804A: Permeabilized Fiber ATPase Apparatus

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The 804A ATPase apparatus was designed to enable physiology researchers to measure consumption of ATP along with contractile force and length changes of permeabilized fibers. The 804A works in conjunction with an Aurora Scientific model 315D or 322D high-speed length controller and an Aurora 400C series force transducer.

The 9-well bath plate is corrosion resistant and can be used for permeabilized fiber experiments such as force-pCa and kTr. Bath 1 is the largest bath which includes right angle prisms on either side, allowing simple fiber attachment and accurate measurement of cross-section. The first bath also has a glass bottom which allows the fiber to be imaged with an inverted microscope or to set the resting sarcomere length (SL) using our High-Speed Video Sarcomere (HVSL) system. A laser diffraction system is also included for setting SL of cardiac tissue. The 804A also includes an add-on ATPase cuvette that is used to measure ATP consumption. The ATPase cuvette includes optics to excite the fluorophores in solution with UV light (970A) and to quantify the absorbed light signal from the bath using a photodetector (971A).

Utilizing a microprocessor-based controller and our 600A software, the researcher can define which bath the fiber is placed in at specified time points, simplifying a force-pCa test. Position sensors ensure bath placement accuracy to better than +/- 0.05mm. A built-in syringe pump provides nL accuracy for ADP injection into the ATPase measurement cuvette. Each apparatus includes XYZ micrometer stages with built-in mounts for Aurora Scientific high-speed length controllers and force transducers.

  • 10-well bath plate: (290µL imaging/mounting + eight 150µL mechanics baths + 21µL removable bath/cuvette for ATP consumption measurements)
  • Complete system available that includes ATPase apparatus, Length Controller, Force Transducer, dual-wavelength (340 and 430nm) LED Light Source, dual-wavelength Photodetector, data acquisition hardware, and 600A Digital Controller software
  • ADP syringe pump included with software control of ADP injection: minimum 36nL injection volume
  • Peltier-based temperature control (4-40°C)
  • Software control of bath movements and timing
  • Fast bath-to-bath transitions, 1.1 seconds (bath-to-bath), 2.5 seconds (end-to-end)
  • Compatible with all Aurora High-Speed Length Controllers and Force Transducers
  • Laser-based sarcomere length measurement included

Components

825A Thermocouple Meter / TEC Controller:
Thermometer and TEC Controller controls bath temperature from 4 to 38°C

826A Water Circulator:
Water circulator that provides heating/cooling for 804A apparatus. Includes water reservoir, pump, and radiator with fan

970A UV Light Source:
Dual-wavelength UV LED light source (340nm and 430nm) provides excitation for the fluorophores in the ATPase cuvette via an included fiber optic cable

971A ATPase Photodetector:
Dual-wavelength UV photodetector provides measurement of the intensity of the 340nm and 430nm wavelengths from the ATPase cuvette via an included fiber optic cable.

Stories of Success

804A – Permeabilized Fiber ATPase Apparatus

CHALLENGE

In 2016 Aurora Scientific was approached by Dr. Pieter de Tombe and Dr. Ramzi Khairallah to partner in the development and commercialization of a new apparatus for making measurements of ATP consumption in permeabilized tissue. This initiative was in response to a growing demand for these types of measurements after a long period of sparse publications on the subject. With no commercial ATPase systems available, and with few researchers able to construct their own systems, a need for a new product was apparent. This new product required motor-driven movement of the bath plate and sensitive, affordable, and easy-to-use, optics for measuring ATPase activity.

SOLUTION

After many prototypes and iterations, a version of the apparatus that could mount onto an inverted microscope was developed. Aurora Scientific was able to leverage their experience with precise motion control from the existing 802D fiber apparatus to develop a reliable and relatively inexpensive moving chamber. Together with Dr. de Tombe and Dr. ’Khairallah’ s extensive knowledge of optics, this solution has opened the door to new technologies that will achieve much improved ATP resolution.

RESULTS

The 804A Permeabilized Fiber ATPase Apparatus represents a true collaborative effort between industry and academia in the spirit of scientific discovery. Aurora Scientific is proud of its development and is grateful for the efforts of all those involved.

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Specifications

Bath Plate
Number of Baths 10 including: 1 Relax/Mounting, 8pCa, 1 ATPase
Bath Dimensions [mm] Bath 1 – Relax: 16L x 6W x 3D
Baths 2-9 – pCa: 16L x 3W x 3D
Bath 10 – ATPase: 4L x 1W x 5.2D
Bath Positioning Accuracy [mm] +/-0.05
Bath Volumes [µL] Bath 1 – Relax: 295
Baths 2-9 – pCa: 124
Bath 10 – ATPase: 21
Inter-Bath Distance [mm] Bath 1-2: 14
Baths 2-9: 7
Bath 10: 14
Indexing Time [second] Bath1–2: 1.3
Adjacent Baths 2-9: 1.1
Bath 9-10: 1.3
Bath 1-10: 2.7
Z-Axis Excursion [mm] High Sensor-Up Sensor: 5.75
Up Sensor-Down Sensor: 4.0
Down Sensor-Lowest Position: 3.75
Total Maximum Excursion: 13.5
Right-Angle Prism Dimensions [mm] 6L x 3D x 3H
Material Apparatus: 6061-T6 Aluminum – Black Anodized
Bath Plate: 6061-T6 Aluminum – Hard Anodized
ADP Injection Syringe
Available Volumes [µL] 10, 25, 50
Type Hamilton type RN1700, cemented needle
Syringe Dimensions [mm] 1701 – 10µL: 0.46 ID x 60 Stroke
1702 – 25µL: 0.73 ID x 60 Stroke
1705 – 50µL: 1.03 ID x 60 Stroke
Flowrate Range [pL/sec] Max:
Min:
Pump Stroke [mm] 52
Minimum Injection Volume [pL] 38
Maximum Injection Volume [µL] 1701 – 10µL: 8.7
1702 – 25µL: 21.7
1705 – 50µL: 43.3
Laser SL Measurement
LED Laser Power [mW] 5
Beam Dimensions [mm] 3 x 1
Wavelength [nm] 635
Laser Class – CDRH IIIb
Measurement Range [µm] 1.7 – 10.0
Temperature Control
Type 2 x Peltier modules
Temperature Control Range [°C] 4 – 40
TEC Power [W] 33
Voltage [V] 12
Temperature Sensor AD590
General Specifications
Operating Temperature [°C] 10 – 30
Power Required 100, 120, 220, 240 VAC, 50/60 Hz. available
Power Consumption [W] 20
Weight [kg] 2.5
Dimensions [mm] Microscope Mount Configuration: W x D x H*****
Table Mount Configuration: W x D x H*****

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