Thesis-PDF - IAP/TU Wien

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Figure 5.8: A small centrifuge was built (with the help of the supportive staff of the mechanical workshop at the <strong>TU</strong>-<strong>Wien</strong>) for the condensing of cell parts in the solution for the AFM. Here it is seen equipped with a vial of growth medium containing living algal cells (left) and a counterweight vial filled with water to the right. Figure 5.9: A schematic drawing of the imaging setup under liquid. The plastic petridish (a) is attached to the scanning stage (b) via double-sided scotch tape or plasticine (c). At the bottom inside the petridish, a piece of glass slide (d) is attached via double-sided scotch tape. Liquid medium (e) fills the petridish so that the cantilever (h) as well as the laser travel line (g) leaving the AFM head (f - only the lower part of the head is drawn) are fully immersed in liquid when scanning over the surface. A petridish measured about 6 cm in diameter. 76
Chapter 6 Results High resolution topographic images of E. gracilis acquired with the Atomic Force Microscope are presented in this chapter. Algal features found are related to Euglenoid morphology as presented in chapter 4. Three major goals were reached during the work on E. gracilis. First, a preparation method suitable for AFM investigation of E. gracilis had to be developed. This was found in a special drying process (see section 5.3) that left whole intact algae to be investigated. Second, AFM data of unprecedented resolution of E. gracilis were acquired, on the algal pellicle and even on certain cell parts. A third goal was reached in providing additional AFM data to what is already available from SEM and TEM investigations. Investigated features of the alga included e.g. imaging of the mucilage in between pellicle strips, a feature usually not visible through SEM or TEM due to sample preparation techniques that remove soft biological material. Our AFM investigation of E. gracilis also revealed new features that have not been described before. 6.1 Sample Preparation - Challenges and Solutions The preparation method as presented in 5.3 solved a number of difficulties encountered with earlier attempts to image the alga. Firstly, the algal cells burst during the drying process if no precautions are taken. Fig. 6.1 shows such a burst E. gracilis cell. Obviously, this hinders imaging of intact cell surfaces and in addition enzymatic activity might damage biological membranes, alter their properties or slowly dissolve them altogether (autolysis). Although certain cell features still can be made out this information is unreliable. A further challenge in the invented drying process had been the amount of residue of the evaporated nutrient solution. As can be seen in Fig. 6.2, the embedding residue does not only prevent cells from bursting but can also prevent 77
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DIPLOMARBEIT Atomic Force Microscop
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»Later I have been asked many time
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Contents Abstract 4 1 Introduction
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5.2.1 Optical Microscopy and Fluore
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theory (see e.g. [1]). Now I call o
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understand phenomena at a larger sc
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of whole dried Euglena cells in air
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these assembly functions for us. 2
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Figure 2.2: Rise in public spending
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The presented three types of nanote
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2.4.1 Self-Assembly and Emergent St
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After diligent investigation such s
Page 25 and 26: 1 µN) between a probe and the samp
Page 27 and 28: 3.2.1 Static Mode In Static Mode (a
Page 29 and 30: crystal and the cantilever tip (pha
Page 31 and 32: Figure 3.6: Highly ordered and fres
Page 33 and 34: Figure 3.8: Schematic drawing of th
Page 35 and 36: Figure 3.11: The cantilever deflect
Page 37 and 38: Figure 3.14: This force curve shows
Page 39 and 40: Figure 4.1: A group of Euglena orga
Page 41 and 42: several species with rigid pellicle
Page 43 and 44: Figure 4.2: The rightmost cell can
Page 45 and 46: cavity and is used for regulatory f
Page 47 and 48: as phobic reactions, because the ce
Page 49 and 50: Figure 4.7: Scheme of the photorece
Page 51 and 52: Figure 4.9: Measured monoclinic uni
Page 53 and 54: Figure 4.11: Twice the same Euglena
Page 55 and 56: 4.15), their ends are termed the (-
Page 57 and 58: Figure 4.15: Kinesin and dynein and
Page 59 and 60: Figure 4.17: (Image reproduced from
Page 61 and 62: into fully functional chloroplasts
Page 63 and 64: Figure 4.19: How a microtubule-kine
Page 65 and 66: Chapter 5 Materials and Methods The
Page 67 and 68: Cell Treatment - Demembranation In
Page 69 and 70: Figure 5.3: This image shows cells
Page 71 and 72: Figure 5.4: The AFM setup at the In
Page 73 and 74: Figure 5.6: If the user menu AFM_TU
Page 75: Figure 5.7: Single steps of the sam
Page 79 and 80: Figure 6.2: An embedded E. gracilis
Page 81 and 82: Figure 6.5: Scanning under liquid s
Page 83 and 84: Flagellum The flagellum of an E. gr
Page 85 and 86: Pellicle The pellicle of E. gracili
Page 87 and 88: Figure 6.11: The pellicle of an E.
Page 89 and 90: Crystalline Cell Parts Crystalline
Page 91 and 92: Figure 6.18: A layered structure th
Page 93 and 94: It might also be possible that comb
Page 95 and 96: Appendix A Acronyms SPM Scanning Pr
Page 97 and 98: [13] A. Junk, F. Riess: From an ide
Page 99 and 100: [45] J. Ruan, B. Bhushan: Atomic-sc
Page 101 and 102: [70] M. Rief, M. Gautel, F. Oesterh
Page 103 and 104: [97] P. Gualtieri, L. Barsanti: Alg
Page 105 and 106: Annex 1 This Igor script was used i
Page 107 and 108: w[i][j][laynr]=(w[i][j][3]-used_off
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