Direct measurements of intracellular pressure were made using the 900A micropressure system (World Precision Instruments), as described previously (Petrie and Koo, 2014 ). Briefly, a 0.5-μm micropipette (World Precision Instruments) was filled with 1 M KCl solution and the resistance of the circuit was set to zero or null. The micropipette was positioned with an MPC-325 micromanipulator (Sutter Instrument) within an environmental chamber (10% CO2 and 37°C) on a Zeiss LSM700 laser scanning microscope using a 32×, 0.4 NA Ph1 objective. To make an intracellular pressure measurement, the microelectrode was inserted through the plasma membrane at a 45° angle, maintained in the cytoplasm for ≥5 s, and then removed. The pressure was measured perinuclearly, between the nucleus and the leading edge. The cytoplasmic hydraulic pressure was calculated as the mean pressure over this time interval.
900a micropressure system
Manufactured by World Precision Instruments
The 900A micropressure system is a lab equipment designed to measure and control pressure at microscale levels. It provides precise pressure measurement and regulation capabilities for various applications.
Automatically generated - may contain errors
Lab products found in correlation
Microelectrode holder half cell, by World Precision Instruments (2 mentions)
Lsm 700 laser scanning microscope, by Zeiss (1 mentions)
Mpc 325 micromanipulator, by Sutter Instruments (1 mentions)
Mpc 325, by Sutter Instruments (1 mentions)
Axiovert 200m microscope, by Zeiss (1 mentions)
Glass micropipette, by World Precision Instruments (1 mentions)
Lsm 700 microscope, by Zeiss (1 mentions)
4 protocols using 900a micropressure system
1
Intracellular Pressure Measurement Protocol
2
Measuring Endocochlear Potential in Mice
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Animals were anesthetised using 2.0% isoflurane. Tracheostomy was performed using a cannula. The bulla was opened using a retroauricular approach. An Ag–AgCl reference electrode was placed on the neck muscles, and a glass microelectrode filled with 2 M KCl was inserted into the scala media of the second turn to measure EP. The microelectrode was connected to an amplifier with high-input impedance (900 A Micropressure System; World Precision Instruments). All experiments were performed in an electrically shielded booth.
3
Measuring Intracellular Pressure with Micropipettes
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The 900A micropressure system (World Precision Instruments) was used to make direct measurements of intracellular pressure according to the manufacturer’s instructions and as described in Petrie and Koo (2014) (link). A 0.5-µm micropipette (World Precision Instruments) was filled with a 1-M KCl solution, placed in a microelectrode holder half-cell (World Precision Instruments), and connected to a pressure source regulated by the 900A system. A calibration chamber (World Precision Instruments) was filled with 0.1 M KCl and connected to the 900A system, and the resistance of each microelectrode was set to zero and then secured in a MPC-325 micromanipulator (Sutter Instrument) within an environmental chamber (37°C and 10% CO2) on an Axiovert 200M microscope (ZEISS). To measure intracellular pressure, the microelectrode was driven at a 45° angle into the cytoplasm, maintained in place for ≥5 s, and removed. The pressure measurement was calculated as the mean pressure reading during this interval of time. In individual polarized, elongated cells, a pressure measurement was first made behind the nucleus, followed by a measurement in front of the nucleus in the same cell.
4
Intracellular Pressure Measurement Technique
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Direct measurements of intracellular pressure were performed using the 900A micropressure system (World Precision Instruments) according to the manufacturer’s instructions as described previously (Petrie and Koo, 2014 (link)). Briefly, a glass micropipette with a 0.5-µm-diameter tip (World Precision Instruments) was filled with a 1-M KCL solution, placed in a microelectrode holder half-cell (World Precision Instruments), and connected to a pressure source regulated by the 900A system. The calibration chamber (World Precision Instruments) was filled with 0.1 M KCl and connected to the 900A system, and the resistance of each microelectrode was set to 0 and attached to a MPC-200 micromanipulator (Sutter Instrument) within an environmental chamber (37°C and 10% CO2) on an LSM 700 microscope (Zeiss). To measure intracellular pressure, the microelectrode was driven at a 45° angle into the cytoplasm, maintained in place for ≥5 s, and removed. The pressure measurement was calculated as the mean pressure reading during this time interval. The experimenters were blinded to the identity of the five cell lines to eliminate potential bias.
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