerator. Mix the recombinant Pseudomonas aeruginosa exotoxin A as antigen with Freund's adjuvant at a ...

Embodiment 2

[0088] Example 2. Analysis of affinity activity between nanobody and antigen Pseudomonas aeruginosa exotoxin A

[0089] 2.1 Chip antigen coupling: Prepare the antigen with different pH sodium acetate buffer (pH 5.5, pH 5.0, pH 4.5, pH 4.0) to prepare 20 μg / mL working solution, and prepare 50 mM NaOH regeneration solution at the same time, use Biacore The template method in the instrument of the T100 protein interaction analysis system analyzes the electrostatic binding between antigens at different pH conditions and the surface of the chip (GE Company). Use a neutral pH system and adjust the antigen concentration as needed for coupling conditions. The chip was coupled according to the template method that comes with the instrument: select the blank coupling mode for channel 1, select the Target coupling mode for channel 2, and set the target to the designed theoretical coupling amount. The coupling process takes about 60 min.

[0090] 2.2 Exploration of analyte concentration...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a nano antibody 3H12 for resisting pseudomonas aeruginosa exotoxin A. The nano antibody 3H12 has a unique CDR region and has extremely high affinity with the pseudomonas aeruginosa exotoxin A. The invention further discloses a method for screening the nano antibody 3H12. The method is simple to operate, short in screening period, low in economic cost and suitable for industrial large-scale screening, the risk of bacteriophage pollution is avoided, the screening efficiency is improved, the workload is greatly reduced, and the industrial large-scale screening requirement is met.

Description

[0001] The patent application of the present invention is a divisional application of the Chinese invention patent application "a method for screening nanobodies by flow cytometry" with the application date of February 12, 2018 and the application number CN201810143471.6. technical field [0002] The invention discloses a nanobody and belongs to the technical field of polypeptides. Background technique [0003] In 1993, Hamers-Casterman et al. found that a class of only heavy chain dimer antibody H was found in camelids (camel, dromedary and llama). 2 , which are mainly IgG2 and IgG3 types. Such antibodies are called heavy chain-only-like antibodies (HCAbs) due to the lack of light chains, and their antigen-binding sites are composed of a domain called the VHH region, so Such antibodies are also referred to as single domain antibodies or single domain antibodies (sdAbs). Because this type of antibody is the variable region sequence after the constant region is removed, the...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/12C12N15/13C12N15/70
CPCC07K16/1214C07K2317/569C07K2317/565G01N15/1404
Inventor 宋海鹏刘原源于建立周宇杭李飞古一
Owner 深圳市国创纳米抗体技术有限公司