The 2PK+ Whole-Cell / Patch Pipette Perfusion Kit from ALA is the original kit for perfusing pipettes during patch clamp recording. The 2PK+ is designed to perfuse patch pipettes during single channel recording and to internally perfuse cells during whole-cell recordings. An indispensable tool for pharmacological and second-messenger studies of ion channel function, the 2PK+ is a proven system with a track record of publications in peer-reviewed literature.
Key features of the 2PK+ include:
- Optional rotating carousel, the MRC-6, for multiple solution exchanges
- Perfusion pipette holder
- Pressure and vacuum controller to balance pressure on patch while driving fluid flow
- Quartz capillary tube provides optimal noise reduction and durability for proximal positioning to pipette tip
The 2PK+ increases the type and variety of experiments that can be performed because it enables the change of solution inside the pipette and in the whole-cell itself. The use of this kit should not increase electrical noise during recordings. The generator will control and measure pressure/vacuum to an accuracy of +/-0.2mmHg throughout the range of +/- 40mmHg. This precise control of pressure and vacuum helps prevent cell damage and enables experiments with limited electrical noise. It needs a minimum gas input of 15PSI/100kPa to perform properly.
In the whole-cell perfusion mode, positive pressure and our unique pressure vessel are used to force the perfusion solution into the recording pipette which then diffuses into the cell. The perfusion solution is sealed into the pressure vessel where it is forced by pressure through a quartz capillary and then into the pipette where it is deposited near the tip. Since positive pressure is being introduced, the negative pressure port on the pipette holder can be used to balance the pipette pressure. This allows the cell (after proper tuning of the vacuum/pressure generator) to experience no net change in pressure (i.e. it is neither pressurized nor are its contents sucked out). The simple streaming of solution within the pipette causes solution to flow out the tip and into the cell. Depending on cell size, flow rates, solution viscosity, and pipette size, cell perfusion times can vary from seconds to minutes.
In the patch perfusion mode, the vacuum port is used to suck the perfusion solution into the patch pipette through the quartz perfusion capillary. The vacuum is slowly increased to start the solution flow. Once perfusion starts, the vacuum is slowly decreased until a steady state of flow is reached. In this mode the pressure port may not be used.
Each 2PK+ system includes the following:
- Pressure/vacuum generator
- Perfusion pipette holder (specify amplifier used)
- Perfusion pressure vessel
- Quartz perfusion capillaries
The 2PK+ system can be combined with the MRC-6 Multi-Reservoir Carousel for fast multiple exchanges of internal pipette solutions.
The 2PK+ system makes it easy to perfuse solutions into recording pipettes. However, due to the exacting nature of the application system (positive or negative pressure under precise control) the solution reservoirs must be manually changed. Twenty to thirty seconds can be required for these changes, and vibrations can cause the cell attachment to suffer. The MRC-6conveniently holds six reservoirs on a carousel, which allows for quick exchanges of internal pipette solutions when perfusing with the 2PK+. These reservoirs are then available for use in seconds while maintaining pneumatic and mechanical stability. The MRC-6 greatly facilitates the acquisition of dose response curves and wash-in/wash-out experiments performed inside the recording pipette.