Multiplex, Quantitative, High-Resolution Imaging of Protein:Protein Complexes via Hybridization Chain Reaction.
ACS Chem Biol
; 19(2): 280-288, 2024 02 16.
Article
en En
| MEDLINE
| ID: mdl-38232374
ABSTRACT
Signal amplification based on the mechanism of hybridization chain reaction (HCR) facilitates spatial exploration of gene regulatory networks by enabling multiplex, quantitative, high-resolution imaging of RNA and protein targets. Here, we extend these capabilities to the imaging of proteinprotein complexes, using proximity-dependent cooperative probes to conditionally generate a single amplified signal if and only if two target proteins are colocalized within the sample. HCR probes and amplifiers combine to provide automatic background suppression throughout the protocol, ensuring that even if reagents bind nonspecifically in the sample, they will not generate amplified background. We demonstrate proteinprotein imaging with a high signal-to-background ratio in human cells, mouse proT cells, and highly autofluorescent formalin-fixed paraffin-embedded (FFPE) human breast tissue sections. Further, we demonstrate multiplex imaging of three different proteinprotein complexes simultaneously and validate that HCR enables accurate and precise relative quantitation of proteinprotein complexes with subcellular resolution in an anatomical context. Moreover, we establish a unified framework for simultaneous multiplex, quantitative, high-resolution imaging of RNA, protein, and proteinprotein targets, with one-step, isothermal, enzyme-free HCR signal amplification performed for all target classes simultaneously.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
ARN
/
Diagnóstico por Imagen
Límite:
Animals
/
Humans
Idioma:
En
Revista:
ACS Chem Biol
Año:
2024
Tipo del documento:
Article
País de afiliación:
Estados Unidos
Pais de publicación:
Estados Unidos