Simplified protocol for detection of protein-ligand interactions via surface-enhanced resonance Raman scattering and surface-enhanced fluorescence.
A simple and effective protocol for detections of protein-protein and protein-small molecule interactions has been developed. After interactions between proteins and their corresponding ligands, we employed colloidal silver staining for producing active substrates for surface-enhanced Raman scattering (SERS) and surface-enhanced fluorescence (SEF). Tetramethylrhodamine isothiocyanate (TRITC) and Atto610 were used for both Raman and fluorescent probes. We detected interactions between human IgG and TRITC-anti-human IgG, and those between avidin and Atto610-biotin by surface-enhanced resonance Raman scattering (SERRS) and SEF. The detection limits of the proposed SERRS-based method are comparable to those of the proposed SEF-based one, 0.9 pg/mL for anti-human IgG and 0.1 pg/mL for biotin. This protocol exploits several advantages of simplicity over other SERS and SEF-based related methods because of the protein staining-based strategy for silver nanoparticle assembling, high sensitivity from SERRS and SEF, and high stability in photostability comparing to fluorescence-based protein detections. Therefore, the proposed method for detection of protein-ligand interactions has great potential in high-sensitivity and high-throughput chip-based protein function determination.
Han
Xiao X
XX
State Key Laboratory of Supramolecular Structure and Materials, Jilin University, Changchun 130012, PR China.
Kitahama
Yasutaka
Y
Tanaka
Yuhei
Y
Guo
Jie
J
Xu
Wei Q
WQ
Zhao
Bing
B
Ozaki
Yukihiro
Y
D000818:Animals; D002417:Cattle; D005453:Fluorescence; D006801:Humans; D008024:Ligands; D011485:Protein Binding; D012680:Sensitivity and Specificity; D013059:Spectrum Analysis, Raman / Q000379:methods*; D013499:Surface Properties; D013997:Time Factors
D008024:Ligands