Rediscovery of dielectrophoresis for label-free manipulation and interrogation of single protein molecules.

Samenvatting

Proteins are the workhorses of living cells. They mediate all cellular functions from cell division, metabolism and transport to programmable cell death. Studying protein activity reveals mechanisms underlying these processes in living systems and establishes a link between diseases and protein anomalies.
In the past decade protein analysis has been downscaled to the single-molecule level, capturing real-time pictures of biological processes. However, most of the available single-molecule techniques for studying proteins require laborious fluorescent labelling and chemical tethering, lowering throughput and prompting questions about the biological relevance of the results. In turn, current label-free approaches are not selective and provide only limited insight into protein behavior.
I propose a radically novel method to interrogate proteins and their behavior, through manipulating a single protein molecule without chemical tethering while simultaneously optically monitoring its physical properties (mass, hydrodynamic radius and low-frequency polarizability) in a label-free fashion.
The core of my approach is controlling the position of the single protein molecule through dielectrophoresis, the actuation of a polarizable object’s position by a highly inhomogeneous electric field. I will apply dielectrophoresis to protein studies by employing graphene nanoelectrodes, which can generate strong enough electric fields to trap single protein molecules. To identify and characterize the trapped molecule I will use interferometric scattering (iScat) microscopy, by which the molecular mass of single protein molecule can be inferred. Moreover, single-molecule manipulation will allow me to measure protein hydrodynamic radii and polarizations combining iScat with lock-in detection. The synergy of iScat and dielectrophoresis opens new avenues for biophysics research as it enables selective label-free protein analysis at a single-molecule level.
My experience with nanofabrication and single-molecule biosensing combined the expertise of the Orrit lab in single-molecule optics make a perfect fit to complete this project.

Kenmerken

Projectnummer

VI.Veni.192.162

Hoofdaanvrager

Dr. S. Pud

Verbonden aan

Universiteit Leiden, Faculteit der Wiskunde en Natuurwetenschappen, Leids Instituut voor Onderzoek in de Natuurkunde (LION)

Uitvoerders

Dr. S. Pud

Looptijd

01/01/2020 tot 01/02/2023