p53 Detection by Fluorescence Lifetime on a Hybrid Fluorescein Isothiocyanate Gold Nanosensor

Abstract

p53 is a tumor suppressor whose detection in the body is highly valuable as marker for early cancer diagnosis and prognosis. In this report we have studied constructs based on gold nanoparticles (NPs) functionalized with specific anti-p53 antibodies and with a fluoresceine derivative, FITC. The interaction of surface plasmons on gold NPs few nm in size, with fluorophores bound within few nanometers from the surface, induces changes in the fluorophore excited state lifetime. This parameter follows linearly the p53 concentration in solutions up to 200-400 pM, depending on the size of the NP, with an uncertainty approximatley =25 pM. We have evaluated the specificity of the nanosensor for p53 by testing it against bovine serum albumin, beta-lactoglobulin and lysozyme solutions. The titration of total cell extracts from p53 positive or p53-null cells with the anti-p53 antibody decorated gold NPs indicates that this construct is promising for possible applications to in vivo screening.

Thermal and Chemical Stability of Thiol Bonding on Gold Nanostars

“…PEG-coated GNPs display improved pharmacokinetic properties; PEG enhances in vitro stability of GNPs in saline buffers or culture media and it allows the coated nanoparticles to evade macrophage-mediated uptake and removal from systemic circulation in vivo . PEG coating is also employed for nanoparticle specific functionalization, as many commercial PEGs in addition to a thiol function suitable for grafting on gold feature also a remote function (e.g., −OH, −COOH, −NH 2 ) that may be used for further chemical modification. , GNPs modified with fluorescent dyes are also proposed for biomedical applications as image-enhancing agents , and as biosensors, , with the dyes typically grafted to the gold surface through a remote thiol moiety. It must also be added that GNPs have relevant applications in hyperthermal treatments, as when irradiated on their LSPR they undergo thermal relaxation, with a highly localized temperature increase.…”

A Molecular Thermometer for Nanoparticles for Optical Hyperthermia

“…The ESL of RhB + on nanoparticles as a function of the bulk temperature (controlled by a thermostatted bath, see Materials and Methods in SI) was measured on diluted suspensions of RhB + decorated nanoparticles under two-photon excitation at λ = 800 nm (80 MHz rep. rate; 250 fs pulse width, beam waist ≅ 0.8 μm, average power ⟨ P ⟩ ≅ 2–4 mW). In this case the fluorescence traces are characterized by bursts (Figure B) that correspond to the two-photon primed emission of a number of dyes adsorbed on the diffusing nanoparticles , on a flat background that is due to residual unbound dyes probably intermixed to PSS polymer. It is noteworthy that rare fluorescence bursts were detected from suspensions of nanoparticles decorated with PAH and incubated with RhB, similarly to what observed for undecorated nanoparticles (Figure B).…”

Gold Nanostar Synthesis and Functionalization with Organic Molecules

“…The immobilization of dye molecules onto nanoparticles induces dramatic changes in its optical properties for chemical and biological applications, including fl uorescence quenching of small dye molecules on gold nanoparticles. This effect can be exploited, for example, for protein sensing approaches [ 70 ]. Complementary oligonucleotides for single-stranded DNA-linked metal nanoparticles or bar-coded metal nanowires and fl uorescent-dye-doped nanoparticles have been developed for medical diagnostics and labeling.…”