Novel quick separation method of bacilluscereus

Abstract

The invention discloses a novel quick separation method of bacilluscereus, which provides the basis for the subsequent research for a target bacterium and relates to the technical field of biology. The novel quick separation method comprises the steps of: carrying out covalent coupling on a multi-arm star-shaped polymer and an antibody, coating the antibody-modified multi-arc star-shaped polymer on a long-chain biotin molecule, capturing the target bacterium in a sample solution by the antibody and long-chain biotin co-modified multi-arc star-shaped polymer, recognizing and coupling the long-chain biotin multi-arm star-shaped polymer in the sample solution by a streptavidin-modified nano magnetic bead, separating and heavily-suspending the captured bacterium; a heavily-suspended liquid can be directly analyzed in subsequent. Compared with a conventional bacterium magnetic separation method, the novel quick separation method is more suitable for carrying out magnetic separation on the bacterium in a complex matrix; the separation efficiency of the target bacterium in a sample is increased.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for isolating food-borne pathogenic bacteria based on nano magnetic beads. Background technique [0002] Foodborne pathogen contamination is one of the major problems of food safety in my country. According to WHO statistics, about one-third of people in developed countries are infected with food-borne diseases every year, and 2.2 million people in the world die every year due to food-borne diseases. In my country, the number of cases of food poisoning is between 200,000 and 400,000 per year, most of which are caused by food-borne pathogens except for accidents. Bacillus cereus ( Bacillus cereus ), as one of the common food-borne pathogens, mainly contaminates meat, milk, vegetables, fish, rice and pasta, etc., and has been recognized by Norway and the Netherlands as the most easily detected pathogenic microorganisms in food. Poisoning incidents occur from time to time, e...

AI Technical Summary

Problems solved by technology

However, the current separation technology based on micron-scale immunomagnetic beads has many limitations: 1) The specific surface area of ​​micron-sized magnetic beads is relatively small, which reduces the capture efficiency of magnetic beads; Bacterial cells are combined through a multiphase reaction (multiphase reaction), and it usually takes longer to specifically capture bacterial cells in the food matrix; 3) Micron magnetic beads have poor monodispersity and are prone to self-disruption in the food matrix solution. Aggregation or formation of precipitates; 4) Traditional immunomagnetic separation techniques often directly couple antibody molecules to immunomagnetic beads, which often leads to greatly reduced antibody activity and changes in the spatial direction of antibodies. The steric hindrance effect between antibodies reduces the capture efficiency of antibodies. 5) The nature of the food matrix is ​​complex and the concentration of non-target pathogenic bacteria is large. Micron magnetic beads are prone to non-specific adsorption, and it is difficult to realize the detection of food samples. Specific separation of target bacteria; 6) Too high concentration of micron magnetic beads will cause damage to bacterial cells (magnetic field causes magnetic beads on the cell surface to attract each other, causing cells to be squeezed or even ruptured), resulting in failure of separation; 7) Magnetic beads When coupling antibodies, hydrophobic adsorption or chemical coupling is generally used to couple active antibodies on the surface of magnetic beads

The distance between the antibody and the surface of the magnetic bead is too close, the nature of the magnetic bead itself and the residual hydrophobic or strong hydrophilic groups on the surface are likely to cause changes in the spatial conformation of the antibody, resulting in a decrease in the biological activity of the antibody

Images