Date of Graduation

5-2011

Document Type

Dissertation

Degree Name

Doctor of Philosophy in Cell & Molecular Biology (PhD)

Degree Level

Graduate

Department

Biological Sciences

Advisor

Jin-Woo Kim

Committee Member

Douglas Rhoads

Second Committee Member

Russell Deaton

Third Committee Member

Steve Tung

Fourth Committee Member

Joshua Sakon

Abstract

Carbon nanotubes (CNT) have recently been in the limelight for its potential role in disease diagnostics and therapeutics. Even before these medical applications can be realized, there is a need to address issues like opsonization, phagocytosis by macrophages and sequestration to liver and spleen for eventual elimination from the body. We believe coating CNT with biocompatible and opsonin resistant moieties will not only help CNT in traversing the blood stream to reach the target organ, but also improve biodistribution tremendously. We set out to achieve this by firstly identifying a new compound, mAmp, which is a fluorescent derivative of the antibiotic, Ampicillin. Besides possessing a varied plethora of properties that complements CNTs already superlative traits, mAmp also affords opsonin resistance to CNT. We wanted to test and compare the four categories of opsonin repellants that we employed: synthetic - Polyethylene glycol (PEG), semisynthetic - mAmp, seminatural - Dextran sulfate (DSS) and natural - Protein A (PrA) + Factor H (FH). We developed novel strategies to conjugate these moieties to CNT and in the process also implemented attachment of Antibodies, specific recognition moieties, to these hybrids to make them ready for precision targeting downstream.

Of the four materials used, DSS posed considerable difficulties in achieving a pure DSS-CNT hybrid mainly due to its size and lack of defined purification strategies to separate polysaccharides in a mixture. We responded to this challenge by devising a simple lectin based affinity chromatography system that employs CNT as the support material. As proof of principle we tested the four hybrids on Staphylococcus aureus, in their ability to evade the bacterium in the absence of specific antibody and ability to specifically attach to the bacterium in its presence. We then tested the particles on human macrophages in the presence of opsonins, C3b and IgG. It was henceforth proved that coating CNT with the opsonin resistant moieties provided excellent immunity versus macrophages and considerable stealth character to CNTs.

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