Label-Free Single-Molecule Pulldown for the Detection of Released Cellular Protein Complexes.
Guangzhong MaPengfei ZhangXinyu ZhouZijian WanShaopeng WangPublished in: ACS central science (2022)
Precise and sensitive detection of intracellular proteins and complexes is key to the understanding of signaling pathways and cell functions. Here, we present a label-free single-molecule pulldown (LFSMP) technique for the imaging of released cellular protein and protein complexes with single-molecule sensitivity and low sample consumption down to a few cells per mm 2 . LFSMP is based on plasmonic scattering imaging and thus can directly image the surface-captured molecules without labels and quantify the binding kinetics. In this paper, we demonstrate the detection principle for LFSMP, study the phosphorylation of protein complexes involved in a signaling pathway, and investigate how kinetic analysis can be used to improve the pulldown specificity. We wish our technique can contribute to uncovering the molecular mechanisms in cells with single-molecule resolution.
Keyphrases
- single molecule
- label free
- induced apoptosis
- signaling pathway
- atomic force microscopy
- living cells
- sensitive detection
- protein protein
- cell cycle arrest
- high resolution
- binding protein
- amino acid
- pi k akt
- loop mediated isothermal amplification
- stem cells
- epithelial mesenchymal transition
- endoplasmic reticulum stress
- quantum dots
- cell therapy
- cell proliferation
- cell death
- single cell
- machine learning
- mass spectrometry
- real time pcr
- high speed