Huisoo Jang

Chromatic biosensor for detection of phosphinothricin acetyltransferase by use of polydiacetylene vesicles encapsulated within automatically generated immunohydrogel beads.

We developed a simple and sensitive colorimetric biosensor in the form of microparticles by using polydiacetylene (PDA) vesicles encapsulated within a hydrogel matrix for the detection of phosphinothricin acetyltransferase (PAT) protein, which is one of the most important marker proteins in genetically modified (GM) crops. Although PDA is commonly used as a sensing material due to its unique colorimetric properties, existing PDA biosensors are ineffective due to their low sensitivity as well as their lack of robustness. To overcome these disadvantages, we devised immunohydrogel beads made of anti-PAT-conjugated PDA vesicles embedded at high density within a poly(ethylene glycol) diacrylate (PEG-DA) hydrogel matrix. In addition, the construction of immunohydrogel beads was automated by use of a microfluidic device. In the immunoreaction, the sensitivity of antibody-conjugated PDA vesicles was significantly amplified, as monitored by the unaided eye. The limit of detection for target molecules reached as low as 20 nM, which is sufficiently low enough to detect target materials in GM organisms. Collectively, the results show that immunohydrogel beads constitute a promising colorimetric sensing platform for onsite testing in a number of fields, such as the food and medical industries, as well as warfare situations.

Automated formation of multicomponent-encapuslating vesosomes using continuous flow microcentrifugation

Vesosomes - hierarchical assemblies consisting of membrane-bound vesicles of various scales - are potentially powerful models of cellular compartmentalization. Current methods of vesosome fabrication are labor intensive, and offer little control over the size and uniformity of the final product. In this article, we report the development of an automated vesosome formation platform using a microfluidic device and a continuous flow microcentrifuge. In the microfluidic device, water-in-oil droplets containing nanoscale vesicles in the water phase were formed using T-junction geometry, in which a lipid monolayer is formed at the oil/water interface. These water-in-oil droplets were then immediately transferred to the continuous flow microcentrifuge. When a water-in-oil droplet passed through a second lipid monolayer formed in the continuous flow microcentrifuge, a bilayer-encapsulated vesosome was created, which contained all of the contents of the aqueous phase encapsulated within the vesosome. Encapsulation of nanoscale liposomes within the outer vesosome membrane was confirmed by fluorescence microscopy. Laser diffraction analysis showed that the vesosomes we fabricated were uniform (coefficient of variation of 0.029). The yield of the continuous flow microcentrifuge is high, with over 60% of impinging water droplets being converted to vesosomes. Our system provides a fully automatable route for the generation of vesosomes encapsulating arbitrary contents. The method employed in this work is simple and can be readily applied to a variety of systems, providing a facile platform for fabricating multicomponent carriers and model cells.

An antibacterial microfluidic system with fish gill structure for the detection of Staphylococcus via enzymatic reaction on a chromatic polydiacetylene material caused by lysostaphin

AbstractThe authors describe a microfluidic system functionalized with a chromatic nanomaterial (polydiacetylene; PDA) and conjugated to the antimicrobial enzyme lysostaphin (LST) as a means for specific detection of Staphylococcus pathogens and to simultaneously perform antimicrobial functions. The LST-loaded PDA vesicles were deposited in a fish gill-like structure on the inner surface of the microchannels. They undergo a color transition from blue to red and enhancement of fluorescence under external mechanical stimulus that is caused by the interaction between Staphylococcus and LST which has antimicrobial activity against Staphylococcus. Due to its fish gill-mimicking structure, the PDA coated channel has a high surface-to-volume ratio, and this maximizes the binding efficiency between Staphylococcus suspended in the fluid and the LST-PDA coating on the microchannels. Consequently, >80% of the Staphylococcus are eliminated in the channels within a short reaction time. As a result, the LST-PDA-coated channel surfaces undergoes color change from blue to red, and red fluorescence pops up. In contrast, no enzymatic reaction and no color transition is observed when an E. coli suspension is applied. The results show that this multifunctional microfluidic system can specifically detect, and can exert an antimicrobial effect on, Staphylococcus. Graphical abstractSchematic of a microfluidic system functionalized with a chromatic polydiacetylene (PDA) material and conjugated to the antimicrobial enzyme lysostaphin (LST) for specific detection of Staphylococcus pathogens and to simultaneously perform antimicrobial functions.