Quantum Dots as Förster Resonance Energy Transfer Acceptors of Lanthanides in Time-Resolved Bioassays

Abstract

We report a flexible and modular design for biosensors based on exploiting semiconductor quantum dots (QDs) and their excellent Forster resonance energy transfer (FRET) acceptor properties along with the long-lived fluorescent lifetimes of lanthanide donors. We demonstrate the format's wide application by developing a broad adenosine diphosphate (ADP) sensor with quantitative and highthroughput capabilities as a kinase/ATPase assay method. The sensor is based on a Terbium (Tb)-labeled antibody (Ab) that selectively recognizes ADP versus ATP. The Tb-labeled Ab (Ab-Tb) acts as a FRET donor to a QD, which has an ADP modified His 6 -peptide conjugated to its surface via metal-affinity coordination. This strategy of using self-assembly, modified peptides to present antibody epitopes on QD surfaces is readily transferable to other assays of interest. We utilize time-resolved FRET (TR-FRET) to measure the amounts of Ab-Tb bound to the QD by looking at the emission ratio of the QD and Tb in a time-gated manner, minimizing background signal. With the addition of free ADP the antibody is competitively separated from the QD and a change in the ratiometric emission signal correlates with the free ADP concentration. The sensor obtained a detection limit below 10 nM of free ADP and quantitation limit of 35 nM ADP using 8 nM of sensor. Quantitative values were obtained for a model enzyme (glucokinase) kinetics, as well as demonstrations of the assays capability to distinguish enzyme inhibitors. We discuss future outlooks and note areas for improvement in similar design strategies.

Sensing with photoluminescent semiconductor quantum dots

“…Reprinted with permission from Ref. [21] Copyright 2018 American Chemical Society. (a) Schematic of the donor QD labeled with three different peptide dye acceptors.…”

“…Increasing concentrations of GLK favored faster reaction rates, and kinetic parameters could be determined from their experiments yielding specificity constant of 210 ± 100 Mm −1 s −1 , k cat app = 19 ± 6 s −1 and k M app = 0.09 ± 0.03 mM. Inhibitory constant (K i ) for the coenzyme palmityl-CoA was determined to be 1.0 ± 0.2 μM in their buffer conditions, for a reported K i of 2μM in literature [21].…”

Tuning resonance energy transfer with magneto-optical properties of graphene

“…In Fig. (5) we depict the comparison of the RET rate using (8) and (9). It is clearly seen that the low field approximation captures a sort of average behavior, but fails to show the marked oscillations present in (8).…”

“…Different energy transfer mechanisms have been extensively discussed not only in physics, but also in several areas like chemistry, biology and engineering. An efficient energy transfer allows for a variety of applications, such as photovoltaics [6], luminescence [7,8], sensing [9], quantum information [10,11], and many others. Due to these numerous applications and to advances in different areas combined with the great development of new technologies, controlled modification of the RET rate has also become a topic of huge interest.…”

“…That led us to a result with a clear interpretation in terms of images -as r 2 + (2z) 2 is the distance between an emitter and the image of the other -but it should be recalled that such interpretation was not to be obviously expected: we are in the low conductivity regime, so these dressed images probably owe their appearance more to the plane symmetry than to the (short) distance regime. All in all, it is established that the oscillations as a function of B are a direct consequence of the interaction between the emitters and the MPPs, but to get a rough trend of the RET rate for low fields, one can turn to (9).…”