Molecular-receptor-specific, non-toxic, near-infrared-emitting Au cluster-protein nanoconjugates for targeted cancer imaging

Abstract

Molecular-receptor-targeted imaging of folate receptor positive oral carcinoma cells using folic-acid-conjugated fluorescent Au₂₅ nanoclusters (Au NCs) is reported. Highly fluorescent Au₂₅ clusters were synthesized by controlled reduction of Au⁺ ions, stabilized in bovine serum albumin (BSA), using a green-chemical reducing agent, ascorbic acid (vitamin-C). For targeted-imaging-based detection of cancer cells, the clusters were conjugated with folic acid (FA) through amide linkage with the BSA shell. The bioconjugated clusters show excellent stability over a wide range of pH from 4 to 14 and fluorescence efficiency of ~5.7% at pH 7.4 in phosphate buffer saline (PBS), indicating effective protection of nanoclusters by serum albumin during the bioconjugation reaction and cell-cluster interaction. The nanoclusters were characterized for their physico-chemical properties, toxicity and cancer targeting efficacy in vitro. X-ray photoelectron spectroscopy (XPS) suggests binding energies correlating to metal Au 4f_7/2˜83.97 eV and Au 4f_5/2~87.768 eV. Transmission electron microscopy and atomic force microscopy revealed the formation of individual nanoclusters of size ~1 nm and protein cluster aggregates of size ~8 nm. Photoluminescence studies show bright fluorescence with peak maximum at ~674 nm with the spectral profile covering the near-infrared (NIR) region, making it possible to image clusters at the 700-800 nm emission window where the tissue absorption of light is minimum. The cell viability and reactive oxygen toxicity studies indicate the non-toxic nature of the Au clusters up to relatively higher concentrations of 500 µg ml^-1. Receptor-targeted cancer detection using Au clusters is demonstrated on FR^+ve oral squamous cell carcinoma (KB) and breast adenocarcinoma cell MCF-7, where the FA-conjugated Au₂₅ clusters were found internalized in significantly higher concentrations compared to the negative control cell lines. This study demonstrates the potential of using non-toxic fluorescent Au nanoclusters for the targeted imaging of cancer.