Label-Free Surface-Enhanced Raman Scattering Bioanalysis Based on Au@Carbon Dot Nanoprobes

Summary

In this study, we developed a low-cost surface-enhanced Raman scattering (SERS)-based fingerprint nanoprobe with favorable biocompatibility to show label-free live cell bioimaging and detect two bacterial strains, showing in detail how to get SERS spectra of living cells in a non-destructive method.

Abstract

Surface-enhanced Raman scattering (SERS) technology has attracted more and more attention in the biomedical field due to its ability to provide molecular fingerprint information of biological samples, as well as its potential in single-cell analysis. This work aims to establish a simple strategy for label-free SERS bioanalysis based on Au@carbon dot nanoprobes (Au@CDs). Here, polyphenol-derived CDs are utilized as the reductant to rapidly synthesize core-shell Au@CD nanostructures, which allows powerful SERS performance even when the concentration of methylene blue (MB) is as low as 10^-9 M, due to the cooperative Raman enhancement mechanism. For bioanalysis, Au@CDs can serve as a unique SERS nanosensor to identify the cellular components of biosamples (e.g., cancer cells and bacteria). The molecular fingerprints from different species can be further distinguished after combination with the principal component analysis. In addition, Au@CDs also enable label-free SERS imaging to analyze intracellular composition profiles. This strategy offers a feasible, label-free SERS bioanalysis, opening up a new prospect for nanodiagnosis.

Introduction

Single-cell analysis is essential for the study of revealing cellular heterogeneity and assessing the comprehensive state of the cell. The cell's instant response to the microenvironment also warrants single-cell analysis¹. However, there are some limitations to the current techniques. Fluorescence detection can be applied to single-cell analysis, but it's limited by low sensitivity. Other challenges arise from the complicated fluorescence background of cells and the fluorescence photobleaching under long-term irradiation². Surface-enhanced Raman scattering (SERS) may qualify in terms of single-cell analysis owin....

Protocol

1. Fabrication of Au@CDs

NOTE: Figure 1 illustrates a fabrication procedure for Au@CDs.

1. Prepare CD solution using citric acid (CA) and gallic acid (GA) via a typical hydrothermal treatment procedure¹⁸. Add 100 µL of 3.0 mg mL^-1 of the prepared CD solution into 200 µL of 10 mM chloroauric acid (HAuCl₄) (see Table of Materials) at room temperature for 10 s .......

Discussion

In summary, Au@CDs with an ultrathin CD shell of 2.1 nm have been successfully fabricated. The nanocomposites show superior SERS sensitivity than pure Au NPs. Also, Au@CDs possess excellent performance in reproducibility and long-term stability. Further research includes taking Au@CDs as substrates to perform SERS imaging of A549 cells³¹ and to detect two bacterial strains³². It has been proved that Au@CDs can be used as an ultrasensitive SERS probe mainly based on the.......

Materials

Name	Company	Catalog Number	Comments
10x PBS buffer (Cell culture)	Langeco Technology	BL316A
6 well cell culture plate	LABSELECT	11110
Cell Counting Kit-8 (CCK-8)	GLPBIO	GK10001
Citric acid	Shanghai Aladdin Biochemical Technology	C108869
CO2 incubator	Thermo Fisher Technologies	3111
Constant temperature magnetic agitator	Sartorius Scientific Instruments	SQP
Cryogenic high speed centrifuge	Shanghai Boxun	SW-CJ-2FD
DMEM high glucose cell culture medium	Procell	PM150210
Electronic balance	Sartorius Scientific Instruments	SQP
Enzyme marker	Thermo Fisher Technologies	3111
Fetal bovine serum	Zhejiang Tianhang Biological Technology	11011-8611
Figure 1	Figdraw.
Fourier infrared spectrometer	Thermo, America	Nicolet 380
Freeze dryer	Tecan	Infinite F50
Gallic acid	Shanghai Aladdin Biochemical Technology	G104228
Handheld Raman spectrometer	OCEANHOOD, Shanghai, China	Uspectral-PLUS
HAuCl4	Guangzhou Pharmaceutical Company (Guangzhou)
High resolution transmission electron microscope	Thermo Fisher Technologies	FEI Tecnai G2 Spirit T12
High temperature autoclave	Shanghai Boxun	YXQ-LS-50S
Inverted microscope	Nanjing Jiangnan Yongxin Optical	XD-202
LB Broth BR	Huankai picoorganism	028320
Medical ultra-low temperature refrigerator	Thermo Fisher Technologies	ULTS1368
Methylene blue	Sigma-Aldrich
Pancreatin Cell Digestive Solution	beyotime	C0207
Penicillin streptomycin double resistance	Shanghai Boxun	YXQ-LS-50S
Pure water meter	Millipore, USA	Milli-Q System
Raman spectrometer	Renishaw
Sapphire chip	beyotime
Thermostatic water bath	Changzhou Noki
Ultra-clean table	Shanghai Boxun	SW-CJ-2FD
Uv-visible light absorption spectrometer	MADAPA, China	UV-6100S
Wire 3.4	Renishaw