Conjugated Polymer Nanoparticles for Two-Photon Imaging of Endothelial Cells in a Tissue Model
✍ Scribed by Nur Aida Abdul Rahim; William McDaniel; Kevin Bardon; Shalini Srinivasan; Vernella Vickerman; Peter T. C. So; Joong Ho Moon
- Publisher
- John Wiley and Sons
- Year
- 2009
- Tongue
- English
- Weight
- 222 KB
- Volume
- 21
- Category
- Article
- ISSN
- 0935-9648
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✦ Synopsis
Two-photon (2P) imaging holds great promise for in vivo microscopic physiological studies in areas such as neurobiology, immunology, and tissue engineering. [1][2][3] Advances in 2P endomicroscope design have further demonstrated the possibility of developing noninvasive diagnostic procedures for the detection of malignancy in organs such as the oral cavity and the intestine. [1,4,5] Many of these applications are currently limited by a lack of fluorescent probes with large 2P action cross-sections. The availability of improved 2P fluorescent probes will enable tissue imaging with higher sensitivity, greater penetration depth, reduced tissue photodamage and tissue autofluorescence backgrounds, and lower instrument cost. Despite a number of previous efforts to design and synthesize new organic fluorophores with high 2P action cross-sections, [6][7][8][9][10] the utility of these probes in biomedicine has been limited by their hydrophobicity and cytotoxicity. A recent development is the realization that quantum dots (QDs) have large 2P action cross-sections on the order of 10 000 Goeppert-Mayer units (GM). [11] These QDs exhibit high photostability and narrow emission spectral width. Despite their excellent photophysical properties, broad biomedical applications of QDs are limited by drawbacks such as the existence of ''dark'' dots, [12] heavy-metal core-related cytotoxicity, and difficulties in surface modification. [13] Therefore, the synthesis and fabrication of alternative probes with high 2P action cross-sections is a priority.
Conjugated polymers (CPs) are attractive candidate materials that address the requirements for 2P microscopy imaging. CPs are organic materials that are readily synthesized by well-established methods. [14] Characteristic photophysical features of CPs are high fluorescence quantum yields (QYs), large extinction coefficients, and efficient energy-transfer properties. [15,16] Despite these characteristics, their intrinsic hydrophobicity originating from the p-conjugated backbone limits their potential application in biological systems. To overcome this hydrophobicity issue in cellular-imaging applications, we previously demonstrated the facile fabrication of conjugated polymer nanoparticles (CPNs) using ultrafiltration techniques. [17] Ultrafiltration of acetic acid-treated poly( p-phenylene ethynylene) (PPE, Scheme 1) followed by dialysis against pure water results in stable and bright CPNs 80-100 nm in size. Live cells readily uptake CPNs, which accumulate in the cytosol with no noticeable inhibition of cellular viability, even after 7 days of incubation.
In this Communication, we report on CPNs' superior 2P characteristics and 2P imaging of endothelial cells in a model-tissue culture system. Improved fabrication methods allowed dramatic reduction of CPN size, down to 8 nm. CPNs exhibit extremely large 2P cross-sections and photostability comparable to QDs. Furthermore, the hydrophilicity and nontoxicity of CPNs allow for long-term monitoring of angiogenesis by endothelial cells in a tissue model, supporting CPNs' potential in biological and biomedical applications.
CPNs were fabricated by ultrafiltration of an amine-containing PPE solution that was acidified with tartaric acid (1 M). [17] After dialysis of the solution against water, transparent yellow colored (l max, absorption ¼ 429 nm, l max, emission ¼ 485 nm, and QY: 0.09) CPNs were obtained (see Supporting Information for spectroscopic data, Fig. S1). The Z-average size of CPNs obtained from this tartaric acid treatment was significantly smaller (8 nm) than that obtained using the acetic acid treatment of the same batch of PPE solution (85 nm, see Supporting Information, Fig. S2), indicating that CPN size can be modulated by the choice of COMMUNICATION www.advmat.de