Synthesis of Fluorescent Gold Nanodot–Liposome Hybrids for Detection of Phospholipase C and Its Inhibitor

Abstract

We report the synthesis of fluorescent 11-mercaptoundecanoic acid-gold nanodot-liposome (11-MUA-Au ND/Lip) hybrids by incorporation of gold nanoparticles (∼3 nm) and 11-MUA molecules in hydrophobic phospholipid membranes that self-assemble to form small unilamellar vesicles. A simple and homogeneous fluorescence assay for phospholipase C (PLC) was developed on the basis of the fluorescence quenching of 11-MUA-Au ND/Lip hybrids in aqueous solution. The fluorescence of the 11-MUA-Au ND/Lip hybrids is quenched by oxygen (O2) molecules in solution, and quenching is reduced in the presence of PLC. PLC catalyzes the hydrolysis of phosphatidylcholine units from Lip to yield diacylglycerol (DAG) and phosphocholine (PC) products, leading to the decomposition of Lip. The diacylglycerol further interacts with 11-MUA-Au NDs via hydrophobic interactions, leading to inhibition of O2 quenching. The 11-MUA-Au ND/Lip probe provides a limit of detection (at a signal-to-noise ratio of 3) of 0.21 nM for PLC, with high selectivity over other proteins, enzymes, and phospholipases. We have validated the practicality of using this probe for the determination of PLC concentrations in breast cancer cells (MCF-7 and MDA-MB-231 cell lines) and nontumor cells (MCF-10A cell line), revealing that the PLC activity in the first two is at least 1.5-fold higher than that in the third. An inhibitor assay using 11-MUA-Au ND/Lip hybrids demonstrated that tricyclodecan-9-yl potassium xanthate (D609) inhibits PLC (10 nM) with an IC50 value of 3.81 ± 0.22 μM. This simple, sensitive, and selective approach holds great potential for detection of PLC in cancer cells and for the screening of anti-PLC drugs.

Photoluminescent Gold Nanomaterials as Sensitive Probes

“…11-MUA-Au NDs conjugated with platelet-derived growth factors (PDGF) were used for quantitation of PDGF receptor, with great selectivity and sensitivity (LOD of 0.25 nM) [10]. Au NDs prepared from Au nanoparticles using different etching agents have been used for quantiation of various analytes, including S 2-, NO 2 -, concanavalin A (Con A), Phospholipase C, and so on [11][12][13][14]. Based on analyte-induced PL quenching, lysozyme-Au NCs were used for quantitation of Hg 2+ , with LODs of 10 nM [15].…”

Synthesis, Optical Properties, and Sensing Applications of Gold Nanodots

“…Absorption bands located at 393 nm and 275 nm were evidence for the existence of Au complexes with 11‐MUA and TSH, respectively, on the surfaces of Au NDs. Conjugating liposomes (Lip) with 11‐MUA–Au NDs led to the formation of photoluminescent 11‐MUA–Au ND/Lip hybrids, which were used to detect biomolecules like phospholipase C (PLC) 21. A mixture of 1,2‐dimyristoyl‐ sn ‐glycero‐3‐phosphocholine (DMPC), 11‐MUA, and cholesterol was used to prepare multilamellar vesicles (MLVs) containing THPC–Au NPs.…”

“…This probe allows detection of protein G via a competitive assay over a concentration range of 100–750 nM with an LOD of 85 nM. 11‐MUA–Au ND/Lip hybrids were prepared for the detection of phospholipase C (PLC) down to 0.2 nM, in which 11‐MUA–Au NDs were distributed in the interior vesicle of the liposome 21. Due to self‐quenching and high local oxygen concentration inside the liposome, the PL intensity of 11‐MUA–Au ND/Lip hybrids is lower than that of 11‐MUA–Au NDs.…”

Synthesis of ultrastable and multifunctional gold nanoclusters with enhanced fluorescence and potential anticancer drug delivery application

“…However, the former method is sensitive to salt concentration, and the latter is usually not suitable for ultrasmall nanoparticles especially when using long polyethylene glycol chains. On the other hand, noble metal nanoclusters, consisting of several to several hundred atoms, also have drawn much attention due to their unique physiochemical properties [5][6][7], catalytic activities [8], ion detection capability [9][10][11][12][13] and potential applications in bio-labeling and sensing [14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29][30]. At the same time, Au NCs can be synthesized using different capping molecules, such as proteins [31][32][33], DNA and nucleotides [34,35], peptides [11,[36][37][38][39], polymers [40][41][42][43][44] and small molecules such as dihydrolipoic acid [10], diphosphine [45], histidine [46,47], p...…”