TOF-SIMS IMAGING OF SINGLE CELLS IN A TREHALOSE MATRIX

Open Access
Author:
Parry, Shawn
Graduate Program:
Chemistry
Degree:
Doctor of Philosophy
Document Type:
Dissertation
Date of Defense:
November 08, 2007
Committee Members:
  • Nicholas Winograd, Committee Chair
  • Robert Allen Schlegel, Committee Member
  • Albert Welford Castleman Jr., Committee Member
  • David Lawrence Allara, Committee Member
Keywords:
  • Mass Spectrometry
  • Trehalose
  • TOF-SIMS
Abstract:
Mapping the chemical composition of single cells at the submicron level could provide abundant insight into cellular functions, such as understanding signaling pathways and cellular phagocytosis. Secondary Ion Mass Spectrometry (SIMS) imaging can be used to create in situ chemical maps of a host of biologically relevant species. Macrophage cells are indispensable constituents of the animal immune system. Acting as cellular garbage disposals, macrophages keep the blood stream clean by recycling lipid droplets, waste particulate, dead cells, and foreign species through a process called phagocytosis. Shown in Chapters 3, 4, and 5 of this thesis are lipid membrane images and calcium depth profiles of phagocytic and inactive or dying macrophages. These images represent a novel approach at identifying the dynamic cellular events related to phagocytosis and demonstrate SIMS ability to simultaneously image multiple cell-related signals at submicron resolutions – difficult to achieve with other methodologies. This combination of imaging and profiling is significant because it provides the possibility of defining previously unknown phagocytic mechanisms. A clearer picture of phagocytosis will enhance understanding of the immune system. Along with the pertinence and scope of single-cell imaging, Chapter 1 of this thesis presents the unique capabilities of SIMS in the biological imaging modality. The novel application of a disaccharide matrix to ultra-high vacuum (UHV) analyses, such as used in SIMS, is detailed in Chapters 2 and 3. Finally included, is the direction of SIMS imaging to relevant biological applications. The aim of this thesis is to demonstrate single-cell SIMS imaging capabilities, provide insight into membrane function during phagocytosis, and illustrate a complimentary technique to cryogenics for UHV analyses.