PA4000/PA-101S Cell Fusion System (Computer Required)
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The PA-101 AC (Alternating Current) Generator option is used in conjunction with the PA-4000 electroporator to produce waveforms necessary for fusion of dissimilar cells. This technology, originally developed by Cyto Pulse and currently in use at many commercial antibody production companies, has now been customized to fit the needs of laboratory research around the globe. Years of design process has resulted in a more optimal system than any other leading method such as chemical polyethylene glycol (PEG), mechanical, or comparable electrofusion. Three main applications are as follows:

• Dendritic and tumor hybrid immunotherapy
• Hybridoma antibody production
• Nuclear transfer for fertility treatment

This system provides the greatest control and process feedback available, while retaining the ability to modularize later into further applications such as bacterial transfection or high throughput electroporation. In addition, the intuitive Pulse Agile Interface software quickly guides even first-time users to direct success.

• Approximately 10X or more efficient than PEG
• Patented AC waveforms to induce dielectrophoresis
• Unique coaxial electrode for optimal pearl chain formation
• Large chambers 0.75 mL volume
• Large chambers 0.75 mL volume
• Fully customizable protocol parameters
• Computer controlled operation
• Log file automatically saved after every run

Three unique components are brought together to form the PA-101 fusion system: Cyto Pulse patented AC waveforms, coaxial electrodes, and Cytofusion Medium.

Electrofusion Protocol

There are a number of techniques used to fuse dissimilar cells such as B cells and myeloma cells to produce viable hybrids. Cyto Pulse fusion technology consists of a four-step procedure for cell electrofusion: 1) cell alignment by non-uniform alternating electric field, known as dielectrophoresis, 2) membrane disruption and subsequent fusion of adjoining cells by DC pulse, 3) aided maturation of fused cells by reapplication of the alternating electric field, and 4) further maturation conditions prior to culture. Optimum parameter values are dependent on the cell types being fused.

Coaxial Electrode

Cyto Pulse coaxial electrodes are specifically designed to provide ideal conditions for cell alignment and fusion. Years ahead of the competition, their revolutionary new design has proven to be the most efficient method of inducing dielectrophoresis by consistent non-uniform electric field. The coaxial shape is what causes AC waveforms to impose a greater electric field near the center of the electrode, thus moving cells uniformly together into pearl-chains. In addition, because the electric field is the same throughout the chamber from top to bottom, there is no wasted volume as opposed to Petri dish-wire electrodes.

 

• Chamber volume 750 µL
• Clear Pyrex optically clear bottom for easy cell visibility by regular or inverted microscope
• Durable stainless steel body prevents sample contamination
• 1% electrode gap tolerance ensures high repeatability
• Alcohol clean or autoclave up to 10 times

Cytofusion® Medium

Critical to the cell fusion process, Cyto Pulse Cytofusion® Medium provides users with a low conductivity solution to prevent the persistent complication of cell death from excessive heat buildup.  It is and is extremely important to wash cells at least twice in order to avoid turbulence during the alignment process. In addition, Cytofusion® Medium is formulated to optimally balance divalent cations while tightly controlling osmolarity, pH, sterility, and endotoxin levels for ensuring consistent results from fusion to fusion.

System Monitoring

There is no mystery to what's inside the box when all necessary information can be accessed from the outside - Included as standard features is pulse voltage monitors ready to attach with an external oscilloscope in order to view real-time pulsing conditions. Using this connections is a true safeguard to know exactly how the applied waveform reflects the programmed waveform, so you never again have to wonder if your protocol ran as intended.

In addition, information relevant to each executed protocol is automatically saved as a log file, including magnified voltage levels of all pulses.

Fault Detection

All Cyto Pulse Sciences equipment is designed to withstand extreme user conditions. In the event of a fault, advanced control circuitry immediately detects the source of any problem and proceeds to halt the system to prevent permanent damage. The safety of Cyto Pulse customers is our number one priority.

• Electrode Interlock - improperly attached cuvettes or electrodes will cause the protocol start function to be disabled until corrected
• Internal Fault - damaged internal system components will cause high voltage to be turned off and fault screen to be displayed

Specifications PA-4000 with PA-101 connected

General
Pulse Waveform	Sinusoidal (AC), Rectangular (Square)
Interface	Computer with Windows®
Electrodes	Coaxial, Cuvette, In-vivo
Pre-pulse AC
Begin Amplitude	5 - 75 volts pk-pk
End Amplitude	5 - 75 volts pk-pk
Duration	0 - 140 sec
Frequency	0.2 - 2 MHz
DC Pulse
Pulse Amplitude	20-800 volts @ 5 volt step
Pulse Amplitude Accuracy	±5%
Max E-Field (2.5 mm gap)	3.2 kV/cm
Pulse Duration	0.001 - 0.1 ms
Pulse Interval	0.125 sec
Number of Pulses	1 - 10
Number of Groups	1 - 10
Post-pulse AC
Begin Amplitude	5 - 75 volts pk-pk
End Amplitude	5 - 75 volts pk-pk
Duration	0 - 140 sec
Frequency	0.2 - 2 MHz
Safety / Monitoring
Electrode Interlock	Yes
AC Monitor	BNC Output
Pulse Voltage Monitor	BNC Output
Electrical
AC Line	100 - 240 VAC 50/60Hz
Physical
High Voltage Outputs	SHV
Computer Control	RS-232
Dimensions	PA-4000 12.6" x 11.8" x 8.2" (32 cm x 30 cm x 20.8 cm) PA-101 12.6" x 11.8" x 3" (32 cm x 30 cm x 7.5 cm)
Weight	PA-4000 23 pounds (11 kg) PA-101 8.8 pounds (4 kg)

 

Included in System	PA-4000/PA-101S
PA-4000	PulseAgile® Advanced Rectangular Wave Electroporator
PA4-SW	PulseAgile® Application Software
PA4-UMAN	PA-4000 User Manual
CE-20	Cuvette Holder
CUV-M	Cuvette Multi-pack set of 5 each 1 mm, 2 mm, 4 mm
CS-L-XX	IEC Line Cord, Serial cable, XX=country
PA-101	AC Generator
PA101-UMAN	PA-101 User Manual
FE-10	Coaxial Electrode Fusion Holder
FP-C25/400	Coaxial Electrode, 2.5 mm gap 750 µl volume
CS-OPT	Interface Cable Set: DB25 cable, HV cable
CPS-LCM-C	Low conductivity media, 500 ml bottle