Multiplex Protein Profiling by Low-Signal-Loss Single-Cell Western Blotting with Fluorescent-Quenching Aptamers.
Anal Chem
; 95(30): 11399-11409, 2023 08 01.
Article
en En
| MEDLINE
| ID: mdl-37458448
ABSTRACT
Single-cell western blotting (scWB) is a prevalent technique for high-resolution protein analysis on low-abundance cell samples. However, the extensive signal loss during repeated antibody stripping precludes multiplex protein detection. Herein, we introduce Fluorescent-quenching Aptamer-based Single-cell Western Blotting (FAS-WB) for multiplex protein detection at single-cell resolution. The minimal size of aptamer probes allows rapid in-gel penetration, diffusion, and elution. Meanwhile, the fluorophore-tagged aptamers, coordinated with complementary quenching strands, avoid the massive signal loss conventionally caused by antibody stripping during repeated staining. Such a strategy also facilitates multiplex protein analysis with a limited number of fluorescent tags. We demonstrated FAS-WB for co-imaging four biomarker proteins (EpCAM, PTK7, HER2, CA125) at single-cell resolution with lower signal loss and enhanced signal-to-noise ratio compared to conventional antibody-based scWB. Being more time-saving (less than 25 min per cycle) and economical (1/1000 cost of conventional antibody probes), FAS-WB offers a highly efficient platform for profiling multiplex proteins at single-cell resolution.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Aptámeros de Nucleótidos
Idioma:
En
Revista:
Anal Chem
Año:
2023
Tipo del documento:
Article
País de afiliación:
China