全自动四通道脂双层工作站

Orbit mini - 脂双层工作站

Orbit mini是一款小型的脂双层工作站，可同时记录四个人工脂双层。

Orbit mini的主要特点有：

• 到手可用、占地极小的独立系统
• 内置四通道低噪音放大器
• 可选的温度控制附件 - 自动制冷与加热
• 低噪音高带宽记录
• 通量提高到四通道
• 附带专用记录软件
• 目标蛋白可直接插入和通过蛋白脂质体插入
• 可记录: 电压、配体门控离子通道、温度敏感离子通道
• 也可记录纳米孔、抗菌肽、毒素等...
• 低实验成本的MECA芯片

Orbit mini包含一个内置的小型化四通道放大器，可在高带宽下进行低噪音记录而不需要其他设备辅助 – 可以在任何环境下进行实验。此外独特的设计使得测量室的温度可以自由控制而不会导入额外的噪音。

完整的Orbit mini平台包含内置四通道放大器(Elements s.r.l.)的记录单元和选配的可制热和制冷的温度控制单元。并使用Ionera公司的 MECA 4记录芯片涂抹磷脂并同时进行四个脂双层记录。.
温度控制单元
使用温度控制单元，芯片上的温度可以在0°C到50°C之间变化，因为它包含主动冷却和主动加热功能。

Orbit mini:PEG诱导的α溶血素孔电流阻断的温度控制
（A） 在10°C和40°C下记录的电流轨迹表明，在高温下，开孔电流和堵塞频率显著增加。（B） 阻塞期间剩余电流的事件平均直方图。开孔电流随堵塞物的温度和停留时间而变化。（C） 开孔电流与温度的关系。（D） 堵塞频率与温度的关系。
Orbit mini EDR 软件


MECA 4 记录芯片
MECA记录基板在高惰性聚合物中包含2 x 2圆形微腔阵列。每个腔体包含一个单独的集成Ag/AgCl微电极。双分子层是通过涂漆形成的，功能性双分子层的成功率很高。MECA芯片已通过多种不同的离子通道验证，包括KcsA、gramicidin、α-溶血素、KV1.3、NaV等。

Available chip types

• "Meca 4 Recording Chips 50 µm": Four-well recording chip with 50 µm cavity size (Order # 132001)
• "Meca 4 Recording Chips 100 µm": Four-well recording chip with 100 µm cavity size (Order # 132002)
• "Meca 4 Recording Chips 150 µm": Four-well recording chip with 150 µm cavity size (Order # 132003)
• "Meca 4 Recording Chips Fluo": Four-well recording chip for fluorescence applications with 150 µm cavity size (Order # 132004)

数据与应用：

Alpha-Hemolysin - Temperature Control

Orbit mini and applications:
Data were kindly provided by Ionera.

Temperature control of the Orbit mini: PEG induced current blockages of a alpha-hemolysin pore

(A) Current traces recorded at 10°C and 40°C illustrating a strong increase of the open pore current as well as the blockage frequency at elevated temperature.
(B) Event averaged histograms of the residual current during blockages. The open pore current scales with the temperature as well as the dwell time of the blockages.
(C) Dependence of the open pore current on the temperature.
(D) Dependence of the frequency of blockages on the temperature.

OccK1 - Outer Membrane Protein of the Pathogenic Bacterium Pseudomonas Aeruginosa

Orbit mini and applications:
Data were kindly provided by Ionera.

Representative recording from a single OmpF trimer showing current blocks due to the translocation of antibiotic Enrofloxacin, event averaged histogram.
Conditions: 150 mM KCl, 5 mM MES, pH 6, 10 mM enrofloxacin, + 60 mV

OmpF - Current block

Orbit mini and applications:
Data were kindly provided by Ionera.

Representative recording from a single OmpF trimer showing current blocks due to the translocation of antibiotic Enrofloxacin, event averaged histogram.
Conditions: 150 mM KCl, 5 mM HEPES, 5 mM MES, pH 6, 10 mM enrofloxacin, +60 mV

TRPA1 - Temperature Dependency

Orbit mini data and applications:

(A) Effect of temperature on TRPA1 activity
(B) The open probability (Po) versus the temperature and fitted with Boltzmann equation (EC₅₀ was found at 14˚C).

The Arrhenius plot of the same data resulted in a Q10 of 46 (Literature: Q10 ~ 40).

VDAC - Application of Fluoxetine

Orbit mini and applications:
Data were kindly provided by Ionera.

Current recordings of single VDA channels from selected bilayers in parallel.
VDAC was reconstituted in DPhPC membranes and partially blocked upon the addition of Fluoxetine (Prozac).

(A) Representative parallel recording from 3 VDAC channels in DPhPC membranes. Conditions: 1 M KCl, 25 mM Tris, pH 7,5, voltage change protocol is given below.
(B) Current trace illustration partial blocade of VDAC by Fluoxetine (Prozac).
(C) Current-voltage trace of a single VADC channel under a ramping voltage of -70/+70 mV before (green) and after (blue) addition of 50 µm Fluoxetine

Purified VDAC sample is a generous gift from Prof. Stephan Nussberger to Ionera, Department Biophysics, University of Stuttgart, Germany

发表文献：
2022 - Integrative Study of the Structural and Dynamical Properties of a KirBac3.1 Mutant: Functional Implication of a Highly Conserved Tryptophan in the Transmembrane Domain
2021 - Structure and desensitization of AMPA receptor complexes with type II TARP γ5 and GSG1L
2021 - Saccharomyces cerevisiae as a superior host for overproduction of prokaryotic integral membrane proteins
2021 - Rapid and multiplex preparation of engineered Mycobacterium smegmatis porin A (MspA) nanopores for single molecule sensing and sequencing
2021 - Design, assembly, and characterization of membrane-spanning DNA nanopores
2021 - ATP and large signaling metabolites flux through caspase-activated Pannexin 1 channels