Sample manipulation is one of the critical issues in synchrotron radiation microbeam experiments and in particular for experiments in special environments such as microfluidic cells. A common requirement is to position and move the sample in front of the beam. This is usually accomplished by fixing the sample on a support (e.g. capillary tip, grid), which is then moved precisely into the beam (Figure 57a). Consequently, this limits the degrees of freedom for sample orientation and manipulation and does not allow interactions between two independent objects to be studied in terms of mutual orientation, mutual growth (single cell study, real time crystal growth) [1].

Fig. 57: Sample manipulation in X-ray microdiffraction experiments: a) Mechanical contact/trap and b) Laser trap without mechanical contact. c) Microscope image of a cluster of liposomes trapped optically (dash line) inside a capillary (the walls of the capillary are seen in black on the lateral sides), scale bar 10 µm. d) Plot of the azimuthally integrated intensity (red) and the Lorentz function (black) fitted to experimental data for the diffraction pattern (inset) at one position of the X-ray beam on an optically trapped POPE cluster.

One solution, laser tweezers have emerged as a powerful tool to manipulate a large variety of microparticles immersed in fluids [2]. We have now built a laser tweezer setup based on a custom inverted microscope and an infrared laser to trap, manipulate and aggregate micrometre-scale liposome particles inside a 100 micrometre glass capillary [3]. The micro-focused synchrotron radiation and laser beams are aligned to intersect each other, allowing the X-ray diffraction signal to be associated with the micrometre-sized region of interest inside the capillary (Figure 57b). This setup has been demonstrated at the ID13 microfocus beamline by trapping about 50 multilamelar palmitoyl-oleoyl-phosphatidylethanolamine (POPE) liposome clusters of about 10 µm diameter (Figure 57c) and performing a scanning diffraction experiment with a 1 µm beam at about 13 keV. The azimuthally integrated intensity of the diffraction pattern obtained from the liposome cluster at one position of the X-ray beam is shown in Figure 57d. Multiple laser traps can be created using diffractive optical elements implemented on a spatial light modulator [4]. This allows manipulation of small separated clusters of liposomes and their fixation within the optical path of the X-ray beam. Using two traps, two different clusters could be brought into contact and, consequently, a reaction between them could be induced. The proof of concept is illustrated by Figure 58. Multiple traps can be also used to trap and manipulate larger particles such as starch granules, as shown in Figure 58c. We have improved the initial laser tweezers setup, allowing 2D optical trapping in a cylindrical capillary (Figure 58a), to a setup with full 3D optical trapping in a 80 µm squared capillary (Figure 58c). This enables positioning and orienting the sample in any point of the capillary [5]. The scanning diffraction image taken from a 20 x 24 µm starch granule is illustrated in Figure 58d together with refraction from the wall of the capillary thus demonstrating the trapping capability far away from the capillary wall.

Two clusters of liposomes trapped at separate positions

Fig. 58: Two clusters of liposomes trapped at separate positions, indicated by red and blue lines: a) microscope image of the traps (infrared laser) and b) X-ray scanning diffraction image (azimuthaly integrated intensities, mesh 3 µm x 5 µm) from the two clusters, with the corresponding intensity profiles. c) Microscope image of a large starch granule trapped with three laser traps (red spots) and d) scanning diffraction image (diffraction patterns) of the starch ({001}-9 nm).

The results presented here have given a first insight into this fascinating new field - to look to single supra-molecular assemblies with micrometre- and submicrometre-sized X-ray beams, which will find multiple applications in third- and in particular future fourth-generation synchrotron radiation sources.



D. Cojoc (a), H. Amenitsch (b), C. Riekel (c), E. Ferrari (a), M. Rappolt (b), M. Burghammer (c), S. Santucci (a), G. Grenci (a), B. Sartori (b), B. Marmiroli (b).
(a) TASC National Laboratory, Trieste (Italy)
(b) Institute of Biophysics and Nanosystems Research, Austrian Academy of Sciences, Graz (Austria)
(c) ESRF


[1] C. Riekel et al., Curr. Opin. Struct. Biol. 63 556 (2005).
[2] D.G. Grier, Nature 424 810 (2003).
[3] H. Amenitsch et al., AIP Conf. Proc. 879 1287 (2006).
[4] D. Cojoc et al., Appl.Phys. Lett. 91 234107 (2007).
[5] D. Cojoc et al. Nat. Meth. submitted (2008).