Development of a micromixing system for treatment of ORL 48 microtissues in different concentrations of cytochalasin B

Mixing and dilution are essential procedures in pharmaceutical operation to process two or more components in a separate or thoroughly mixed condition until homogenous solution was obtained. However, conventional serial dilution method used in laboratory assessment causes high usage of reagents, higher complexity procedures and costly. Micromixing method provides a better platform that enables mixing and dilution of liquid-based reagents which is convenient solutions preparation, easy liquid handling and time-saving. In this study, a polydimethylsiloxane (PDMS) micromixer was designed, simulated and prototyped using vinyl tape method and successfully applied to mix and dilute Cytochalasin-B in culture media (CB-DMEM, 30.0 μM) with 0.05 % ethanol solutions (diluent) to produce four different concentrations of CB-DMEM (5.3, 10.6, 14.8, and 20.2 μM). The different concentrations of CB-DMEM were applied on to ORL-48 microtissues produced by using flicking technique. The morphological responses, cell viability and cell proliferation of ORL-48 monolayer cells (2D) and microtissues (3D) treated in four different CB concentrations were assessed via phase contrast microscopy, live/dead staining and Alamar Blue® staining respectively. The results show that both 2D and 3D of ORL-48 microtissues were only morphologically affected (fibroblastic spreading to round shape) while cell viability and cell proliferation show that CB treatment solely does not causes apoptosis (≈ 90 % cells are alive and able to proliferate). The micromixer employed in solution preparation of CB-DMEM (5.3, 10.6, 14.8, and 20.2 μM) provide a convenient and faster method to prepare cytochemical solution for drug screening and experiments. Besides that, application of micromixer consumes less volume of reagents and cost efficient

Development of a twin-head infusion pump for micromixing

Mixing is a crucial process in most of the industrial technology such as the operation of chemicals and fermentation reactors, combustion engines, polymer blends, and pharmaceutical formulations [1]. For handling a smaller volume of liquid, micromixing is a suitable method that can be applied. Micromixing (micromixer) is one of the microfluidic functions for mixing and blending liquids as precursors for biological process such as cell activation, enzyme reaction, and drug delivery system [2, 3]. There are several advantages of applying microfluidic device (micromixer) in the chemical technological processes such as processing accuracy, efficiency, minimum usage of reagents and ease of disposing of devices and fluids [3]. Basically, micromixers are categorised into passive and active micromixers. Passive micromixer consists of no moving parts and free from additional friction. It does not use external forces, fully dependent on molecular diffusion and chaotic advection for mixing process [4]. In contrast to active micromixers, external forces are applicable to active micromixers by implementing moving elements either within the microchannels, a time-variant, or a pressure field [5]. To create the pressure field differences for moving the liquid within the micromixer, an infusion pump is usually applied

Micromixers and applications

Mixing and dilutions are processes to combine or putting together one or more reagents such as biomolecules, enzyme, proteins and emulsions into the desired concentrations for further applications in chemical and biological applications. The mixing and diluting of chemical solutions in the laboratory require the use of large quantity of plasticwares, consume time and involve with laborious procedures. Micromixing based on microfluidic is a new innovative means to micromix reagents in microliters volume. The reactions of mixing in both active and passive micromixers are dependent on the mass transport phenomena, viscosity of fluid, molecular diffusion and convection. Reynold number indicates if the fluid flow through a channel is steady or turbulent, while Péclet number indicates the magnitude order between convective and diffusive transport. The success of micromixing is also greatly influenced by the velocity, concentration and pressure of fluids. Different methods for fabricating microfluidic device that include etching, thermoforming, polymer ablation, casting and soft-lithography will be reviewed. In addition, the applications of different designs of micromixers in fluid and particles mixing are illustrated in this chapter

Comparison of biophysical properties characterized for microtissues cultured using microencapsulation and liquid crystal based 3D cell culture techniques

Growing three dimensional (3D) cells is an emerging research in tissue engineering. Biophysical properties of the 3D cells regulate the cells growth, drug diffusion dynamics and gene expressions. Scaffold based or scaffoldless techniques for 3D cell cultures are rarely being compared in terms of the physical features of the microtissues produced. The biophysical properties of the microtissues cultured using scaffold based microencapsulation by flicking and scaffoldless liquid crystal (LC) based techniques were characterized. Flicking technique produced high yield and highly reproducible microtissues of keratinocyte cell lines in alginate microcapsules at approximately 350 ± 12 pieces per culture. However, microtissues grown on the LC substrates yielded at lower quantity of 58 ± 21 pieces per culture. The sizes of the microtissues produced using alginate microcapsules and LC substrates were 250 ± 25 μm and 141 ± 70 μm, respectively. In both techniques, cells remodeled into microtissues via different growth phases and showed good integrity of cells in field-emission scanning microscopy (FE-SEM). Microencapsulation packed the cells in alginate scaffolds of polysaccharides with limited spaces for motility. Whereas, LC substrates allowed the cells to migrate and self-stacking into multilayered structures as revealed by the nuclei stainings. The cells cultured using both techniques were found viable based on the live and dead cell stainings. Stained histological sections showed that both techniques produced cell models that closely replicate the intrinsic physiological conditions. Alginate microcapsulation and LC based techniques produced microtissues containing similar bio-macromolecules but they did not alter the main absorption bands of microtissues as revealed by the Fourier transform infrared spectroscopy. Cell growth, structural organization, morphology and surface structures for 3D microtissues cultured using both techniques appeared to be different and might be suitable for different applications