Pacific Rim Symposium on Surfaces, Coatings and Interfaces (PacSurf 2018) | |
Biomaterial Surfaces & Interfaces | Tuesday Sessions |
Session BI-TuE |
Session: | 35 Years of NESAC/BIO II |
Presenter: | Hua Tian, Pennsylvania State University |
Authors: | H. Tian, Pennsylvania State University N. Winograd, Pennsylvania State University |
Correspondent: | Click to Email |
The cross-talk between molecular network is central to signaling pathways that mediate cellular functionalities of all aspects. To understand the molecular mechanisms, it is necessary to unfold a complete spectrum of molecular species (e.g., lipids, metabolites and proteins) in parallel. Previously, independent molecular extraction is conducted to identify each classes using ensembles of cells. This poses a major limitation to study interconnection between lipid metabolism/protein based signaling from a global perspective and overlooks cell heterogeneity. Moreover, the spatial distributions, a vital piece for understanding biological processes is lost. It is a great technique challenge to detect all biomolecules in single cells near their nature state, and currently there is no method to directly detect metabolites in situ because of their rapid and dynamic nature and impossibility of amplifying.
The development of high resolution CCIB-SIMS in our lab has positioned us to image multiple biomolecules in cryofixed cells in a single run. The approach takes advantage of three aspects of GCIB-SIMS - low chemical damage, high yield of intact biomolecules, and the possibility of sub-micron lateral resolution. In this work, we utilize a DC beam buncher-ToF SIMS instrument to achieve high lateral resolution. Moreover, this configuration simplifies depth profiling since erosion and spectral acquisition are performed with a single beam.
To illustrate this instrumental protocol, single HeLa cells expressing purine de novo biosysthesis (PDNB) are imaged in 3D using a novel 70 keV (CO2)14000+ beam with a spot size of 1 µm. Purine de novo biosynthesis (PDNB) is essential for supporting cellular proliferation, survival and metabolic adaptation under varying nutritional environment. Using isotope tracer experiments, the stable PDNB intermediates are localized as distinct isolated punctate within cellular boundary. The simultaneous imaging of enzyme to catalyze the pathway is also developed to show the interactions of enzyme and protein. The approach provides a complete chemical picture of single cells at near original physiological and morphological state, opening the opportunities for single cell omics and heterogeneity studies using SIMS.