Imaging and Manipulation of Extracellular Traps by Atomic Force Microscopy.

Neutrophil extracellular traps (NETs) are part of an immunological response and one of the mechanisms by which neutrophils protect the host from pathogen invasion and proliferation. Notwithstanding their protective role, NETs have also been linked to the development of a variety of disorders, including cardiovascular and autoimmune diseases. Since the first reports on NETs in 2004 it has been possible to image NETs by a variety of imaging techniques. Despite this, such reports seldomly include contact probe methods, and therefore lack the unique insights such techniques typically provide. In fact, more than 10 years have passed since the discovery of NETs, and although their importance as part of a unique cellular response mechanism has become very clear, studies that attempt to address them by atomic force microscopy (AFM) remain very limited. Particularly striking is the almost absent information on the mechanical properties of NETs, and factors that may influence them. The fact that NETs are a particularly adhesive network of filaments poses a considerable technical challenge for contact probe methods and can limit advances involving either imaging or manipulation of NETs by AFM. The current set of protocols aims at aiding a knowledgeable AFM operator to obtain AFM images and to perform force spectroscopy experiments with such samples. A variety of different topics, including sample preparation and data analysis, are discussed.