Presented by Zac Eichmeyer
And BEMsS (BioElectromagnetic student Society)
 The V.P. is an instrument capable of measuring current
densities in a biological medium
 Capable of measuring 10 nA/cm2
 It was developed in the 70’s to measure current pulses
from an Oocyte
 Non-Destructive measurements
 Can measure activities in live cells
 Offers an order of magnitude increase from current
detection methods, and is more versatile than dyes
How does the V.P. Work?
 The V.P. works by vibrating a high
impedance probe horizontally ~10mm
 The vibration is controlled by a
Piezoelectric Actuator
 The Probe is connected to a
Preamplifier which conditions the
signal for the Lock-In Amplifier
 The Lock-in amplifier locks the
probes measurement to that of its
vibration so to only read current
density at its max displacement
 Reducing Noise, and giving a voltage
reading at the max of its amplitude
 The data is then collected via a data
logger and fed into a computer for
analyzing
 The V.P. Requirements:
 The probe
 High impedence, greater than 5 MW
 Low length to mass ratio
 Piezoelectric Actuator (the bender)
 Operate between ± 45V
 Bend at least 30mm
 The Lock-in Amplifier
 Lock on to a 150 Hz – 1kHz signal
 The Pre-Amp
 Low Noise
 The DAQ
 Capable of sampling faster than the vibration frequency
 <1 kHz max
 The Oscillator
 Capable of driving a Piezoelectric Actuator at a frequency less
then 1kHz
 The Mechanical Manipulator
 Connects the probe to the microscope
 Rigid
 Have 1 degree of motion (up and down)
 Thermally and electrically neutral
 To be able to measure current densities <100 nA/cm2
 To eliminate/control extraneous noise
 To build the appropriate test setup to implement the
probe with a microscope
 Implement into current BEMSS research
 Measure current densities in other biological samples
 Track Ion concentrations as they relate to Neutrophils
•Project Overview (Top Down)
•The V.P. Overview(subsystems)
•Probes
•Piezoelectric Actuator (Bender)
•Pre Amp
•Lock In Amplifier
•Oscillator
•DAQ
•Housing
•Mechanical Manipulator
•Project Overview (Top Down)
 Two Probes
 Ref Probe
 Used to find a Common
 Sensing Probe
 Used to find the Potential in a given medium
 Construction:
 Plate probe in gold
 Gold is more bio-friendly than steel
 Connect to Housing
 Drivers
 Length to mass ratio
 Impedance of the probe
 Material
 Sensing Probe
 The Elgiloy Stainless Steel Microelectrodes
 Dimensions (D x L) 225mm x 80mm
 Impedence is 12 MW
 Tip Diamter, 1-2 mm
 A Piezoelectric Actuator is a device which can deflect
when given a voltage
 Will use as the vibration device
 Capable of vibrating
 Between 100 Hz to 1 kHz
 Amplitude of ± 10 mm
 Operate with a Vpp of 60 Volts
 Must be Mountable onto the microscope test setup
 Selection: Physik Instumente-PICMA® Ceramic-
Encapsulated Piezo Bender Actuator, PL112.10
 Size (L x W x T):
 Deflection:
 Input Voltage

17.8 x 9.6 x .65 [mm]
160 mm (nominal)
Nominal Voltage ± 45 V
Max Voltage ± 90 V
 Pre-Amplifier Drivers
 This conditions the signal to feed it into the Lock-in Amp
 Low Noise
 Differential input
 High Input Impedance
 At least a 10x gain
 Selection: Model 100 Preamplifier
 Available at Applicable electronics
 Lock-in Amplifier
 Needs to lock the signal from the probes to the oscillator
 150 Hz to 1 kHz ref input
 Needs to be able to output out to a DAQ system
 Selection: Stanford Research Systems
 Model SR810 DAP Lock-In Amplifier
 The Oscillator needs to provide a stable signal in the
range of 150 Hz to 1 kHz at 45 Vpp
 The probe will need to vibrate at its resonance frequency
 This range covers all possible frequencies for the probes
 Must be relatively stable
 Not crucial because the Lock In amplifier will compensate
 Selection: Hewlett Packard Model 204C Oscillator
 Holds the Probe and allows the actuator to vibrate the
probe
 Requirements
 Electrically and thermally neutral
 Rigid
 Allow light through
 Selection:
 Custom Made according to specification
 Material: Lucite
 Test Setup Drivers
 Needs to be mechanically stable
 Any vibration will be detected as extraneous noise
 Needs to be precise
 Accurate to ~5 mm
 Needs to mount to a microscope setup
 The Data Acquisition System
 A/D conversion (Measure voltage)
 Data logging capabilities
 Needs to sample at least every 10 ms
 The HOBO Data Logger
 HOBO U12 Temp/RH/2 External Channel Logger
 USB connection to Computer
 Analog Voltage measurement
 12 bit resolution (.6mV)
 0-2.5 v sensor range
 Accuracy 1%
 Three Phases : Preliminary, Intermediate and Final
 Preliminary
 Probes
 Measure Impedance
 Measure Mass
 Piezoelectric Actuator
 Verify vibration modes
 Oscillator
 Verify stability and range
 DAQ
 Connect to computer and start to log
 Mechanical manipulator
 Connect to Microscope, show degree of freedom
 Lock-in Amplifier
 Verify its ability to lock onto a signal
 Intermediate:
 Mechanical
 Housing
 Make sure everything attaches snuggly
 Verify the vibration mode with the Probe
 Show light can still get through
 Mechanical manipulator
 Attach housing, show stability
 Electrical
 Pre-Amp
 Verify the gain
 Show stability
 Probes
 Attach to Pre-amp
 Calibrate (as shown in the appendix)
 Final phase:
 Mechanical
 Attach everything to the microscope test setup
 Show functionality
 3 degrees of movement (2 by the microscope slide)
 Electrical
 Connect the Pre-amp and Oscillator to Lock in Amp
 Verify its sampling correctly
 Connect the Lock in amp to the DAQ
 Verify its outputting accurately
 Measure well known cell actions
 To prove calibration
 Start measuring voltages off of live cells
 Mile Stone 1 Deliverables
 Construct housing
 Make and Prep the Probes
 Complete all of the preliminary testing phase
 Milestone 2 Deliverables
 Complete all of the intermediate testing phase
 Physically attach all pieces together
 Begin Live cell tests
 Design Expo
 A fully working VP system, that can be demonstrated
live
Legend
PDR
CDR
Milestone 1
Milestone 2
Design Expo
BUDGET
Item
Quantity
Cost
Total
Probes
3
128
$ (384.00)
Bender
1
287
$ (287.00)
Pre-amp
1
$ (145.00)
Miscellaneous
$ (168.00)
Cost
$ (984.00)
Funding
Resources
UROP
1000
Total
24
Lock-in Amplifier
Waveform generator
BEMSS
 Expand it into two or three dimensions
 Computer controlled
 Mechanically threw the computer
 Improve sampling rate
 Min>50ms
 Integrate into BEMSS reasearch
 The VP is an instrument capable of measuring current
densities as low as 10 nA/cm2
 Used primarily for measurements in biological mediums
• The V.P. Overview(subsystems)
• Probes
• Piezoelectric Actuator (Bender)
• Pre Amp
• Lock In Amplifier
• Oscillator
• DAQ
• Housing
• Mechanical Manipulator

Pictures:

Nuccitelli, Jaffe, “AN ULTRASENSITIVE VIBRATING PROBE FOR MEASURING, STEADY EXTRACELLULAR CURRENTS”, www.jcb.org

UMASS Vibrating Probe Facility- http://marlin.bio.umass.edu/biology/kunkel/conte/polarized-probes.html

Reid, Nuccitell, Zhao, “Non-invasive measurement of bioelectric currents with a vibrating probe” Nature Protocols, Vol 2, #3, 2007, p 661-
669

Information, background and resource

Nuccitelli, Jaffe, “AN ULTRASENSITIVE VIBRATING PROBE FOR MEASURING, STEADY EXTRACELLULAR CURRENTS”, www.jcb.org

“A simple method for preparing a Vibrating Probe” T Nakagawa, http://pcp.oxfordjournals.org

Reid, Nuccitell, Zhao, “Non-invasive measurement of bioelectric currents with a vibrating probe” Nature Protocols, Vol 2, #3, 2007, p 661-669
Any Questions?