Specific Recognition And Detection Of Lung Cancer Cells Based On Quantum Dots And Aptamers

Aptamers are short, single-stranded oligonucleotides generated from an invitro process known as SELEX (systematic evolution of ligands by exponentialenrichment). They can recognize a wide variety of targets with high affinity andselectivity. In addition, aptamers also possess some additional advantages, such astheir relatively small size, the ease of synthesis and modification, high stability,low immunogenicity and so on, widely used in bioanalysis, chemical biology, etc.Quantum dots (QDs) have been widely applied in various fields of moleculardiagnosis, target therapy, biomedical imaging and biosensor due to their superioroptical, chemical and electronic properties, such as high fluorescence quantumyield, multicolor and photochemical stability, allowing a single excitation source toexcite QDs of different emission colours, etc. In this thesis, taking non-small celllung cancer A549as an object, based on QDs and aptamers, in order to improve theefficiency of cell recognition, and develop simple, convenient detection methodsfor cancer cells. Several works have been mainly performed as following:1. Quantitative analysis of membrane receptor protein with fluorescentaptamer probesThe density of membrane receptor proteins binding with aptamers on A549cell were quantified according to the fluorescence decrease of aptamer probesbefore and after A549cell incubation. The results demonstrate that the number ofmembrane receptor proteins per A549cell is8.4×105on average, and the densityis ca.1050receptors/μm2, which permit larger QDs as markers of aptamers totarget A549cell. This work offers the possibility of QDs-based cell recognition.2. Specific recognition of non-small cell lung cancer A549based on QDs andaptamersQDs were introduced as a robust fluorescent labeling for cell recognition bytwo-step recognition method (recognition of aptamer before QDs labeling). It wascompared that the recognition effects of two aptamers S11e and S6at differenttemperatures. The incubation time and concentration of QDs were optimized. Theresults show that A549cell can be well recognized by S11e and S6at4℃and37℃, respectively. Incubation time of60min and QDs concentration at300nMguarantee a best recognition result. This work provides helpful information for further cell detection.3. Quantitative detection of A549cell based on QDs and aptamersThe recognition difference of two-step recognition method and one-steprecognition method (QDs-aptamer conjugates are to recognize cells) was studiedbased on QDs and aptamers. The results indicate that the former method avoidesthe influence of steric hindrance brought from QDs, guarantee in better efficiencyof cell recognition than that of the latter method. Meanwhile, the ratio of signal tobackgroud is three times higher than that of one-step recognition method. Inaddition, compared with traditional fluorescent dye, QDs show strong fluorescenceand high photostability. Based on the high specific recognition unit and strongsignal unit, sensitive detection of A549cell is realized, and the detection limit is6×10~3cells/mL. This work provides a sensitive and specific approach for cancer celldetection.