Precision Mems Nano-cutting Device for Cellular Microsurgery

An important tool for biological research and microsurgery is a microdevice for the cutting and isolation of subcellular neuronal components such as axons and dendrites for analysis or microsurgery. We have fabricated an easy-to-use, inexpensive and robust MEMS device with a nanoscale cutting tool that performs highly reproducible cutting of axons and dendrites. The device consists of a knife with an 20 nm-sharp edge ranging from 10-200 microns in length and is formed from molding conformally deposited silicon nitride over a potassium hydroxide-etched trench in <100>-oriented single crystal silicon. Knife surfaces are coated with a thin layer of liquid perfluorinated polyether to prevent adhesion of debris from cut targets. The knife is assembled onto a microfabricated suspension and frame consisting of serpentine flexures of single crystal silicon. These supporting structures help to properly orient the knife and control cutting force. We have used this assembled nano-cutting device to make reliable cuts of individual living dendrites and unmyelinated and myelinated axons from both adult and embryonic animal tissue. The cutting device was able to target and cut specific cell processes within a complex field and without disturbing surrounding structures. The cuts were sharp and repeatable, and microdevice’s performance was undiminished with repeated use. INTRODUCTION The integration of microand nanoscale technologies with traditional biological analytical methods will enable unprecedented studies of cellular and subcellular structures and their roles in health and disease. The fields of genomics and proteomics, for example, stand to benefit greatly from the introduction of a new generation of tools to manipulate and analyze microscale biological samples. A key element of this integration is appropriately scaled devices for the cutting, dissection, and isolation of tissue, cells, or subcellular components for analysis or for microsurgery. To date, subcellular cutting in biology has utilized conventional razor blades, glass micropipettes [1-5], or focused laser ablation systems [6]. Razor blades and glass pipettes result in crude, non-repeatable cuts, and often damage large portions of the cell or surrounding tissues. Laser tools, while providing accurate and repeatable cutting, tend to leave large areas of ablation and are limited by high cost as well as complexity of operation and in some cases, carry the risk of damaging other parts of cells. To provide an alternative to these traditional methods, we have fabricated an easy-to-use, inexpensive and robust nanocutting device using MEMS-based techniques and demonstrated its use in the precise cutting of biological samples. This microscale device consists of a knife component with a nano-sharp cutting edge that is held by a flexing suspension and frame (Fig. 1). The knife itself is a pyramid-like structure converging to a sharp cutting edge along its zenith and is formed by conformal deposition of silicon nitride over a sharp, V-shaped mold etched anisotropically into single crystal silicon by aqueous potassium hydroxide (KOH) solution. The frame and suspension holding the knife provide multiaxial compliances for the motion of the knife during actuation of cutting and is fabricated in single crystal silicon by a singlemask etch process. In biological testing, we have used this assembly to make reliable cuts of individual living dendrites (1 μm diameter), unmyelinated axons (< 2 μm diameter), as well as myelinated axons (5-10 μm diameter). The cutting device remained robust upon repeated use without diminished