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on the glass slide. In order to achieve sufficient attachment of the material onto the glass surface the material was given time to adsorb to the surface (incubation time). Depending on preparation type and liquid, this time varied from seconds to over an hour. Poly-l-lysine coated slides were sometimes used in this setup to improve adhesion of the algae to the slides (see Fig. 5.9). 5.3.4 Experimental Preparations For experimental purposes other types of preparations were tried out. It turned out that the most effective sample preparation method is the one given in 5.3.2. Experimental preparations include prior calcium treatment of cells for ejection of the reservoir system and the flagellar apparatus. Through calcium diffusion internal cell pressure was raised until ejection. With the ejection of the flagellar apparatus the paraflagellar body (crystalline photoreceptor) attached to the longer flagellum becomes accessible. This procedure proved viable for good results with the optical microscope. Unfortunately, investigation with fluorescence microscopy showed that the fluorescence characteristics of the photoreceptor had deteriorated. The three-dimensional integrity of the photoreceptor was no longer guaranteed and therefore no additional investigation of these samples under the AFM was carried out. Another experimental procedure tried was the chitosan matrix procedure. Fixated, demembranated cells were included in a chitosan matrix (its main component is chitin). Subsequent washing with alcohol, and freeze fracturing gave surfaces that were possible to scan with scanning probe techniques. Although this approach yielded some images on cell surface features, it is a time consuming process and the data obtained by ambient AFM investigation of dried specimen was clearly of better quality. 74
Figure 5.7: Single steps of the sample preparation. A drop of liquid containing cells is applied to the glass slide and covered by a coverslip (1). The evaporation starts at the edges around the coverslip (2 - detail). The coverslip is removed through gentle dragging after about 5 minutes to allow for complete drying (3). Although cells usually burst during the drying process (4 - detail), some areas where the edges of the coverslip have been (designated in 3) still contain embedded intact Euglena cells. 75
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DIPLOMARBEIT Atomic Force Microscop
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»Later I have been asked many time
Page 5 and 6:
Contents Abstract 4 1 Introduction
Page 7 and 8:
5.2.1 Optical Microscopy and Fluore
Page 9 and 10:
theory (see e.g. [1]). Now I call o
Page 11 and 12:
understand phenomena at a larger sc
Page 13 and 14:
of whole dried Euglena cells in air
Page 15 and 16:
these assembly functions for us. 2
Page 17 and 18:
Figure 2.2: Rise in public spending
Page 19 and 20:
The presented three types of nanote
Page 21 and 22:
2.4.1 Self-Assembly and Emergent St
Page 23 and 24: 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: Figure 5.6: If the user menu AFM_TU
Page 77 and 78: Chapter 6 Results High resolution t
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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