Aptamers are DNA or RNA oligonucleotide sequences that selectively bind to their target with high affinity and specificity. They are obtained using an iterative selection protocol called SELEX. Several small molecules and proteins have been used as targets. Recently, a variant of this methodology, known as cell-SELEX, has been developed for a new generation of aptamers, which are capable of recognizing whole living cells. We have used this methodology for the selection of aptamers, which show high affinity and specificity for several cancer cells. In this chapter, we describe (1) the process followed for the generation of aptamers capable of recognizing acute leukemia cells (CCRF–CEM cells) and (2) the method of enhancing the selectivity and sensitivity of these aptamers by conjugation with a dual-nanoparticle system, which combines magnetic nanoparticles (MNP) and fluorescent silica nanoparticles (FNP). Specifically, the selected aptamers, which showed dissociation constants in the nanomolar range, have been coupled to MNPs in order to selectively collect and enrich cells from complex matrices, including blood samples. The additional coupling of the aptamer to FNPs offers an excellent and highly sensitive method for detecting cancer cells. In order to prove the potential of this rapid and low-cost method for diagnostic purposes, confocal microscopy was used to confirm the specific collection and detection of target cells in concentrations as low as 250 cells. The final fluorescence of the cells labeled with the nanoparticles was quantified using a fluorescence microplate reader.
Fluorescent silica nanoparticle magnetic nanoparticle aptamer cell-SELEX cancer cells
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This work was supported by NIH, NCI, and NIGMS grants and by State of Florida Center for NanoBiosensors. M.-C.E. acknowledges financial support from the Departament d’Universitats, Recerca i Societat de la Informació de la Generalitat de Catalunya, Spain.
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1.Department of Chemistry and Department of Physiology and Functional Genomics Shands Cancer Center and UF Genetics Institute, and McKnight Brain Institute, Center for Research at the Bio/Nano InterfaceUniversity of FloridaGainesvilleUSA
2.Department of Chemistry and Department of Physiology and Functional Genomics Shands Cancer Center and UF Genetics Institute, and McKnight Brain Institute, Center for Research at the Bio/Nano InterfaceUniversity of FloridaGainesvilleUSA