Ultraefficient Cap-Exchange Protocol To Compact Biofunctional Quantum Dots for Sensitive Ratiometric Biosensing and Cell Imaging

Abstract

An ultraefficient cap-exchange protocol (UCEP) that can convert hydrophobic quantum dots (QDs) into stable, biocompatible, and aggregation-free water-dispersed ones at a ligand:QD molar ratio (LQMR) as low as 500, some 20–200-fold less than most literature methods, has been developed. The UCEP works conveniently with air-stable lipoic acid (LA)-based ligands by exploiting tris(2-carboxylethyl phosphine)-based rapid in situ reduction. The resulting QDs are compact (hydrodynamic radius, Rh, < 4.5 nm) and bright (retaining > 90% of original fluorescence), resist nonspecific adsorption of proteins, and display good stability in biological buffers even with high salt content (e.g., 2 M NaCl). These advantageous properties make them well suited for cellular imaging and ratiometric biosensing applications. The QDs prepared by UCEP using dihydrolipoic acid (DHLA)-zwitterion ligand can be readily conjugated with octa-histidine (His8)-tagged antibody mimetic proteins (known as Affimers). These QDs allow rapid, ratiometric detection of the Affimer target protein down to 10 pM via a QD-sensitized Förster resonance energy transfer (FRET) readout signal. Moreover, compact biotinylated QDs can be readily prepared by UCEP in a facile, one-step process. The resulting QDs have been further employed for ratiometric detection of protein, exemplified by neutravidin, down to 5 pM, as well as for fluorescence imaging of target cancer cells.

Quantum Dots: A Review from Concept to Clinic

“…[120] In addition to amphiphilic oligomers, a couple of surfactants are also used to make desirable modifications, including cetyltrimethylammonium bromide (CTAB), [131] gemini surfactants, [132] and pegylated-phospholipids. [133] As previously noted, two types of amphiphilic polymers have been used with reference to this strategy, including phospholipidtype low-molecular-weight polymers and high-molecular-weight polymers. [134] The phospholipid-type low-molecular-weight polymers are able to form oil-in-water micelles using hydrophobic interactions between the lipid hydrophobic portion and the hydrophobic surface ligands of the QDs.…”

Alternative reagents to antibodies in imaging applications

“…The reduced circulation time and more rapid tumour penetration afforded by smaller binding proteins compared to antibodies provides the potential for a faster and timelier imaging procedure and thus should reduce patient time in hospital. Another approach with promise for in vivo tumour imaging involves dye-conjugated Affimers that have been used in Förster resonance transfer (FRET) experiments (Conway et al 2014 ; Wang et al 2017b ).…”

“…The binding region is generated from sequences in two variable loops presented between pairs of β-sheets. Affimers have been raised against a diverse set of targets, thereby demonstrating their utility in many different molecular biology applications, including those related to bio-imaging (Fisher et al 2015 ; Kyle et al 2015 ; Raina et al 2015 ; Sharma et al 2016 ; Arrata et al 2017 ; Koutsoumpeli et al 2017 ; Tiede et al 2017 ; Wang et al 2017b ).…”

Nanostructure and Corresponding Quenching Efficiency of Fluorescent DNA Probes

“…The field of fluorescent DNA probes improved considerably with the introduction of inorganic nanoparticles, such as quantum dots (QDs) [ 4 , 5 ] and gold (Au) nanoparticles (AuNPs) [ 6 , 7 , 8 ] as energy donors and acceptors [ 9 ], respectively. As a new kind of luminescent inorganic fluorophores, QDs are being widely used in chemical sensors [ 10 ], DNA detection [ 11 ], cell labeling [ 12 ], and imaging [ 13 ] because they have a broad and continuous excitation spectrum, a narrow size-tunable symmetric emission spectrum, and a high fluorescence quantum yield. AuNPs are widely used in gene delivery [ 14 ] and cell labeling [ 15 ] because they are bioinert, nontoxic, and readily synthesized and functionalized.…”