Array-Based Protein Sensing Using an Aggregation-Induced Emission (AIE) Light-Up Probe

Abstract

Protein detection and identification are important for the diagnosis of diseases; however, the development of facile sensing probes still remains challenging. Here, we present an array-based "turn on" protein-sensing platform capable of detecting and identifying proteins using aggregation-induced emission luminogens (AIEgens). The water-soluble AIEgens in which fluorescence was initially turned off showed strong fluorescence in the presence of nanomolar concentrations of proteins via restriction of the intramolecular rotation of the AIEgens. The binding affinities between the AIEgens and proteins were associated with various chemical functional groups on AIEgens, resulting in distinct fluorescent-signal outcomes for each protein. The combined fluorescence outputs provided sufficient information to detect and discriminate proteins of interest by linear discriminant analysis. Furthermore, the array-based sensor enabled classification of different concentrations of specific proteins. These results provide novel insight into the use of the AIEgens as a new type of sensing probe in array-based systems.

Scheme 1. Representative Illustration of a Protein Sensor Based on Fluorescence-Pattern Recognition

(a) Mechanism of AIE fluorescence in the presence of proteins. (b) Turn on fluorescence sensors using an array involving AIEgens and five proteins. Following normalization of the fluorescence intensity, LDA was performed to classify each protein.

Figure 1

Schematic of the synthesis and chemical structure of four AIEgens possessing AIE and excited state of intramolecular proton-transfer properties (AIE-1–4). N₂H₄·H₂O, hydrazine monohydrate; TFA, trifluoroacetic acid.

Figure 2

Optical studies of AIE phenomena in water-THF mixtures and water alone. (a) UV–vis absorption spectra and (b) fluorescence spectra of AIE-1 in water (black line) and in a mixture of water/THF (v/v, 5:95) (red line). λ_ex = 365 nm; [AIE-1] = 40 μM.

Figure 3

Fluorescence spectrum of AIE-2 in the absence and presence of esterase. The inset shows images of AIE-2 solution before and after the addition of 500 nM esterase. λ_ex = 365 nm; [AIE-2] = 40 μM.

Figure 4

Array-based sensing of five proteins (500 nM) using four AIEgens (40 μM) in water at room temperature. (a) Fluorescence pattern of the synthesized AIEs (AIE-1–4) in the presence of the five proteins (BSA, esterase, transferrin, fibrinogen, and β-galactosidase) for subsequent fluorescence experiments. Each value is an average of six parallel measurements. (b) LDA analysis using two-dimensional (2D) plots with 95% ellipse confidence.

Figure 5

Array-based sensing and discrimination of BSA and esterase at different concentrations. (a) Fluorescence patterns of AIE-1 to AIE-4 (40 μM) in the presence of BSA and esterase at 200, 500 nM, 1, and 2 μM. Each value represents an average of six parallel measurements. (b) LDA analysis of the results using 2D with 95% ellipse confidence.