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Y. Dong et al.
13.1 Introduction
As systematically introduced in the aforementioned chapters, aptamers have been
widely utilized in molecular imaging and point-of-care testing, drug development,
targeted delivery, personalized medicine, customized therapy, medical appliances,
and a broad-spectrum of fundamental medical/biomedical researches, reflecting
the great potential of in vitro diagnostics, prognostics, and theranostics for public
health. However, well-performed aptamers are still limited. In order to facilitate
effectively aptamer screening, characterizing, and further expand the application
scope of aptamers, some frontier technologies, as exemplified by UVMag-SELEX,
aptamer biochip, and aptamer-oriented native mass spectrometry, emerge as the most
intriguing investigations nowadays.
13.2 UVMag-SELEX
Usually, aptamers are generated by Systematic Evolution of Ligands by Exponential Enrichment (SELEX), an in vitro selection procedure [1]. An oligonucleotide
library containing 10
13 –10
15 single-stranded sequences is chemically synthesized for
the beginning, and each sequence may contain 20–60 nt oligonucleotides in random
region flanked by short constant regions. The selection normally experiences 6–20
selection rounds of target incubation, unbound separation, bound elution, and amplification. Oligonucleotides that specifically bind to the target molecule are further
enriched to identify aptamer candidates with affinity characterization of the dissociation constants (Kd values). Generally, Kd values of aptamers are rather low at
micromolar or nanomolar levels, which ensures the high affinity of aptamer to its
corresponding target practically. Different SELEX variants [2, 3], e.g., Cell-SELEX
[4, 5], fluorescence-magnetic beads (FluMag-SELEX) [6], capillary electrophoresis
(CE-SELEX) [7], microfluidic-SELEX [8], in silico-SELEX [9], were successively
developed in the past few decades. Most of the reported methods need labeling treatment to the oligonucleotides so that aptamer candidates with high affinity toward
the target can be identified throughout the selection cycles; however, the labeling
process might negatively affect aptamer’s specific adaptive folding or require expensive equipment. In addition, most aforecited SELEX variants are applicable to find
aptamers toward a single target molecule, for aptamers toward broad-spectrum or
class-specific target molecules such as tetracycline or sulfonamide antibiotics, and
these SELEX variants will not work exclusively.
UVMag-SELEX was developed in recent years by the author’s research group
accordingly. DNA quantification by wide applicable UV colorimetry and process
monitoring of SELEX can simply be carried out by a miniaturized UV spectrometer. No labeling was needed, and the DNA monitoring was fulfilled with ease using
label-free UV spectrometry, obviating the utilization of gel electrophoresis, and thus
shortening selection time and enhancing SELEX efficiency. Hence, UVMag-SELEX
Y. Dong et al.
13.1 Introduction
As systematically introduced in the aforementioned chapters, aptamers have been
widely utilized in molecular imaging and point-of-care testing, drug development,
targeted delivery, personalized medicine, customized therapy, medical appliances,
and a broad-spectrum of fundamental medical/biomedical researches, reflecting
the great potential of in vitro diagnostics, prognostics, and theranostics for public
health. However, well-performed aptamers are still limited. In order to facilitate
effectively aptamer screening, characterizing, and further expand the application
scope of aptamers, some frontier technologies, as exemplified by UVMag-SELEX,
aptamer biochip, and aptamer-oriented native mass spectrometry, emerge as the most
intriguing investigations nowadays.
13.2 UVMag-SELEX
Usually, aptamers are generated by Systematic Evolution of Ligands by Exponential Enrichment (SELEX), an in vitro selection procedure [1]. An oligonucleotide
library containing 10
13 –10
15 single-stranded sequences is chemically synthesized for
the beginning, and each sequence may contain 20–60 nt oligonucleotides in random
region flanked by short constant regions. The selection normally experiences 6–20
selection rounds of target incubation, unbound separation, bound elution, and amplification. Oligonucleotides that specifically bind to the target molecule are further
enriched to identify aptamer candidates with affinity characterization of the dissociation constants (Kd values). Generally, Kd values of aptamers are rather low at
micromolar or nanomolar levels, which ensures the high affinity of aptamer to its
corresponding target practically. Different SELEX variants [2, 3], e.g., Cell-SELEX
[4, 5], fluorescence-magnetic beads (FluMag-SELEX) [6], capillary electrophoresis
(CE-SELEX) [7], microfluidic-SELEX [8], in silico-SELEX [9], were successively
developed in the past few decades. Most of the reported methods need labeling treatment to the oligonucleotides so that aptamer candidates with high affinity toward
the target can be identified throughout the selection cycles; however, the labeling
process might negatively affect aptamer’s specific adaptive folding or require expensive equipment. In addition, most aforecited SELEX variants are applicable to find
aptamers toward a single target molecule, for aptamers toward broad-spectrum or
class-specific target molecules such as tetracycline or sulfonamide antibiotics, and
these SELEX variants will not work exclusively.
UVMag-SELEX was developed in recent years by the author’s research group
accordingly. DNA quantification by wide applicable UV colorimetry and process
monitoring of SELEX can simply be carried out by a miniaturized UV spectrometer. No labeling was needed, and the DNA monitoring was fulfilled with ease using
label-free UV spectrometry, obviating the utilization of gel electrophoresis, and thus
shortening selection time and enhancing SELEX efficiency. Hence, UVMag-SELEX