DOI: 10.1002/asia.201000242 In vivo Fluorescence Imaging of Tumors using Molecular Aptamers Generated by Cell-SELEX Hui Shi, [a] Zhiwen Tang, [b] Youngmi Kim, [b] Hailong Nie, [a] Yu Fen Huang, [b] Xiaoxiao He, [a] Ke Deng, [a] Kemin Wang,* [a] and Weihong Tan* [a, b] Introduction As a rapidly emerging field within biomedical research and clinical diagnosis, molecular imaging is critically important for in vivo studies characterizing the complex pathogenic ac- tivity that accompanies tumor growth. [1–4] Traditional imag- ing technologies mainly rely on morphological information. Molecular imaging, on the other hand, typically utilizes spe- cific molecular probes as the source of image contrast in order to distinguish particular biological events. Therefore, in the context of integrative biology, molecular imaging po- tentially offers clinicians the ability to detect disease at the early stages, as well as measure and characterize its patho- genesis, thus increasing the range of treatment options. [1] These are also the very functions which make molecular probes promising candidates for optimized in vivo imaging of molecular events inside the whole body. Up to now, a wide variety of targeting molecules have been tested, with varying degrees of success, for their poten- tial molecular imaging applications in clinical and laboratory settings. [5–8] These include small molecules, such as receptor ligands or enzyme substrates, and higher molecular-weight affinity ligands, such as monoclonal antibodies or recombi- nant proteins. However, a new and more promising means of targeting protein surfaces has emerged in the form of nu- cleic acid ligands, which are called aptamers. [9–10] Aptamers have now been recognized as a novel class of reliable affini- ty ligands that rival antibodies in their therapeutic and diag- nostic potential. Aptamers are RNA or DNA oligonucleo- tides that are able to fold by intramolecular interaction into Abstract: Poor sensitivity and low spe- cificity of current molecular imaging probes limit their application in clinical settings. To address these challenges, we used a process known as cell- SELEX to develop unique molecular probes termed aptamers with the high binding affinity, sensitivity, and specif- icity needed for in vivo molecular imaging inside living animals. Impor- tantly, aptamers can be selected by cell-SELEX to recognize target cells, or even surface membrane proteins, without requiring prior molecular sig- nature information. As a result, we are able to present the first report of ap- tamers molecularly engineered with signaling molecules and optimized for the fluorescence imaging of specific tumor cells inside a mouse. Using a Cy5-labeled aptamer TD05 (Cy5- TD05) as the probe, the in vivo efficacy of aptamer-based molecular imaging in Ramos (B-cell lymphoma) xenograft nude mice was tested. After intrave- nous injection of Cy5-TD05 into mice bearing grafted tumors, noninvasive, whole-body fluorescence imaging then allowed the spatial and temporal distri- bution to be directly monitored. Our results demonstrate that the aptamers could effectively recognize tumors with high sensitivity and specificity, thus es- tablishing the efficacy of these fluores- cent aptamers for diagnostic applica- tions and in vivo studies requiring real- time molecular imaging. Keywords: aptamers · cancer · cell recognition · fluorescence · imaging agents [a] Dr. H. Shi, Dr.H. Nie, Prof. X. He, K. Deng, Prof. K. Wang, Prof. W. Tan State Key Lab of Chemo/Biosensing and Chemometrics College of Biology College of Chemistry and Chemical Engineering Hunan University Changsha 410082 (China) E-mail : kmwang@hnu.edu.cn [b] Dr. Z. Tang, Dr. Y. Kim, Prof. Y.F. Huang, Prof. W. Tan Center for Research at the Bio/Nano Interface Department of Chemistry and Department of Physiology and Functional Genomics Shands Cancer Center Genetics Institute, and McKnight Brain Institute University of Florida Gainesville, Florida 32611-7200 (USA) Fax: (+ 1) 352-846-2410 E-mail : tan@chem.ufl.edu Chem. Asian J. 2010, 5, 2209 – 2213  2010 Wiley-VCH Verlag GmbH&Co. KGaA, Weinheim 2209