Fast Mixing and Reaction Initiation Control of Single-Enzyme Kinetics in Confined Volumes

Abstract

Supporting InformationPhotolithography PDMS molds were fabricated with multilayer photolithographic techniques. Briefly, negative tone photomasks for the control and mixer layers were printed on transparency films at 20,000 dots per inch resolution by CAD/Art Services (Bandon, OR). A chrome mask with arrays of 5 μm dots was purchased from MEMS and Nanotechnology Exchange (Reston, VA). To make the mold for the mixer/chamber layer, SU8 2005 was first spun at 2000 rpm on a piece of isopropanol-cleaned silicon. After soft baking(1 min at 65°C and 2 min at 95°C), the 5 μm dot arrays as well as alignment marks were transferred from the chrome mask to the resist by UV exposure at 4.5 mW/cm 2 for 90 sec, followed by postexposure baking (1 min at 65°C and 1 min at 95°C), and development in SU8 developer. 5 μm dot patterns were further stabilized by hard baking at 150°C for 5 min. Another layer of 25 μm thick SU8 2025 photoresist was then spin-coated on top of the dot pattern and soft baked (1 min at 65°C and 3 min at 95°C). Each 5 μm chamber had a volume of approximately 100 femtoliters (4.4 μm diameter with 6.5 μm height), as inferred from scanning electron microscopy (SEM) images of the microchambers.To align the staggered herringbone mixer patterns relative to the array of dots, four corners of the silicon chip were carefully cleared with SU8 developer to reveal the alignment marks on the mold. The mixer patterns were then transferred to the second resist layer through the transparency mask by UV

Shear-Driven Redistribution of Surfactant Affects Enzyme Activity in Well-Mixed Femtoliter Droplets

“…Figures S8A, B are fluorescence plots of 100 M RGP substrate alone, and 100 M substrate with 200 nM resorufin, contained in 5 m diameter chambers (~100 fL) formed by micromolding in PDMS, 4 under the same illumination conditions as the fL droplet experiments carried out in the microfluidic devices as described in the paper. If there was autohydrolysis of RGP due to reaction with singlet oxygen species generated from the photobleaching of resorufin, we would expect to see significant differences in the relative increases in fluorescence in S8B versus S8A over the time course of five minutes.…”

Experimental observations of flow instabilities and rapid mixing of two dissimilar viscoelastic liquids

“…2,29 Furthermore, the unique advantage of this mixing technology is that the viscoelastic properties responsible for elastic flow instabilities could be readily found in most chemical and biological samples like organic and polymeric solutions, [23][24][25] blood 28 and crowded proteins and enzymes samples. 26,27 In this paper, we used the same microdevice previously reported 2,3 to experimentally show that the viscoelasticity ratio (between mainstream and sidestream liquids) has a direct bearing on the magnitude of the induced elastic flow instabilities, and in particular the transverse flow instability at downstream of a contraction. The possible underlying mechanism is discussed to elucidate the complex interplay between elasticity/viscosity and the elasticity ratio between the mainstream and side-streams solutions.…”

“…21,22 In polymer and chemical syntheses in micro-reactors, [23][24][25] dissimilar viscosities could significantly influence the reaction rate. Other applications like rapid mixing of crowded biological solutions 26,27 (e.g. highly concentrated protein or enzyme solutions) and injecting of reagent into whole blood 28 also require the mentioned domain knowledge to achieve a precise incubation period and rapid-mixed samples.…”

Recent developments in microfluidic large scale integration

“…Improved designs for fundamental components include normally closed valves (NCVs) [16 ,17], structured valves [18], cell traps [19], and capacitors and diodes ( Figure 1a) [20,21]; higher-level components that have been introduced include a chamber array [22], a high performance separation column [23 ], a gradient selector (Figure 1b) [23 ], a long term gradient generator [24 ], a 2D spatial gradient controller [25], an agar filled chamber [26] and a mutilaminate mixer [27].…”