Optical tweezers development

Single molecule pulUng experiments can be described with the formalism developed in Section lll.C.l. In the simplest setting the configurational variable C corresponds to the molecular extension of the complex (handles plus inserted molecule) and the control parameter X is either the force/measured in the bead or the molecular extension of the system, x. For small enough systems the thermodynamic equation of state is dependent on what is the variable that is externally controlled [87]. In the actual experiments, the assumption that either the force or the extension is controlled is just an approximation. Neither the molecular extension nor the force can be really controlled in optical tweezers [88]. For example, in order to control the force a feedback mechanism must operate at aU times. This feedback mechanism has a time delay response so the force is never really constant [89, 90]. By assuming that the force is constant. 

Wakamoto, Y.C., Umehara, S., Matsumura, K., Inoue, I.,Yasuda, K., Development of non-destructive, non-contact single-cell based differential cell assay using on-chip microcultivation and optical tweezers. Sensors Actuators B 2003, 96, 693-700. 

Other methods have appeared more recently for measuring forces between macroscopic surfaces or between a (large) colloidal particle and a surface, immersed in a liquid. These include the total internal reflection microscope in 1990 (TIRM) and the atomic force microscope in 1991(AFM). With TIRM, incredibly weak forces can be measured (-10 N), whilst with AFM forces - 10 N can be determined. With the development of optical tweezers, we now have the ability to measure forces directly between two colloidal particles. Using these latest techniques, not only may interaction forces between surfaces be measured but, by performing dynamic measurements, the hydrodynamic forces can also be examined. We are now at a stage surely undreamt of by Theo Overbeek 50 or so years ago when he made his own measurements. It is surely fitting that he has lived to witness all this, and that he has reached an age almost commensurate with that of the Faraday Society/Division itself ... 

The optical tweezers technique has been a powerful tool for biological application. We believe that precise control of the cellular microenvironment and single-cell analysis provide opportunities to predict the effects of external stimuli including cell-cell, cell-ECM and cell-soluble factor interaction on the cell behavior and fate, which are link to revealing the internal cellular signaling system. There still exists a broad distribution of cell responses even by single-cell analysis. Researchers need to improve and develop the technique to one utilizable for a precise analysis. The... 

Observing single molecules. A major advance in the study of molecular motors has been the development of ways to observe and study single macromolecules. The methods make use of optical traps (optical "tweezers") that can hold a very small ( 1 pm diameter) polystyrene or silica bead near the waist of a laser beam focused through a microscope objective. ° ... 

New techniques are being developed that are able to measure the force-distance relationships of single molecules as they are stretched. Such techniques include atomic force microscopy and optical tweezers. Atomic force microscopy utilizes a microscale cantilever that has a probe to scan the specimen surface. Force is measured in piconewtons and the distance in nanometers. An example of some results from this technique is shown in Figure 6.5, where the force required to stretch poly(ethyleneglycol) molecules of different lengths is shown (Oesterhelt, Rief, and Gaub 1999). 

The implementation of physical mechanisms at the microscale for cell manipulation takes advantage of the scaling laws at sizes comparable to cells sizes. Some of the techniques are widely used and very well developed (see the entries on Cell Patterning on Chip, Dielectrophoretic Motion of Particles and Cells, Optical Tweezers for Manipulating Cells and Particles ). Other techniques are still emerging and our understanding of the practical aspects of these is still evolving. 

To increase throughput and eliminate sample cross contamination, which is common when analyzing cells sequentially, Munce et al. [22] developed a microfluidic device with four parallel CE channels. Calcein-labeled acute myeloid leukemia cells were selected and transported with optical tweezers to injector structures at the entrance of each channel, as shown in Fig. 3a. Once loaded, the cells were lysed by the combined action of an applied electric field and the reduction in channel cross section (Fig. 3b). The injector design also enabled stepwise lysis of the cell, shown in Fig. 3c, where only the cytoplasmic material (containing calcein AM in green) was injected into the capillary, while the nucleus (stained blue with Hoechst 33342) remained in the injector structure. This selective lysis is especially useful to separate the cytoplasm from the... 

A variety of different methods that allow one to directly study forces between few or individual molecules or functional groups relevant to supramolecular chemistry have been developed and refined in the past decades. These techniques include, among others, magnetic beads, optical tweezers, glass microneedles, the biomembrane force probe," force clamp, and AFM approaches. For a recent overview, the reader is referred to the review by Clausen-Schaumann et al, who summarized and compared these prominent methods in detail. As shown in Table these techniques enable the measurement of forces that range from weak entropic forces at several piconewtons to the rupture of covalent bonds at a few nanonewtons covering a dynamic range from microseconds to seconds. 

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