This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Method Article
Direct Synthesis of Gold Nanoparticles on Cysteine-rich Tags in Yeast Cells
Wanzhong He
National Institute of Biological Sciences, Beijing, 102206, China
Corresponding Author
License:
This work is licensed under a CC BY 4.0 License. Read Full License
We developed a novel auto-nucleation suppressed mechanism (ANSM) for direct synthesis of EM-visible gold nanoparticles (AuNPs) on cysteine-rich tags (e.g., metallothionein) in cells for single-molecule detection with electron microscopy (it accompanies our Nature Method manuscript, Jiang et al. 2020 [1]). Both tagged-fusion proteins expressed in cells (e.g.bacteria, yeast and mammalian cells) and antigens stained with antibody-tag fusion proteins can be visualized by this protocol. Here we describe the typical protocols (both the chemical fixation and the high pressure freezing cases) developed for ANSM-based AuNP synthesis in yeast cells expressing metallothionein (MTn) tags (Figure 1). This approach should be useful for EM visualization of single-molecule in yeast cells, and easier adapted for bacterial cells.
Keywords
Cysteine-rich tag, electron microscopy, gold nanoparticle, yeast cell, Schizosaccharomyces pombe, metallothionein, AuNP synthesis, single-molecule chemical fixation, oxidization, high pressure freezing, freeze-substitution fixation, gold enhancement, imaging, ultrastructure, labeling
Figure 1
Figure 2
This is a list of supplementary files associated with this preprint. Click to download.
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 10 Aug, 2020
No community comments so far
Metrics
Comments: 0
HTML Views: ...
Associated Publications
Learn more about our company.
This is a preprint. It has not completed peer review.
Method Article
Direct Synthesis of Gold Nanoparticles on Cysteine-rich Tags in Yeast Cells
Wanzhong He
National Institute of Biological Sciences, Beijing, 102206, China
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 10 Aug, 2020
No community comments so far
Metrics
Comments: 0
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
We developed a novel auto-nucleation suppressed mechanism (ANSM) for direct synthesis of EM-visible gold nanoparticles (AuNPs) on cysteine-rich tags (e.g., metallothionein) in cells for single-molecule detection with electron microscopy (it accompanies our Nature Method manuscript, Jiang et al. 2020 [1]). Both tagged-fusion proteins expressed in cells (e.g.bacteria, yeast and mammalian cells) and antigens stained with antibody-tag fusion proteins can be visualized by this protocol. Here we describe the typical protocols (both the chemical fixation and the high pressure freezing cases) developed for ANSM-based AuNP synthesis in yeast cells expressing metallothionein (MTn) tags (Figure 1). This approach should be useful for EM visualization of single-molecule in yeast cells, and easier adapted for bacterial cells.
Figure 1
Figure 2