Fluorogenic Quantum Dot-Gold Nanoparticle Assembly for Beta Secretase Inhibitor Screening in Live Cell

Choi, Yong Je; Cho, Yoo Jin; Kim, Minjung; Grailhe, Régis; Song, Rita

doi:10.1021/ac301574b

Cited by 55 publications

(57 citation statements)

References 41 publications

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“…6AuNP, rather than smaller AuNPs, was chosen as the acceptor to provide for improved analytical performance. 8 In these experiments the 6AuNP concentration was kept constant at 50 nM, with the amount of QDs varied to obtain the specified AuNP to QD ratios. It is important to note that in configuration 1 it was the QDs that were kept at a constant concentration, with the concentration of AuNP varied to obtain the desired ratios.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

Energy Transfer Assays Using Quantum Dot–Gold Nanoparticle Complexes: Optimizing Oligonucleotide Assay Configuration Using Monovalently Conjugated Quantum Dots

Uddayasankar

Krull

2015

Langmuir

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The energy transfer between quantum dots (QDs) and gold nanoparticles (AuNPs) represents a popular transduction scheme in analytical assays that use nanomaterials. The impact of the spatial arrangement of the two types of nanoparticles on analytical performance has now been evaluated using a nucleic acid strand displacement assay. The first spatial arrangement (configuration 1) involved the assembly of a number of monovalently functionalized QD-oligonucleotide conjugates around a single central AuNP that was functionalized with complementary oligonucleotide sequences. The assembly of these complexes, and subsequent disassembly via target oligonucleotide-mediated displacement, were used to evaluate energy transfer efficiencies. Furthermore, the inner filter effect of AuNPs on the fluorescence intensity of the QD was studied. AuNPs of three different diameters (6, 13, and 30 nm) were used in these studies. Configuration 2 was based on the placement of monovalently functionalized AuNP-oligonucleotide conjugates around a single QD that was functionalized with a complementary oligonucleotide. The optimal assay configuration, established by evaluating energy transfer efficiencies and inner filter effects, was obtained by arranging at most 15 QDs around the 13 nm AuNP (configuration 1). These assays provided a 2.5-fold change in fluorescence intensity in the presence of target oligonucleotides. To obtain the same response with configuration 2 required the placement of three 6 nm AuNPs around the QD. This resulted in configuration 2 having a 5-fold lower fluorescence intensity when compared to configuration 1. The use of low-cost detection systems (digital camera) further emphasized the higher analytical performance of configuration 1. Response curves obtained using these detection systems demonstrated that configuration 1 had a 10-fold higher sensitivity when compared to configuration 2. This study provides an important framework for the development of sensitive assays using gold nanoparticles and quantum dots.

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Section: ■ Results and Discussionmentioning

confidence: 99%

Energy Transfer Assays Using Quantum Dot–Gold Nanoparticle Complexes: Optimizing Oligonucleotide Assay Configuration Using Monovalently Conjugated Quantum Dots

Uddayasankar

Krull

2015

Langmuir

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“…This restored the QD PL and allowed the measurement of enzyme activity in vitro and in living cells. 300 Wei et al presented a DNA-templated heterobivalent QD FRET nanosensor. This probe exhibited dual motifs for extracellular and intracellular targeting and imaging of cancer cells.…”

Section: Cell Environmentmentioning

confidence: 99%

Quantum dots: bright and versatile in vitro and in vivo fluorescence imaging biosensors

2015

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Semiconductor quantum dots (QDs) have become important fluorescent probes for in vitro and in vivo bioimaging research. Their nanoparticle surfaces for versatile bioconjugation, their adaptable photophysical properties for multiplexed detection, and their superior stability for longer investigation times are the main advantages of QDs compared to other fluorescence imaging agents. Here, we review the recent literature dealing with the design and application of QD-bioconjugates for advanced in vitro and in vivo imaging. After a short summary of QD preparation and their most important properties, different QD-based imaging applications will be discussed from the technological and the biological point of view, ranging from super-resolution microscopy and single-particle tracking over in vitro cell and tissue imaging to in vivo investigations. A substantial part of the review will focus on multifunctional applications, in which the QD fluorescence is combined with drug or gene delivery towards theranostic approaches or with complementary technologies for multimodal imaging. We also briefly discuss QD toxicity issues and give a short outlook on future directions of QD-based bioimaging.

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“…24 Using a peptide substrate sandwiched by CdSe/ZnS quantum dots (QDs) and gold nanoparticles, Choi et al recently extended FRET to BACE1 inhibitor screening in living cells. 30 However, traditional FRET assays have relatively small stokes shifts and the near-UV wavelengths for fluorophore excitation could overlap with the absorption peaks of many small molecules. 24 As a consequence, the choices of the donor/acceptor pairs in many applications is limited and varies from study to study.…”

Section: Introductionmentioning

confidence: 99%

“…Such steric hindrance might be the reason why relatively long reaction times (≥ 3 h) are needed for FRET-based BACE1 inhibition assays. 24,30 …”

Section: Introductionmentioning

confidence: 99%

Sensitive and Continuous Screening of Inhibitors of β-Site Amyloid Precursor Protein Cleaving Enzyme 1 (BACE1) at Single SPR Chips

Hao

Xia

et al. 2013

Anal. Chem.

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Developments of new methods that meet the demand of high-throughput, high-fidelity screening of hit compounds are important to searching modalities of important diseases such as neurological disorders, HIV, and cancer. A surface plasmon resonance (SPR)-based method capable of continuously screening enzyme inhibitors at a single chip with antibody-amplified signal enhancement is developed. The proof of concept is demonstrated by monitoring the cleavage of chip-confined peptide substrates (a segment of the amyloid precursor protein (APP) with the Swiss mutation) by β-site APP cleaving enzyme 1 (BACE1). In the presence of a non- inhibitor, BACE1 clips the peptide substrate at the cleavage site, detaching a fragment that is homologous to the N-terminus of the amyloid beta (Aβ) peptide. Consequently, a subsequent injection of the Aβ antibody does not lead to any molecular recognition or SPR signal change at the chip. In contrast, abolishment of the BACE1 activity by a strong inhibitor leaves the peptide substrate intact, and the subsequent antibody attachment produces an easily detectable SPR signal. Compared to the widely used FRET (fluorescence resonance energy transfer) assay, the method reported here is more cost effective, as unlabeled peptide is used as the BACE1 substrate. Furthermore, the assay is more rapid (each screening cycle lasts for ca. 1.5 h) and can be continuously carried out at a single, regenerable SPR chip for more than 30 h. Consequently, excellent reproducibility (RSD% < 5%) and throughput can be attained. Two inhibitors were screened and their half maximal inhibitory concentrations (IC50) determined by the SPR method are in excellent agreements with values deduced from ELISA and mass spectrometry.

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Fluorogenic Quantum Dot-Gold Nanoparticle Assembly for Beta Secretase Inhibitor Screening in Live Cell

Cited by 55 publications

References 41 publications

Energy Transfer Assays Using Quantum Dot–Gold Nanoparticle Complexes: Optimizing Oligonucleotide Assay Configuration Using Monovalently Conjugated Quantum Dots

Energy Transfer Assays Using Quantum Dot–Gold Nanoparticle Complexes: Optimizing Oligonucleotide Assay Configuration Using Monovalently Conjugated Quantum Dots

Quantum dots: bright and versatile in vitro and in vivo fluorescence imaging biosensors

Sensitive and Continuous Screening of Inhibitors of β-Site Amyloid Precursor Protein Cleaving Enzyme 1 (BACE1) at Single SPR Chips

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