All-quantum dot based Förster resonant energy transfer: key parameters for high-efficiency biosensing.

While colloidal quantum dots (QDs) are commonly used as fluorescent donors within biosensors based on Förster resonant energy transfer (FRET), they are hesitantly employed as acceptors. On the sole basis of Förster theory and the well-known behaviour of organic dyes, it is often argued that the QD absorption band over the UV-visible range is too wide. Discarding these preconceptions inherited from classical fluorophores, we experimentally examine the FRET process occurring between donor and acceptor CdTe QDs and provide a mathematical description of it. We evidence that the specific features of QDs unexpectedly lead to the enhancement of acceptors' emission (up to +400%), and are thus suitable for the design of highly efficient all-QD based FRET sensors. Our model enables us to identify the critical parameters maximizing the contrast between positive and negative biosensing readouts: the concentrations of donors and acceptors, their spectral overlap, the densities of their excitonic states, their dissipative coupling with the medium and the statistics of QD-QD chemical pairing emerge as subtle and determinant parameters. We relate them quantitatively to the measured QD-QD FRET efficiency and discuss how they must be optimized for biosensing applications.

Quenched or alive quantum dots: The leading roles of ligand adsorption and photoinduced protonation.

HYPOTHESIS: The fluorescence emission of water-soluble CdTe quantum dots (QDs) capped with mercaptocarboxylic acids (MCAs) is known to be pH-dependent. However, this behaviour is quite different from a study to another, so that literature suffers from a lack of coherence. Here we assume that the QD fluorescence efficiency is actually driven by the acid-base equilibrium of MCA thiol groups, and that light-excited QDs open a non-radiative relaxation path through photoinduced protonation. EXPERIMENTS: We address this issue by examining colloidal CdTe QDs with (time-resolved) fluorescence spectroscopy under various conditions of acidity and light excitation. FINDINGS: It appears that the emission of QDs is quenched below a critical pH value of 6.87, and that light excitation power strengthens this quenching. We thus demonstrate the existence of an additional photochemical process and developed a mathematical modeling accounting for all our experimental results. With only three parameters, it is possible to accurately predict the fluorescence decay of QDs over time, at any pH. Further, we also related the critical pH value of 6.87 to QD surface properties, explaining why observations may differ from a study to another and making the literature much more coherent.

How Quantum Dots Aggregation Enhances Förster Resonant Energy Transfer.

As luminescent quantum dots (QDs) are known to aggregate themselves through their chemical activation by carbodiimide chemistry and their functionalization with biotin molecules, we investigate both effects on the fluorescence properties of CdTe QDs and their impact on Förster Resonant Energy Transfer (FRET) occurring with fluorescent streptavidin molecules (FA). First, the QDs fluorescence spectrum undergoes significant changes during the activation step which are explained thanks to an original analytical model based on QDs intra-aggregate screening and inter-QDs FRET. We also highlight the strong influence of biotin in solution on FRET efficiency, and define the experimental conditions maximizing the FRET. Finally, a free-QD-based system and an aggregated-QD-based system are studied in order to compare their detection threshold. The results show a minimum concentration limit of 80 nM in FA for the former while it is equal to 5 nM for the latter, favouring monitored aggregation in the design of QDs-based biosensors.

A global method for handling fluorescence spectra at high concentration derived from the competition between emission and absorption of colloidal CdTe quantum dots.

We investigate the effects of the concentration of CdTe quantum dots (QDs) on their fluorescence in water. The emission spectra, acquired in right angle geometry, exhibit highly variable shapes. The measurements evidence a critical value of the concentration beyond which the intensity and the spectral bandwidth decrease and the fluorescence maximum is redshifted. To account for these observations, we develop a model based on the primary and secondary inner filter effects. The accuracy of the model ensures that the concentration dependent behaviour of QD fluorescence is completely due to inner filter effects. This result is all the more interesting because it precludes the assumption of dynamic quenching. As a matter of fact, the decrease of the emission intensity is not a consequence of collisional quenching but an effect of competition between fluorescence and absorption. We even show that this phenomenon is linked not only to the QD concentration but also to the geometric configuration of the setup. Hence, our analytical model can be easily adapted to every fluorescence spectroscopy configuration to quantitatively predict or correct inner filter effects in the case of QDs or any fluorophore, and thus improve the handling of fluorescence spectroscopy for highly concentrated solutions.

Protoporphyrin IX-Functionalized AgSiO2 Core-Shell Nanoparticles: Plasmonic Enhancement of Fluorescence and Singlet Oxygen Production.

Metal-enhanced processes arising from the coupling of a dye with metallic nanoparticles (NPs) have been widely reported. However, few studies have simultaneously investigated these mechanisms from the viewpoint of dye fluorescence and photoactivity. Herein, protoporphyrin IX (PpIX) is grafted onto the surface of silver core silica shell NPs in order to investigate the effect of silver (Ag) localized surface plasmon resonance (LSPR) on PpIX fluorescence and PpIX singlet oxygen (1 O2 ) production. Using two Ag core sizes, we report a systematic study of these photophysical processes as a function of silica (SiO2 ) spacer thickness, LSPR band position and excitation wavelength. The excitation of Ag NP LSPR, which overlaps the PpIX absorption band, leads to the concomitant enhancement of PpIX fluorescence and 1 O2 production independently of the Ag core size, but in a more pronounced way for larger Ag cores. These enhancements result from the increase in the PpIX excitation rate through the LSPR excitation and decrease when the distance between PpIX and Ag NPs increases. A maximum fluorescence enhancement of up to 14-fold, together with an increase in photogenerated 1 O2 production of up to five times are obtained using 100 nm Ag cores coated with a 5 nm thick silica coating.

Compound Droplets on Fibers.

Droplets on fibers have been extensively studied in the recent years. Although the equilibrium shapes of simple droplets on fibers are well established, the situation becomes more complex for compound fluidic systems. Through experimental and numerical investigations, we show herein that compound droplets can be formed on fibers and that they adopt specific geometries. We focus on the various contact lines formed at the meeting of the different phases and we study their equilibrium state. It appears that, depending on the surface tensions, the triple contact lines can remain separate or merge together and form quadruple lines. The nature of the contact lines influences the behavior of the compound droplets on fibers. Indeed, both experimental and numerical results show that, during the detachment process, depending on whether the contact lines are triple or quadruple, the characteristic length is the inner droplet radius or the fiber radius.

A one-step short-time synthesis of Ag@SiO2 core-shell nanoparticles.

A performance of shell-thickness precise control in silver-silica coating core-shell nanoparticles is presented. 60nm sized citrate-stabilized silver nanoparticles are directly silica coated using a modified Stöber process. Tetraethyl orthosilicate is used as a silica precursor and ammonium hydroxide as catalyst in an alcoholic solvent to promote the seeded silica growth. By simply varying the synthesis reaction time from 4 to 60min, the silica shell thickness is increased from 5.1nm to 76.4nm. This well-controlled synthesis is then transposed to 40, 80 and 100nm sized silver cores in order to show the independence of the silica shell growth on the nanoparticle core size. Optical properties, i.e. localized surface plasmon resonance, of the produced silver-silica core-shell are also investigated.

Linear and nonlinear optical properties of functionalized CdSe quantum dots prepared by plasma sputtering and wet chemistry.

We develop an innovative manufacturing process, based on radio-frequency magnetron sputtering (RFMS), to prepare neat CdSe quantum dots (QDs) on glass and silicon substrates and further chemically functionalize them. In order to validate the fabrication protocol, their optical properties are compared with those of QDs obtained from commercial solutions and deposited by wet chemistry on the substrates. Firstly, AFM measurements attest that nano-objects with a mean diameter around 13 nm are located on the substrate after RFMS treatment. Secondly, the UV-Vis absorption study of this deposited layer shows a specific optical absorption band, located at 550 nm, which is related to a discrete energy level of QDs. Thirdly, by using two-color sum-frequency generation (2C-SFG) nonlinear optical spectroscopy, we show experimentally the functionalization efficiency of the RFMS CdSe QDs layer with thiol derived molecules, which is not possible on the QDs layer prepared by wet chemistry due to the surfactant molecules from the native solution. Finally, 2C-SFG spectroscopy, performed at different visible wavelengths, highlights modifications of the vibration mode shape whatever the QDs deposition method, which is correlated to the discrete energy level of the QDs.

Adsorption properties of the penicillin derivative DTPA on gold substrates.

Despite the large number of articles and patents dealing with penicillin and other beta-lactam antibiotics, there have been no reports about the self-assembly of such substances as monolayers on gold surfaces. The main reason stems from the high reactivity of the beta-lactam ring, which hinders the development of molecules possessing this entity together with a metal-anchoring function. Herein, we present the synthesis of a novel molecule, 6-[(R,S)-5-(1,2-dithiolan-3-yl)pentanoyl-amino]-penicillanic acid, which combines the beta-lactam ring and a metal-anchoring group. Using spectroscopic tools, we demonstrate the chemisorption of this compound on gold as self-assembled monolayers without any alteration of the penicillin pharmacophore and document its reactivity towards a penicillin-binding protein, BlaR-CTD. Our work is a preliminary step towards the development of new biosensors and well-ordered protein arrays, both based on the high affinity of penicillin for penicillin-binding proteins.

Sum-frequency generation spectroscopy of DNA monolayers.

The anchoring of thiolated single-stranded DNA (HS-ssDNA) monolayers onto platinum substrates was investigated by sum-frequency generation spectroscopy. Different buffer solutions were used for the preparation of the adlayers. Vibrational fingerprints in the 2700-3100 cm(-1) spectral range showed the intercalation of Tris/EDTA (TE) buffer molecules within the HS-ssDNA self-assembled monolayer. Buffer contribution to SFG can be quenched either by using SFG inactive molecules like KH(2)PO(4)/K(2)HPO(4)/NaCl (PBS) or by repeated rinsing of the DNA layer with pure water. Comparing the SFG spectra of HS-ssDNA and mercaptohexanol (MCH), which had been self-assembled onto the same substrate, enabled us to infer ordering of the anchor arms and strong disordering of the DNA strands of HS-ssDNA monolayers self-assembled on platinum.

Probing ligand-protein recognition with sum-frequency generation spectroscopy: the avidin-biocytin case.

Infrared/visible sum-frequency generation (SFG) spectroscopy is used to study the recognition of a protein (avidin) by a derived vitamin (biocytin) adsorbed on a calcium fluoride substrate. The specificity of the process is tested by replacing avidin with bovine serum albumin or presaturated avidin. The SFG spectroscopy shows drastic modifications in the CH and NH spectral ranges only upon exposure of the biocytin film to avidin. The comparison of the SFG data with Fourier transform infrared reflection absorption spectra (FT-IRRAS) in the same spectral ranges illustrates the advantages of nonlinear spectroscopy for studying and detecting recognition between biomolecules.

Picosecond laser for performance of efficient nonlinear spectroscopy from 10 to 21 microm.

Laser tunability from 10 to 21 microm is obtained by use of an optical parametric oscillator based on a KTP crystal followed by a difference-frequency stage with a CdSe crystal. An all-solid-state picosecond Nd:YAG oscillator mode locked by a frequency-doubling nonlinear mirror is used for synchronous pumping.