核酸适配子解剖.pdf

Aptamers in Bioanalysis Andrei Drabovich Bioanalytical Chemistry Samuel Lunenfeld Research Institute Lectures 2009 Mount Sinai Hospital / University of Toronto Affinity Probes Affinity probes – biopolymers or small molecules which bind to a target molecule with high affinity and specificity. These probes are used for quantitative analysis of targets which cannot be otherwise detected 1. The probe should be labelled with a detectable tag (fluorescent molecule, radioactive isotope) 2. The known excess of probe is mixed with the unknown amount of target 3. Probe-target complex and excess of the probe are separated and quantified Affinity Probes Contd. Types of affinity probes: 1. Antibodies – proteins used for the analysis of proteins. Produced in vivo by shuffling the parts of the genes as a response to antigene (target) introduction 2. Small molecules (for example, ethidium bromide for dsDNA analysis) 3. Oligonucleotides – DNA or RNA probes for quantitative hybridization analysis of DNA or RNA. Hybridization probes should be complimentary to their targets 4. Oligonucleotide aptamers – ssDNA or ssRNA used for the analysis of proteins and haptens. Selected in vitro from large libraries of random oligonucleotide sequences 5. Peptide aptamers – peptides used for the analysis of proteins and haptens. Selected in vitro from large libraries of random peptide sequences Oligonucleotide Aptamers The concept of aptamers (from apt: fitted, suited; Latin aptus: fastened) was introduced by Szostak’s and Gold’s groups in 1990. RNA aptamer to Malachite Green MG Malachite Green Secondary structure of Tertiary structure of the (MG) the aptamer with the a