Download presentation
Presentation is loading. Please wait.
Published bySherman Booth Modified over 8 years ago
1
Presented By O.A.NAZEER HUSSAIN Roll No: 43 S7 AE&I Guided By: Mr. Suresh Babu V
2
Overview Introduction Experiments Results Discussion Conclusion
3
Introduction Gene Transfection Deliberately introducing nucleic acids into cells. “Infection by transformation” Viral methods Carrier Viruses. ‘Virus Transduction’.
4
Non-Viral methods Chemical based transfection Non chemical methods Paticle based methods Merits Demerits
5
Magnetofection Invented by Christian Plank and Christian Bergemann Simplicity and efficiency. Application of magnetic field Processing time Principle
7
Schematic diagram of forming magnetic gene complexes via electrostatic interaction
8
Transfection agents Highly charged macromolecules Formation of Magnetic Gene vectors Composition. Toxicity. PEI(Poly ethyleneimine)
9
Hypothesis Any surface charged magnetic nanoparticle is capable of enhancing PEI transfection. Theory cationic polymers,cationic liposomes and genes Formation of complexes electrostatic adsorption Preparation of magnetic nanoparticles independent of genes or vectors used.
10
Testing Preparation of Magnetic nanoparticles Creation of charges Surface modifications (MP)-CA, (MP)-CMD, (MP)-APTES, (MP)-betaine etc 25kDA PEI,green fluroscent protein(GFP) plasmids, NIH-3T3 cells
11
EXPERIMENTS A. Preparation of Magnetic nanoparticles with various surface modifications Different modifications were done. 1. CA modification 2.CMD modification 3.APTES modification 4.Betaine modification 5.PEI modification
12
Structural formulas for coating materials used in this method
13
B. Transfection 25kDa PEI,GFP plasmids, NIH-3T3 cells mass ratio O.8 Different for each modification Serum culture medium Magneto FACTOR
14
Contd….. Replaced by medium supplemented with fetal calf serum. Flow cytometry All experiments performed in triplicatae
15
Results A. Materials
16
In the room temperature magnetization curve we can find that No coercive force. Specifice saturation Attraction towards bottom of the cell plate
18
The graph shows the zeta potential of magnetic nanoparticles with various surface modification. Zeta potential CA or CMD negatively charged APTES or betaine or PEI positively charged
19
TEM images of prepared magnetic nanoparticles with different modifications
20
Hydrodynamic diameters(Dh) of prepared magnetic nanoparticles with different modification
22
Transfection efficiencies of standard PEI transfection and magnetofection with various surface modifications. Only 25% cells express GFP after standard PEI transfection Magnetic field improves the efficiencies for all methods except fot betaine Magnetofection better than Standard PEI for MP-PEI and MP-CMD If magnetic nanoparticles are only added and the cell culture plate not placed on a magnetic plate, no enhancing effect is seen(black column)
23
Fluroscent photos of cell after magnetofection(using MP- PEI) or standard PEI transfection with same cell density Standard PEI transfection needs 4hr s of cell incubation
24
Discussions Results prove that surface-charged magentic nanoparticles enhance gene transfection efficiencies by applying a magnetic field CA and CMD have great biocompatibility, have equal or even better transfection efficiency than PEI They can be easily coated on magnetic particles since they have multicarboxyl groups. So they are more preferred than PEI MP-betaine suppress transfection efficiency. Also transfection efficiency is low if MP-PEI is alone used
25
Conclusion If the particle have sufficiently high surface potential, either positive or negative, they enhance gene delivery of PEI Particles’ surface charge is correlated with the efficiency of magnetofection Besides PEI, the carboxyl compounds CA and CMD are good coating agents
26
Refernces F. Scherer,M. Anton, U. Schillinger, J. Henkel, C. Bergemann, A. Kruger,B. Gansbacher, and C. Plank, “Magnetofection: Enhancing and targeting gene delivery by magnetic force in vitro and in vivo,” Gene Ther.,vol.9,no. 2, pp. 102–109, 2002. S. C.McBain, H. H. P. Yiu, A. E. Haj, and J. Dobson, “Polyethyleneimine functionalized iron oxide nanoparticles as agents for DNA delivery and transfection,” J. Mater. Chem., vol. 17, no. 24, pp. 2561–2565, 2007 X. Pan, J. Guan, J.W. Yoo, A. J. Epstein, L. J. Lee, and R. J. Lee, “Cationic lipid-coated magnetic nanoparticles associated with transferrin for gene delivery,” Int. J. Pharma., vol. 358, no. 1–2, pp. 263–270, 2008.
27
THANK YOU
28
QUESTIONS???????
Similar presentations
© 2025 SlidePlayer.com. Inc.
All rights reserved.