Details
| Original language | English |
|---|---|
| Pages (from-to) | 199-205 |
| Number of pages | 7 |
| Journal | TALANTA |
| Volume | 172 |
| Publication status | Published - 18 May 2017 |
Abstract
Sensitive and specific detection and quantification of small molecules often remain challenging. We developed a novel magnetic bead-based aptamer-assisted real-time PCR (Apta-qPCR) assay to provide a versatile platform for quantification of small molecules. The assay has been realized for the detection of ATP as a model system. The assay relies on a combination of qPCR with the target-induced dissociation (TID) of ATP aptamer from an oligonucleotide, complementary to the ATP binding site of the aptamer. The complementary oligonucleotide was immobilized on deoxythymidine (dT)-modified magnetic beads (dT-beads) and hybridized with the aptamer. The presence of ATP resulted in dissociation of the aptamer from the dT-beads and the dissociated aptamer was quantified using qPCR. The Apta-qPCR assay was able to detect 17 nM ATP with a broad dynamic range from 50 nM to 5 mM. The assay is label-free, and real-time PCR-based detection of aptamer facilitates high sensitivity. The presented method is highly versatile and can be applied to various aptamer-target pairs to allow detection of a broad range of target analytes.
Keywords
- Apta-qPCR, Aptamer, ATP, Small molecules, Target-induced dissociation
ASJC Scopus subject areas
- Chemistry(all)
- Analytical Chemistry
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In: TALANTA, Vol. 172, 18.05.2017, p. 199-205.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Aptamer-based detection of adenosine triphosphate via qPCR
AU - Modh, Harshvardhan
AU - Witt, Martin
AU - Urmann, Katharina
AU - Lavrentieva, Antonina
AU - Segal, Ester
AU - Scheper, Thomas
AU - Walter, Johanna Gabriela
N1 - Funding information: The German Research Foundation (DFG - SCHE 279/32-1) supported parts of this work. German Academic Exchange Service (DAAD) is acknowledged for the financial support to Harshvardhan Modh. We thank Didem Ag Seleci (Institute of Technical Chemistry, Leibniz University of Hannover) for providing HeLa cells.
PY - 2017/5/18
Y1 - 2017/5/18
N2 - Sensitive and specific detection and quantification of small molecules often remain challenging. We developed a novel magnetic bead-based aptamer-assisted real-time PCR (Apta-qPCR) assay to provide a versatile platform for quantification of small molecules. The assay has been realized for the detection of ATP as a model system. The assay relies on a combination of qPCR with the target-induced dissociation (TID) of ATP aptamer from an oligonucleotide, complementary to the ATP binding site of the aptamer. The complementary oligonucleotide was immobilized on deoxythymidine (dT)-modified magnetic beads (dT-beads) and hybridized with the aptamer. The presence of ATP resulted in dissociation of the aptamer from the dT-beads and the dissociated aptamer was quantified using qPCR. The Apta-qPCR assay was able to detect 17 nM ATP with a broad dynamic range from 50 nM to 5 mM. The assay is label-free, and real-time PCR-based detection of aptamer facilitates high sensitivity. The presented method is highly versatile and can be applied to various aptamer-target pairs to allow detection of a broad range of target analytes.
AB - Sensitive and specific detection and quantification of small molecules often remain challenging. We developed a novel magnetic bead-based aptamer-assisted real-time PCR (Apta-qPCR) assay to provide a versatile platform for quantification of small molecules. The assay has been realized for the detection of ATP as a model system. The assay relies on a combination of qPCR with the target-induced dissociation (TID) of ATP aptamer from an oligonucleotide, complementary to the ATP binding site of the aptamer. The complementary oligonucleotide was immobilized on deoxythymidine (dT)-modified magnetic beads (dT-beads) and hybridized with the aptamer. The presence of ATP resulted in dissociation of the aptamer from the dT-beads and the dissociated aptamer was quantified using qPCR. The Apta-qPCR assay was able to detect 17 nM ATP with a broad dynamic range from 50 nM to 5 mM. The assay is label-free, and real-time PCR-based detection of aptamer facilitates high sensitivity. The presented method is highly versatile and can be applied to various aptamer-target pairs to allow detection of a broad range of target analytes.
KW - Apta-qPCR
KW - Aptamer
KW - ATP
KW - Small molecules
KW - Target-induced dissociation
UR - http://www.scopus.com/inward/record.url?scp=85019592669&partnerID=8YFLogxK
U2 - 10.1016/j.talanta.2017.05.037
DO - 10.1016/j.talanta.2017.05.037
M3 - Article
C2 - 28602295
AN - SCOPUS:85019592669
VL - 172
SP - 199
EP - 205
JO - TALANTA
JF - TALANTA
SN - 0039-9140
ER -