Preparation Of Micro-scale Magnetic Beads Based On Macromolecular Crosslinking Of Nanoparticles And Their Application In The Seperation Of Biological Samples

In this paper,firstly,the magnetic Fe₃O₄ nanoparticles were prepared and functionalized on their surfaces;Afterwards,the micro-scale magnetic beads were also prepared by macromolecular crosslinking of the as-prepared nanoparticles,and then they were used to isolate and purify target DNA.Aqueous magnetite nanoparticles with rich hydroxyl groups were prepared by coprecipitation method.Fe₃O₄ magnetic nanoparticles were nearly spherical with mean diameter of ca.15nm,they were not very stable under ambient conditions and tended to agglomerating.Based on this,Fe₃O₄ magnetic nanoparticles were modified with polyethyleneimine（PEI）by in-situ method and silane coupling agent 3-（2,3-epoxypropyl oxygen）propyl silane three oxygen radicals（KH560）by post-processing method respectively.With rich amino and epoxy groups on the surface,the dispersion of magnetic nanoparticles had been improved.Three different kinds of magnetic beads（Fe₃O₄-KH560-PEI、Fe₃O₄-PEI-BDE-PEI、Fe₃O₄-PEI-PAA）were synthesized with polyethyleneimine（PEI）and polyacrylic acid（PAA）by crosslinking of functionalized magnetic nanoparticles respectively.Particle size,potential,morphology and magnetic responsiveness,suspension stability and other properties were characterized and compared,comprehensive conclusion was Fe₃O₄-KH560-PEI magnetic beads possessed optimal performance.The responsiveness of particles were significantly improved with the remanent magnetization rate falling to 12.24%and 5.86%after magnetic separation for 1min to 2min respectively.Besides,due to rich amino on the surface,the particles had good dispersion stability with 97.18%of particles suspended in the solution after 30 min.Compared with medical magnetic beads,the magnetic beads had smaller diameter with irregular shape and uneven size,leading to slightly lower magnetic responsiveness than medical magnetic beads.However,smaller particle size can effectively reduce the reaction of the space steric hindrance,improving the sensitivity of response.In addition,with rich amino on the surface,the particles tended to connect with other active substances and enlarged its application.The magnetic beads（Fe₃O₄-KH560-PEI）were used as the carrier,herring sperm DNA as the model DNA to examine the performance for DNA separation.The magnetic beads adsorped DNA under low pH and desorped under high pH to realize the separation and purification of DNA by electrostatic interaction between magnetic beads and DNA.Based on adsorption,desorption influencing factors（such as pH,time,temperature,ionic strength,etc.）,the optimum conditions of adsorption and desorption was pH=3,37℃,no salt,10min and pH=11,37℃,no salt,reaction 10 min respectively.The experimental results showed that DNA could be fast separated using the magnetic beads by magnetic field,the maximum capacity of DNA could be up to 629.49 mg/g.Moreover,Magnetic beads could be recycled and their adsorption rate remained above 50%after six times,which expected to highlight its advantages in practical application.