Single cell analysis inside Environmental Scanning Electron Microscope (ESEM)-nanomanipulator system

Abstract

The conventional approach to characterize cellular biology is called biochemistry. This developed science is used for studying physiological aspects, mainly genetics, by characterizing protein and other biomaterials. Since single cells are difficult to study, a collection of cells are used for characterizing cellular physiology and inturn used to describe behavior of single cell (Brehm-Stecher & Johson, 2004). However, in addition to this advance understanding of cellular genetics, information about mechanical properties of cells is also needed. The molecular structure of the cell-wall is only partially understood, and its mechanical properties are an area of “near-total darkness” (Harold, 2005). Moreover, the approximation of single cell behavior from a group used in conventional approach also requires further justification whether it can be applied to all cell types (Shapiro, 2000). The knowledge of the cell mechanics could be valuable in the future for biomedical applications, for example, variations in cell mechanics of healthy and unhealthy cells can be linked to a specific disease. Available experimental techniques to probe single cells include micropipette aspiration, optical tweezers, magnetic tweezers (Bausch et al., 1999), atomic/molecular force probes (Gueta et al., 2006), nanoindenters, microplate manipulators, optical stretchers (Thoumine et al., 1999) and nanoneedle (Obataya et al., 2005).. The functionality of nanoneedles is not limited only to the stiffness measurements but it can also be used for single cell surgery (Leary et al., 2006) which can be further applied to a novel single cell drug delivery system (Bianco et al., 2005) or as a delivery tool for nanoparticles (Brigger et al., 2002). Conventional drug therapy suffers from the problems of inefficacy or nonspecific effects; hence, nanosystems are being developed for targeted drug delivery (Stylios et al., 2005). In order to successfully deliver materials; e.g. bioactive peptide, proteins, nucleic acids or drugs inside the cell, carriers must be able to penetrate the cell wall or cell membrane without causing death or create any mechanotransduction to the cell (Goodman et al., 2004), i.e. the process of converting physical forces to biochemical signals and integrating them into cellular responses (Huang et al., 2004). Therefore, the knowledge of biomechanics of the cell is crucial in providing prior-estimation of required insertion force to deliver drug material inside the cell. Without having this information, the insertion process may be unsuccessful due to inadequate applied force or the cell may be seriously damaged due to the excessive applied force. This chapter focuses on the following two needs, i.e. the needs for the understanding of the mechanical properties of single cells and the needs for the novel nanotools for the single cells probing. The first need was fed by highlighting our findings on the effects of three factors, i.e. cell sizes, environmental conditions and growth phases, on the strength of the single W303 yeast cells. The second need was served by showing our findings on the development of nanoneedles which can be used for single cell local stiffness characterizations and single cell surgery.