Mechanical forces have been observed to affect various aspects of life, for example, cell differentiation, cell migration, locomotion and behavior of multicellular organisms etc. Such forces are generated either by external entities such as mechanical touch, fluid flow, electric and magnetic fields or by the living organisms themselves. Study of forces sensed and applied by living organisms is important to understand the interactions between organisms and their environment. Such studies may reveal molecular mechanisms involved in mechanosensation and locomotion. Several techniques have been successfully applied to measure forces exerted by single cells and cell monolayers. The earliest technique made use of functionalized soft surfaces and membranes as substrates on which cell monolayers were grown. The forces exerted by the cells could be measured by observing deformation of the substrates. Atomic Force Microscope (AFM) is another sensitive instrument that allows one to exert and measure forces in pico-Newton range. Advances in micromachining technology have enabled development of miniature force sensors and actuators. Latest techniques for mechanical force application and measurement use micromachined Silicon cantilevers in single as well as array form and micropillar arrays. Micropillar arrays fabricated using soft lithography enabled the use of biocompatible materials for force sensors. Together, these techniques provide access to a wide range of forces, from sub micro-Newton to milli-Newton. In the present work, types of forces experienced in biological systems and various force measurement and actuation techniques will be introduced. This will be followed by in depth description of the two major contributions of this thesis, 1) ―Colored polydimethylsiloxane micropillar arrays for high throughput measurements of forces applied by genetic model organisms‖. Biomicrofluidics, January 29, 2015. doi: 10.1063/1.4906905 2) ―Air microjet system for non-contact force application and the actuation of micro-structures‖. Journal of micromechanics and microengineering, December 15, 2015. doi: 10.1088/0960-1317/26/1/017001 Device developed for force measurement consists of an array of micropillars made of a biocompatible polymer Poly Dimethyl Siloxane (PDMS). Such devices have been used by researchers to measure traction forces exerted by single cells and also by nematode worm Caenorhabditis elegans (C. elegans). C. elegans is allowed to move in between the micropillars and the locomotion is video recorded. Deflection of the micropillar tips as the worm moves is converted into force exerted. Transparent appearance of C. elegans and PDMS poses difficulties in distinguishing micropillars from the worm, thus making it challenging to automate the analysis process. We address this problem by developing a technique to color the micropillars selectively. This enabled us to develop a semi-automated graphical user interface (GUI) for high throughput data extraction and analysis, reducing the analysis time for each worm to minutes. Moreover, increased contrast because of the color also delivered better images. Addition of color changed the Young‘s modulus of PDMS. Thus the dye-PDMS composite was characterized using hyper-elastic model. The micropillars were also calibrated using commercial force sensor. Analysis of forces exerted by wild type and mutant C. elegans moving on an agarose surface was performed. Wild type C. elegans exerted a total average force of 7.68 µN and an average force of ~1 µN on an individual pillar. We show that the middle of C. elegans exerts more force than its extremities. We find that C. elegans mutants with defective body wall muscles apply significantly lower force on individual pillars, while mutants defective in sensing externally applied mechanical forces still apply the same average force per pillar compared to wild type animals. Average forces applied per pillar are independent of the length, diameter, or cuticle stiffness of the animal. It was also observed that the motility of the worms with mechanosensation defects, lower cuticle stiffness, and body wall muscle defects was reduced with worms that have defective body wall muscle having the largest degree. Thus, we conclude that while reduced ability to apply forces affects the locomotion of the worm in the micropillar array, the reduced motility/locomotion may not indicate that the worm has reduced ability to apply forces on the micropillars. We also used the colored micropillar array for the first time to measure forces exerted by Drosophila larvae. Our device successfully captured the peristaltic rhythm of the body wall muscles of the larva and allowed us to measure the forces applied on each deflected pillar during this motion. Average force exerted by 1st instar wild type Drosophila larvae was measured to be ~ 1.5 µN per pillar. We demonstrated that a microjet of air can be used to apply forces in micro-Newton range. We developed a standalone system to generate a controlled air microjet. Microjet was generated using a controlled electromagnetic actuation of a diaphragm. With a nozzle diameter of 150 µm, the microjet diameter was maintained to a maximum of 1 mm at a distance of 5 mm from the nozzle. The force generated by the microjet was measured using a commercial force sensor to determine the velocity profile of the jet. Axial flow velocities of up to 25 m/s were obtained at distances as long as 6 mm. The microjet exerted a force up to 1 µN on a poly dimethyl siloxane (PDMS) micropillar (50 µm in diameter, 157 µm in height) and 415 µN on a PDMS membrane (3 mm in diameter, 28 µm thick). We also demonstrate that from a distance of 6 mm our microjet can exert a peak pressure of 187 Pa with a total force of about 84 µN on a flat surface with 8 V operating voltage. Next, we demonstrated that the response of C. elegans worms to the impinging air microjet is similar to the response evoked using a manual gentle touch. This contactless actuation tool avoids contamination and mechanical damage to the samples. Out of the cleanroom fabrication and robust design make this system cost effective and durable. Magnetic micropillars have been used as actuators. We fabricated magnetic micropillar arrays and designed actuation mechanisms using permanent magnet and a pulsed electromagnet. Force of about 19 µN was achievable using a permanent magnet actuation. In a pulsed electromagnetic field micropillar exerted a force of about 10 µN on a commercial force sensor. These techniques have promising applications when actuation needs to be controlled from long distances.

L’utilisation de chromophores absorbant à deux photons (ADP) pour des applications en photothérapie dynamique (PDT) et en imagerie de fluorescence présente de nombreux avantages. Les propriétés non-linéaires de ces chromophores permettent notamment d’améliorer la longueur de pénétration dans les organismes vivants ainsi que la résolution. Pour des applications in-vivo la biocompatibilité de ces chromophores lipophiles doit aussi se poser. Une étude d’ingénierie pour le développement de chromophores pour la PDT-ADP en utilisant des atomes de brome comme groupe générateur d’oxygène singulet est décrite. Différents paramètres dont le nombre et la position des atomes de brome sur la chaîne carbonée, la longueur de conjugaison, la géométrie des chromophores ont été étudiés. Cette étude permet de mettre en évidence l’importance de la position des substituants bromes et de la symétrie sur le rendement de croisement inter-système.Les observations spectroscopiques faites lors de l’étude d’ingénierie ont permis de développer des chromophores pour la microscopie de fluorescence à deux photons. La biocompatibilité est apportée grâce à un polymère d’(hydroxyethyl)acrylate. Ce polymère permet de créer une coquille hydrosolubilisante covalente. Ces chromophores ont été utilisés pour faire de l’imagerie de vascularisation cérébrale de haute résolution. Une observation particulière sur un chromophore, marquage des cellules endothéliales des parois des vaisseaux sanguins intravitaux ainsi que les applications en résultant sont présentées. Des stratégies visant l’amélioration de la sélectivité des systèmes polymères/chromophores pour des applications intravitales, comme le traitement des tumeurs cancéreuses sont décrites. Une stratégie de modification des fonctions hydroxy des chaînes polymères par des groupements imidazoliums est présentée. L’étude de complexation des polymères avec l’ADN et les études spectroscopiques in-cellulo ont été réalisées. / The use of two-photon absorbing (TPA) chromophore for applications in photodynamic therapy (PDT) and fluorescence imaging provides many advantages. The non-linear properties make it possible to increase both observation depth in animals and 3D resolution. Nevertheless, for in-vivo applications, improving bio-compatibility of these inherently lipophilic chromophore is a challenge. The development of new chromophores for PDT-TPA using a molecular engineering approach using bromide substituents as singlet oxygen generators is described. Parameters like position and number of bromide, the conjugated length and chromophore symmetry are studied. The study shows the importance of bromide atom position and of the symmetry on the inter system crossing efficiency. During the engineering study, spectroscopic observation and rationalization permit to envision the design of new chromophores for two photon laser scanning fluorescent microscopy. Bio-compatibility of these chromophores is provided by (hydroxyethyl)acrylate polymer, which provides a covalent water-soluble shell. These chromophores are used to make high resolution image of cerebral vascularization. One of these chromophores shows intravital specific interaction with endothelial cells in blood vessels. Some applications of the chromophore are described. Strategies to increase the intravital selectivity of polymer/chromophores units towards cancer cells and tumor are presented. A modification of hydroxyl function by imidazolium group is described. This new chromophore is evaluated towards its complexation properties with DNA and in cellulo spectroscopic studies.

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