Disruption of Cultured Cells by Nitrogen Cavitation

INTRODUCTION

Cell disruption by nitrogen decompression from a pressurized vessel is a rapid and effective way to homogenize cells and tissues, to release intact organelles, and to prepare cell membranes. Cells are placed in a pressure vessel and large quantities of oxygen-free nitrogen are dissolved in the cells under high pressure (~5500 kilopascals [kPa], equivalent to 800 pounds per square inch [psi]). When the pressure is released suddenly, the nitrogen bubbles out of solution, rupturing the cell membrane and releasing the cell contents. Nitrogen cavitation is well suited for mammalian and plant cells and fragile bacteria, but is less effective with yeast, fungi, spores, or other cell types with tough cell walls. The chemical and physical stresses imposed by nitrogen cavitation on enzymes and subcellular compartments are minimized compared with ultrasonic and mechanical homogenizing methods. Unlike lysis methods relying on shear stresses and friction, there is no heat damage to proteins and organelles during nitrogen cavitation. Indeed, the method is accompanied by an adiabatic expansion that cools the sample instead. Also, labile cell components are protected from oxidation by the inert nitrogen gas. Furthermore, nitrogen does not alter the pH of the suspending medium. The process is fast and uniform because the same disruptive forces are applied within each cell and throughout the sample, ensuring reproducible cell-free homogenates. Finally, variable sample sizes (e.g., from ~1 mL to 1 L or more) can be accommodated with most commercial systems.