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In nature, biological compartments such as cells rely on dynamically controlled permeability for matter exchange and complex cellular activities. Likewise, the ability to engineer compartment permeability is crucial for in vitro systems to gain sustainability, robustness, and complexity. However, rendering in vitro compartments such a capability is challenging. Here, a facile strategy is presented to build permeability-configurable compartments, and marked advantages of such compartmentalization are shown in reconstituting sustained synthetic biology systems in vitro. Through microfluidics, the strategy produces micrometer-sized layered microgels whose shell layer serves as a sieving structure for biomolecules and particles. In this configuration, the transport of DNAs, proteins, and bacteriophages across the compartments can be controlled an guided by a physical model. Through permeability engineering, a compartmentalized cell-free protein synthesis system sustains multicycle protein production; ≈100 000 compartments are repeatedly used in a five-cycle synthesis, featuring a yield of 2.2 mg mL-1 . Further, the engineered bacteria-enclosing compartments possess near-perfect phage resistance and enhanced environmental fitness. In a complex river silt environment, compartmentalized whole-cell biosensors show maintained activity throughout the 32 h pollutant monitoring. It is anticipated that permeability-engineered compartmentalization should pave the way for practical synthetic biology applications such as green bioproduction, environmental sensing, and bacteria-based therapeutics.

Microfluidic devices have been increasingly used for low-volume liquid handling operations. However, laboratory automation of such delicate devices has lagged behind due to the lack of world-to-chip (macro-to-micro) interfaces. In this paper, we have presented the first pipette-free robotic-microfluidic interface using a microfluidic-embedded container cap, referred to as a microfluidic cap-to-dispense (μCD), to achieve a seamless integration of liquid handling and robotic automation without any traditional pipetting steps. The μCD liquid handling platform offers a generic and modular way to connect the robotic device to standard liquid containers. It utilizes the high accuracy and high flexibility of the robotic system to recognize, capture and position; and then using microfluidic adaptive printing it can achieve high-precision on-demand volume distribution. With its modular connectivity, nanoliter processability, high adaptability, and multitask capacity, μCD shows great potential as a generic robotic-microfluidic interface for complete pipette-free liquid handling automation.

Circulating tumor cells are typically found in the peripheral blood of patients, offering a crucial pathway for the early diagnosis and prediction of cancer. Traditional methods for early cancer diagnosis are inefficient and inaccurate, making it difficult to isolate tumor cells from a large number of cells. In this paper, a new spiral microfluidic chip with asymmetric cross-section is proposed for rapid, high-throughput, label-free enrichment of CTCs in peripheral blood. A mold of the desired flow channel structure was prepared and inverted to make a trapezoidal cross-section using a micro-nanotechnology process of 3D printing. After a systematic study of how flow rate, channel width, and particle concentration affect the performance of the device, we utilized the device to simulate cell sorting of 6 μm, 15 μm, and 25 μm PS (Polystyrene) particles, and the separation efficiency and separation purity of 25 μm PS particles reached 98.3% and 96.4%. On this basis, we realize the enrichment of a large number of CTCs in diluted whole blood (5 mL). The results show that the separation efficiency of A549 was 88.9% and the separation purity was 96.4% at a high throughput of 1400 μL/min. In conclusion, we believe that the developed method is relevant for efficient recovery from whole blood and beneficial for future automated clinical analysis. [ABSTRACT FROM AUTHOR]

X-ray structures of the bacterial [K.sup.+] channel KcsA have led to unparalleled progress in our understanding of ion channel structures. The KcsA channel has therefore been a prototypic model used to study the structural basis of ion channel function, including the gating mechanism. This channel was previously found to close at near-neutral intracellular pH ([pH.sub.i]) and to open at acidic [pH.sub.i]. Here, we report the presence of a previously unknown channel inactivation process that occurs after the KcsA channel is activated. In our experiments, mammalian cells transfected with a codon-optimized synthetic gene encoding the KcsA protein expressed [K.sup.+]-selective channels that activated in response to a decrease in [pH.sub.i]. Using patch-clamp and rapid solution exchange techniques, we observed that the KcsA channels inactivated within hundreds of milliseconds after channel activation. At all tested pHs, inactivation always accompanied activation, and it was profoundly accelerated in the same pH range at which activation increased steeply. Recovery from inactivation was observed, and its extent depended on the pHi and the amount of time that the channel was inactive. KcsA channel inactivation can be described by a kinetic model in which pHi controls inactivation through pH-dependent activation. This heretofore-undocumented inactivation process increases the complexity of KcsA channel function, but it also offers a potential model for studying the structural correspondence of ion channel inactivation. gating | ion channel inactivation recovery from inactivation | synthetic gene | model

In mildly hyperosmotic medium, activation of the [Na.sup.+][sub.-.K.sup.+]-2[Cl.sup.-] cotransporter (NKCC) counteracts skeletal muscle cell water loss, and compounds that stimulate protein kinase A (PKA) activity inhibit the activation of the NKCC. The aim of this study was to determine the mechanism for PKA inhibition of NKCC activity in resting skeletal muscle. Incubation of rat slow-twitch soleus and fast-twitch plantaris muscles in isosmotic medium with the PKA inhibitors H-89 and KT-5720 caused activation of the NKCC only in the soleus muscle. NKCC activation caused by PKA inhibition was insensitive to MEK MAPK inhibitors and to insulin but was abolished by the PKA stimulators isoproterenol and forskolin. Furthermore, pinacidil [an ATP-sensitive potassium ([K.sub.ATP]) channel opener] or inhibition of glycolysis increased NKCC activity in the soleus muscle but not in the plantaris muscle. Preincubation of the soleus muscle with glibenclamide (a [K.sub.ATP] channel inhibitor) prevented the NKCC activation by hyperosmolarity, PKA inhibition, pinacidil, and glycolysis inhibitors. In contrast, glibenclamide stimulated NKCC activity in the plantaris muscle. In cells stably transfected with the Kir6.2 subunit of the of [K.sub.ATP] channel, inhibition of glycolysis activated potassium current and NKCC activity. We conclude that activation of [K.sub.ATP] channels in slow-twitch muscle is necessary for activation of the NKCC and cell volume restoration in hyperosmotic conditions. protein kinase A; glibenclamide; glycolysis; [Na.sup.+][sub.-.K.sup.+]-[2Cl.sup.-] cotransporter; Kir6.2