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There is no doubt that cell culture requires oxygen, but not too much. When cells are cultured in traditional cell culture devices such as flasks and petri dishes, cells are exposed to the excessive concentration of dissolved oxygen (DO). The open atmosphere contains about 20% oxygen, and the oxygen from the air is dissolved in cell culture media. The solubility depends on the cell culture temperature, atmospheric pressure, and the salinity of media. Reaching the limit of oxygen solubility results in concentrations of DO in media that are far too high for normal cells integrating the living tissues. This problem may be the source of many misleading results and differences between experimental and in vivo environments.

Petaka G3™ Breakthrough Technology

The Petaka G3™ is a series of proprietary, single-use bioreactors for cell culture that automatically regulate oxygen, carbon dioxide and humidity, without any electronics or hardware. In Petaka G3™ bioreactors, adherent or suspension cells can be transported, cultured, paused, cryostored or centrifuged, reducing the risk of contamination.

Petaka G3™ bioreactors are designed to provide semi-hermetic cell chambers with physiologic respiration, using a unique system of micro-tubular gas diffusion channels. With this system, the threshold of oxygen in the cellular environment is maintained at physiological levels similar to those in living organisms, creating a perfect environment for stem cells and other mammalian cell types. In the Petaka G3™, cell culture begins with the DO concentration equivalent to that of arterial blood (oxygen partial pressure 75 mmHg), which is high enough to promote exponential cell growth. As cells proliferate, the DO concentration automatically decreases to close to the DO concentration received by the cells in normal living mammalian tissues and embryonic development (oxygen partial pressure about 15 mmHg), avoiding cell damage by reactive oxygen species (ROS) formation and facilitating cell differentiation.

Petaka G3™ cell culture systems are able to culture cells in incubators with 21% oxygen concentration. These systems reduce oxygen to 5% or even 2% under certain conditions, depending on the cell type and level of confluence, which avoids the additional use of anoxic chambers or three gas incubators.

Features of Petaka G3™

• Closed system: Petaka G3™ are simple sterile culture devices that do not require expensive equipment. No fluid leakage, spillage or cell dehydration occurs.
• Greater capacity: the internal cell culture chamber has a 150 cm² surface area for cell attachment and a volume of 24 mL. This facilitates high-density cell growth, allowing optimal use of media and special additives.
• Universal size: the bioreactors made from polystyrene occupy the same footprint as a standard microtiter plate and can be stacked on a bench or incubator.
• Availability: the bioreactors are available in 3 formats, including PetakaG3™ LOT, PetakaG3™ FLAT, and PetakaG3™ HOT.

Petaka G3™ Bioreactors

Petaka G3™ bioreactors can be used to culture a variety of cells from vertebrates and invertebrates, especially stem cells, primary cultures, and tumor cells. Petaka G3™ LOT and Petaka G3™ HOT bioreactors with hydrophilic surfaces are developed for anchorage-dependent (adherent) cell growth. Petaka G3™ FLAT with a hydrophobic surface does not allow cells to adhere to its surface and is used for anchorage-independent (suspension) cell growth and 3D cell aggregation. Petaka G3™ can carry different surface molecular matrices (such as collagen, vitronectin, and matrigel), structural matrices (nano-fibers mesh), or a combination of both.

Petaka G3™ LOT Average DO = 1 mg/L (0.5 to 1.1 mg/L depending on cell type), equivalent to average 22.7 mmHg (Torr) in tissues or equivalent to average 3.11% O2 in hypoxia chambers.
Petaka G3™ HOT Average DO = 3 mg/L (2.8 to 3.2 mg/L depending on cell type), equivalent to 66.5 mmHg in tissues or equivalent to 9.33% O2 in hypoxia chambers.

A virtually hermetical system with an air filter of 0.2 micron pore and a silicon port that is sealable after injection

With the Petaka G3™ system, live cells can be transported for long distances at normal ambient temperatures. Petaka G3™ can be used to transport most cell lines for 1-14 days without the need for dry ice or refrigeration. The slim shape of Petaka G3™ allows for easy packaging and protection. In addition, microgravity 3D culture can be performed in a Petaka G3™ bioreactor, where cells are induced to form 3D cell clusters (pseudo-tissues) under microgravity conditions.

Parameters of Petaka G3™

Use Roadmap of Petaka G3™

Frequently Asked Questions

• • What is the capacity of cells in the Petaka G3™?

    • The Petaka G3™ system is not designed to produce tens of billions of cells. Typical suspension cell will be produced 2.0-2.5mm cells/mL at peak density in Petaka G3™, or 50mm cells per Petaka G3™. Adherent cells will be typically cultured 13-18mm per side, so using both sides of the Petaka G3™ for adherent cells it is possible to yield 30mm cells per Petaka G3™. The Petaka G3™ is ideal for higher quality and safer seed culture production, or for storage and transfer of samples for cryopreservation or downstream QC testing. All cell types are different and while the above is achievable it is recommended to make this calculation based on the maximum known cell density of the specific cell type being used.

  • How is it possible to grow cells in the Petaka G3™ without supplemental CO2?

    • Cells naturally generate CO2. However, in plates, dishes and flasks CO2 rapidly diffuses out of the medium, so a closed CO2 incubator system is required. The Petaka G3™ automatically retains CO2 at a level sufficient to maintain pH in a standard bicarbonate-buffered medium. However, a minimum number of cells in the starting culture must be present to supply sufficient CO2 to maintain this pH balance. We recommend at least two-million cells (approximately 25,000 cells/cm²) be seeded in a Petaka G3™ when culturing without supplemental CO2. With this number of starting cells, your culture should have no problem maintaining the pH of the medium. Much lower cell densities can also be cultured in the Petaka G3™ but may require supplemental CO2 for the first 24 hours, or until sufficient cell numbers are reached and sufficient internal CO2 is generated.

  • At what level of dissolved oxygen will the Petaka G3™ maintain my cultures?

    • The proprietary microchannel gas regulator within each Petaka G3™ will reduce the dissolved oxygen from hyperoxic levels (150 mmHg pO2) to levels found in native tissues (25-50 mmHg pO2). The exact level of dissolved oxygen will vary depending on cell type, length of uninterrupted culture and medium conditions. It is important to understand that oxygen flows into the Petaka G3™ is controlled by diffusion, so the bigger the difference between the outside air and the inside culture chamber, the more oxygen will diffuse in. Therefore, as the internal oxygen level is reduced, more oxygen will automatically and passively diffuse inward to maintain equilibrium. Just as in native tissues, cells in the Petaka G3™ will take what they need and establish their preferred local dissolved oxygen levels. The Petaka G3™ does not force an artificial level of dissolved oxygen into the culture environment, this is why we state oxygen is “auto-regulated” and cells maintain physiologic levels of oxygen at all times.

  • What types of cells can I culture with the Petaka G3™?

    • Practically every cell type has been successfully cultivated in the Petaka G3™, including adherent, suspension, immortal, primary and transfected cells, hybridomas, MSCs, ESCs, iPSCs, etc. The Petaka G3™ FLAT is untreated for suspension cell culture, while the Petaka G3™ LOT is surface treated for adherent cells. The Petaka G3™ is not just another flask. It corrects the error of cultivating cells in the non-physiological, hyperoxic conditions with plates, dishes, and flasks. Since most cell lines have been cultivated in hyperoxic conditions for decades, Petaka G3™ users may observe differences as the cells re-adapt to their correct, physiological environment. Practically speaking, this may result in slight morphological changes, changes in doubling time, etc., but this is not a negative effect. It is actually corrective, and in the long-term, your cells will be more reflective of the native cells when grown inside the Petaka G3™.

  • Should I culture with the Petaka G3™ in an upright or horizontal configuration?

    • Suspension cells must be cultured in the Petaka G3™ in a horizontal configuration similar to a flask, because these cells will clump on the bottom when the Petaka G3™ is cultured upright. After adherent cells attach (typically 1-2 hours), the Petaka G3™ may be placed in the horizontal position or may be switched to an upright position for the duration of the culture. However, when the Petaka G3™ is kept “horizontal”, it is strongly advised to elevate the top end of the Petaka G3™ is slightly higher than the bottom (5°-15°) to limit the amount of medium that may enter the capillary breakers at the top of the Petaka G3™.

  • How can I ship cells in the Petaka G3™?

    • Cells may be shipped at ambient temperatures using the Petaka G3™. Cells should be seeded in the Petaka G3™ unit 24 hours prior to shipping to allow them to acclimatize. Petaka G3™ LOT is recommended for this purpose. Since it is not ideal to have relatively high oxygen levels in the transport unit when shipping at temperatures below 37°C.

  • How can I measure the amount of dissolved oxygen inside my Petaka G3™?

    • The Petaka G3™ is unique in that it is possible to measure the dissolved oxygen (pO2) in your culture in real-time, using a sterile, off-the-shelf setup. The Petaka G3™ DO Sense features an oxygen sensing dot manufactured by Ocean Insights, that when paired with the NeoFox™ reader, allows the user to measure and record the dissolved oxygen in the culture medium with precision and high temporal resolution. With the Petaka G3™ DO Sense, it is easy to perform simple bioprocess monitoring or to carry out sophisticated experiments determining single-cell oxygen consumption rates.

  • How is it possible to wash and centrifuge cells without transfer to conical tubes?

    • The Petaka G3™ is the only cell culture device that can be centrifuged directly. This allows the user to pellet, wash and separate cells without the need to transfer cells into conical tubes. Unlike open dishes, plates, and flasks, the Petaka G3™ is designed to withstand up to 650 g during routine centrifugation.