University PhD Dissertation Defense
New AFM Methods for Characterizing Drug Delivery Agents
Elizabeth A. Hager-Barnard
Department of Materials Science & Engineering
Advisor: Prof. Nicholas A. Melosh
Monday July 25th 2011
1:00 pm
(Refreshments at 12:45 pm)
Location: Paul G. Allen Auditorium (CIS-X 101)
http://cis.stanford.edu/directions/
Abstract:
Understanding how cell-penetrating peptides (CPPs) can pass through cell membranes is critical for designing drug delivery agents. While CPPs like HIV-TAT have been widely studied, their ability to penetrate membranes directly, without active transport, is still a matter of considerable debate. New direct contact methods that report the interactions during translocation are needed to end this debate.
Here we show that atomic force microscopy (AFM) is an excellent direct contact method for characterizing CPPs. By pushing CPP-functionalized AFM probes through lipid bilayers we directly measure CPP-lipid interactions during the actual membrane translocation event. Furthermore with our custom AFM probes it is possible to track the CPP location inside the bilayer over time. We exploit this ability in a new method, Penetration Pathway Analysis (PPA), that creates probability maps that illustrate the paths CPP take as they penetrate through lipid bilayers. In addition to tracking CPP position, AFM results can also characterize the mechanics and energetics of bilayer penetration through a method called dynamic force spectroscopy (DFS).
Our results show that PPA combined with DFS is an excellent method for studying drug delivery agents like CPPs. For example, PPA combined with DFS can differentiate between CPPs that can directly penetrate bilayers and those that cannot. By using this new method it will be possible to directly characterize, with microsecond resolution, the dynamics and energetics of the interactions between lipid bilayers and a wide range of peptides and molecular species.
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