title: Tracking and quantifying polymer therapeutic distribution on a cellular level using 3D dSTORM.
abstract: We used a single-molecule localization technique called direct stochastic optical reconstruction microscopy (dSTORM) to quantify both colocalization and spatial distribution on a cellular level for two conceptually different N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer conjugates. Microscopy images were acquired of entire cells with resolutions as high as 25nm revealing the nanoscale distribution of the fluorescently labeled therapeutic components. Drug-free macromolecular therapeutics consisting of two self-assembling nanoconjugates showed slight increase in nanoclusters on the cell surface with time. Additionally, dSTORM provided high resolution images of the nanoscale organization of the self-assembling conjugates at the interface between two cells. A conjugate designed for treating ovarian cancer showed that the model drug (Cy3) and polymer bound to Cy5 were colocalized at an early time point before the model drug was enzymatically cleaved from the polymer. Using spatial descriptive statistics it was found that the drug was randomly distributed after 24h while the polymer bound dye remained in clusters. Four different fluorescent dyes were used and two different therapeutic systems were tested to demonstrate the versatility and possible general applicability of dSTORM for use in studying drug delivery systems.
authors: Department of Bioengineering, University of Utah, 20 S. 2030 E., Rm. 108, Salt Lake City, UT 84112, USA.
authors: Jonathan M
authors: JM
authors: Hartley
authors: Department of Pharmaceutics and Pharmaceutical Chemistry, University of Utah, 20 S. 2030 E., Rm. 205, Salt Lake City, UT 84112, USA.
authors: Rui
authors: R
authors: Zhang
authors: Department of Biology, University of Utah, 257S 1400 E, Salt Lake City, UT 84112, USA; Bruker Nano Surfaces, 630 Komas Drive, Salt Lake City, UT 84108, USA.
authors: Manasa
authors: M
authors: Gudheti
authors: Department of Pharmaceutics and Pharmaceutical Chemistry, University of Utah, 20 S. 2030 E., Rm. 205, Salt Lake City, UT 84112, USA.
authors: Jiyuan
authors: J
authors: Yang
authors: Department of Bioengineering, University of Utah, 20 S. 2030 E., Rm. 108, Salt Lake City, UT 84112, USA; Department of Pharmaceutics and Pharmaceutical Chemistry, University of Utah, 20 S. 2030 E., Rm. 205, Salt Lake City, UT 84112, USA. Electronic address: jindrich.kopecek@utah.edu.
authors: Jindřich
authors: J
authors: Kopeček
mesh_terms: D002051:Burkitt Lymphoma / Q000378:metabolism; D045744:Cell Line, Tumor; D016503:Drug Delivery Systems; D005260:Female; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D008853:Microscopy / Q000379:methods; D058726:Nanoconjugates / Q000737:chemistry*; D061848:Optical Imaging; D010051:Ovarian Neoplasms / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D011109:Polymethacrylic Acids / Q000737:chemistry* / Q000378:metabolism
chemical_list: D005456:Fluorescent Dyes; D058726:Nanoconjugates; D010455:Peptides; D011109:Polymethacrylic Acids; C011533:Duxon
keywords: Drug delivery; N-(2-hydroxypropyl)methacrylamide (HPMA); Super resolution; dSTORM