Surface-modified complex SU-8 microstructures for indirect optical manipulation of single cells

Abstract

Abstract:We introduce a method that combines two-photon polymerization (TPP) and surface functionalization to enable the indirect optical manipulation of live cells. TPP-made 3D microstructures were coated specifically with a multilayer of the protein streptavidin and nonspecifically with IgG antibody using polyethylene glycol diamine as a linker molecule. Protein density on their surfaces was quantified for various coating methods. The streptavidin-coated structures were shown to attach to biotinated cells reproducibly. We performed basic indirect optical micromanipulation tasks with attached structure-cell couples using complex structures and a multi-focus optical trap. The use of such extended manipulators for indirect optical trapping ensures to keep a safe distance between the trapping beams and the sensitive cell and enables their 6 degrees of freedom actuation.

High accuracy indirect optical manipulation of live cells with functionalized microtools

“…First we have to note that cell attachment to this type of structure is as fast as was found for the previous type [16] : within seconds after the structure is pushed against the cell, they are permanently linked. The shape of the manipulator is designed such that the three spheres form a 16m side length triangle; therefore, when the structure is rotated such that the spheres are above each other, the beam trapping the upper sphere is minimally disturbed by the lower sphere.…”

“…In our earlier paper we introduced a microtool for indirect trapping consisting of four spheroids and a flat attachment plate [16] . The four spheroids were trapped by a static HOT holding the cell-structure couple while the microfluidic environment was translated along one of the three spatial directions.…”

“…The cells (K562 cell line, provided by Ervin Welker, Institute of Biochemistry, Biological Research Centre) were attached to the structures exploiting the strong streptavidin-biotin linkage. For this the structures were functionalized to create a thin layer of streptavidin film on them, and the cells were treated with water soluble sulfo-NHS biotin that binds to their surface proteins, as described earlier [16] . In order to visualize cell nucleus, we selectively stained it by Hoechst 33342 fluorophore (abs.…”

Strategies for Optical Trapping in Biological Samples: Aiming at Microrobotic Surgeons

“…Other microrobot designs from our group include wave‐guided optical waveguides and disk‐tools for localized heating in microfluidic channels . Kelemen's group employed SU‐8‐based microtools for indirect optical micromanipulation of single live cells . Figure E shows one of these microtools used for cellular manipulation.…”

Imaging the cell

“…Our second invited speaker, John Briggs (EMBL, Germany), will discuss the use of cryoelectron tomography to determine protein structures within complex environments, an imaging technology which has enabled unprecedented insight into the architecture of macromolecular machines in the complex milieu of cells (Dodonova et al 2017). Our first contributed talk in the session is then from Lóránd Kelemen (Biological Research Centre, Hungary), who will tell us about the application of indirect optical micromanipulation in fluorescent 3D live-cell imaging, an interesting new approach which enables 3D reconstruction of single cells by imaging them at different orientations (Aekbote et al 2015). Our third invited speaker is James McNally (Helmholtz-Zentrum Berlin, Germany), who will talk on the topic of cryo X-ray tomography to enable 3D cellular imaging of the ultrastructure of intact cells without the use of fixation or staining (Schneider et al 2010).…”