The benefits of flow culture

One of the most important challenges of tissue engineering is the reproduction of an "artificial" vascular system able to supply the growing cells with sufficient oxygen and nutrients, as well as to remove the waste matter generated by cells metabolism, is one of the most difficult issues.

Cell cultures developed on three-dimensional scaffolds under static conditions are specially sensitive to oxygen and nutrients depletion. Assuming that they are equally distributed across the volume of the scaffold at the beginning of the culture, cells should develop at a similar rate in the whole scaffold. However, those cells located at the inner areas of the scaffold transform all the available oxygen and nutrients into carbon dioxide and waste matter soon, since renovation rate by diffusion is too slow compared to cell metabolism. Then, it typically happens that scaffolds larger than 1 mm result in a shell of viable cells and matrix surrounding a necrotic core due to the absence of vascular supply.

Thus, the improvement of mass transport and nutrient exchange in three-dimensional in-vitro experiments is crucial to get viable cells populations, for which the imposition of certain flow rate across the scaffold (perfusion) has been experimentally shown very efficient, compared to static cultures.

Likewise, flow stimulation is also crucial for the appropriate development of certain kinds of tissue as, for instance, the vascular tissue. In this case, endothelial cells in the intimal layer are strongly influence by the effect of blood flow, which has been experimentally demonstrated to exert a strong influence of the behaviour of such cells and, in general, of the whole vessel. Thus, culturing of blood vessels under flow conditions is of vital importance, since no other conditions better reproduce conditions experienced by cells in-vivo.

The TEB flow bioreactor series are versatile systems which allow setting up a wide variety of flow culture circuits for many different kinds of scaffolds. Check their capabilities of the two available models.

 

 

Frequently Asked Questions

Here you will find some of the most frequently asked questions about EBERS' products.

Topics

If you cannot find the answers to your questions in this list, please contact us and will will attend you as soon as possible!

 

TEB1000 Series FAQ

  • Can I define and record my own flow profiles? +

    Yes. The TEB1000 allows defining and recording your customized flow profiles. For that purpose, three basic flow profiles are predefined in the bioreactor memory. By defining combinations of those profiles, you can easily create and store your flow profile so it is available for future cultures.

  • Can I use the TEB1000 as an ordinary incubator? +

    EBERS TEB1000 Master Unit maintains the basic capabilities of an ordinary incubator. The incorporation of the additional functionalities does no affect the original performance of the Unit as an incubator since constant 37ºC temperature and C02 concentration are guaranteed inside. However, using the Master Unit as an incubator might have a negative effect on pumps, engines and electronics used for their control, specially in 100% humidity conditions. Thus, we recommend to avoid creating high humidity conditions in the Unit in order to safeguard the electric machinery.

  • Can the TEB1000 be used with previously existing equipment? +

    Yes. Any equipment fitting inside the TEB1000 - including tubing, accessories and culture chambers - may be used combined with the accessories provided by EBERS. No one better than yourself can decide the equipment you need for your experiment.

  • Can you help me to develop my growth protocols? +

    EBERS bioreactors are general purpose and conceived to be used with many different kinds of tissue. Thus, the culturing conditions and needs for each type of culture might differ as a function of a multitude of factors, like cell and scaffold type or additive supply.

    Thus, the experimental conditions growth conditions should be particularized as a function of many variables, which joined to the specified flow conditions, we determine the result of the experiment. Therefore, developing a growth protocol requires of a deep knowledge of the experiment and the desired results, which will probably be far away from our understanding.

    No one better than yourself can determine the most appropriate growing conditions for your particular experiment though, if possible, we would not hesitate in trying to advise you within our limitations.

  • Do I need humidity when using closed flow circuits? +

    Flow circuits are tipically closed. However, it is usually necessary to establish gas interchange between the culture medium and the atmosphere, which can result in unwanted variations of the medium properties. Therefore, using the humidity tray is recommended.

    Read More
  • Does the TEB1000 control humidity? +

    The TEB1000 includes a humidity try which can be used to get high humidity levels in its chamber in order to avoid or minimise evaporation in the culture vessels. However, the TEB does not incorporate any humidity sensor, so it is not able to control de humidity level inside the chamber.

    Read More
  • How can I increase the number of simultaneous experiments? +

    There are several ways in which you can easily increase the number of simultaneous experiments maintaining the same flow conditions. A relation of the most common strategies to increase the productivity of your experiments can be found here.

  • How many samples can I grow simultaneously? +

    The number of samples you can grow simultaneously strongly depends on your experiment configuration, the culture chamber, the scaffold size and the number of pumpheads you are using.

    To give an example, if we are using the EBERS seeding rack with five circuits and two P3D chambers per circuit, we would get a total of 16 simultaneous samples.

  • What is needed to operate the TEB1000? +

    The TEB1000 has been designed to be operated without using any additional equipment than the supplied by EBERS. Controls are fully integrated in the bioreactor, so there is no need of using a notebook to control the pumps functioning.

    Control software is intuitive and easy to use without the need of reading a manual. The main testing parameters are placed at visible locations and are easy to set and modify. All of these features are easily controlled by means a tactile screen placed in the front of the bioreactor.

  • Which are the benefits of culturing cells under flow conditions? +

    One of the most important challenges when growing cells for tissue regeneration is the lack of a vascular network capable to supply the cells with nutrients and oxygen in the used substrate.

    Cell cultures developed on three-dimensional scaffolds in a still culture medium are specially sensitive to this fact. Assuming that oxygen and nutrients are equally distributed across the volume of the scaffold at the beginning of the culture, cells should develop at a similar rate in the whole scaffold. However, those cells located at the center of the scaffold transform oxygen and nutrients in carbon dioxide and waste matter soon, since renovation rate by unforced diffusion is too slow compared to cell metabolism. Then, it is found that scaffolds larger than 1 mm typically result in shell of viable cells and matrix surrounding a necrotic core due to the absence of vascular supply.

    Thus, the improvement of mass transport and nutrient exchange in three-dimensional in-vitro experiments is crucial to get viable cells populations, for which the imposing of certain flow rate the culture media across the scaffold (perfusion) has been experimentally shown very efficacious compared to static cultures.

    Likewise, flow stimulation is also crucial for the appropriate development of certain kinds of tissue as, for instance, the vascular tissue. In this case, endothelial cells in the intimal layer are strongly influence by the effect of blood flow, which has been experimentally demonstrated to exert a strong influence of the behaviour of such cells and, in general, of the whole vessel. Thus, culturing of blood vessels under flow conditions is of vital importance, since no other conditions better reproduce conditions experienced by cells in-vivo.

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TEB500 Series FAQ

  • Can I define and record my own flow profiles? +

     Yes. EBERS TEB500 series allows defining and recording your customized flow profiles. For that purpose, three basic flow profiles are predefined in the bioreactor memory. By defining combinations of those profiles, you can easily create and store your flow profile so it is available for future cultures.
  • Can I use the TEB500 as an ordinary incubator? +

     Yes. The TEB500 has the basic capabilities of an ordinary incubator. The incorporation of the additional functionalities does no affect the original performance of the Unit as an incubator since constant 37ºC temperature, C02 and 02 concentration are guaranteed inside.
  • Can the TEB500 be used with previously existing equipment? +

    Yes. Any equipment fitting inside the TEB500 - including tubing, accessories and culture chambers - may be used combined with the accessories provided by EBERS. No one better than yourself can decide the equipment you need for your experiment.

    Note that in case of metallic of electronic equipments the humidity and gas concentration conditions inside the chamber of the TEB500 may negatively affect the equipment. Please check the compatibilty of your equipments with high humidity conditions before placing it inside the TEB500 chamber.

  • Can you help me to develop my growth protocols? +

    EBERS bioreactors are general purpose and conceived to be used with many different kinds of tissue. Thus, the culturing conditions and needs for each type of culture might differ as a function of a multitude of factors, like cell and scaffold type or additive supply.

    Thus, the experimental conditions growth conditions should be particularized as a function of many variables, which joined to the specified flow conditions, we determine the result of the experiment. Therefore, developing a growth protocol requires of a deep knowledge of the experiment and the desired results, which will probably be far away from our understanding.

    No one better than yourself can determine the most appropriate growing conditions for your particular experiment though, if possible, we would not hesitate in trying to advise you within our limitations.

  • Do I need humidity when using closed flow circuits? +

    Flow circuits are tipically closed. However, it is usually necessary to establish gas interchange between the culture medium and the atmosphere, which can result in unwanted variations of the medium properties. Therefore, using the humidity tray is recommended.

    Read More
  • Does the TEB500 control humidity? +

    The TEB500 includes a humidity try which can be used to get high humidity levels in its chamber in order to avoid or minimise evaporation in the culture vessels. However, the TEB does not incorporate any humidity sensor, so it is not able to control de humidity level inside the chamber.

    Read More
  • How many samples can I grow simultaneously? +

    The number of samples you can grow simultaneously strongly depends on your experiment configuration and, in particular, of the culture chamber, the scaffold size the number of pumpheads you are using. The TEB500 counts on two independent pumping systems equipped with four casette pumpheads. Thus, up to eight independent channels can be run simultaneously.

  • What is needed to operate the TEB500? +

    The basic temperature and gas concentration control functionalities of the TEB500 can be operated from the front screen and touchpad. However, and advanced interface for PC and tablets is also available allowing the user to remotely control all the advanced features of the TEB500, including flow rate, datalogging and alarm management.

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P3D Chambers & Racks FAQ

  • Can be the P3D chambers autoclaved? +

    The P3D chambers are supplied in sterile ready-to-use conditions in order to simplify the building of the perfusion set-up. After the experiment, they can be recovered and cleaned to be used again, however it is worth noting that the P3D chambers have been designed to be single use, so EBERS can not guarantee their proper functioning when used more than once. The repeated use of the chambers can then result in fluid leakages or non-desired results.

    EBERS is not responsible of the misfunctionings occurring when working with re-utilized chambers, which are left to the solely responsibility of the user.

  • Can the P3D chambers be used with hydrogel scaffolds? +

    The P3D chambers have been designed to create flow perfusion conditions on porous scaffold. Thus, the surfaces in the direction which is perpendicular to the flow has been minimised and the dimensions of the chamber have been reduced to closely fit with the scaffold dimensions. This allows getting the most of the flow rate going through the pores of the scaffold and not throught the gaps left between the scaffold and the chamber, which must be taken into account when working with non-porous or low porosity scaffolds.

    If the porosity of the scaffold is very reduced, as it happens with hydrogels, it will exert a strong resistance to the flow, which will results in a very high flow velocity at the pores  and in an increase of the pressure at the scaffold. If the mechanical consistency of the scaffold is low, it will probably be damaged under the combination of high pressure and high velocity at the pores.

    Thus, you are recommended to carefully use low flow rates when dealing with hydrogels or low porosity scaffolds.

  • Can the P3D chambers be used with membrane scaffolds? +

    P3D chambers were designed to be used with porous scaffolds so, if the membrane is porous, it is theoretically possible to insert it in a P3D chamber and make the flow go through its pores. This is not the most common way to work with membranes, which are mainly two dimensional substrates in which the nutrient and oxygen delivery is easyly obtain by diffusion.

    However, the application of flow by means of this set up can be used to mechanically stimulate the cells by means of:

    1. The application of pressure
    2. The application of direct substrate deformation

     

    Both effects are obtained when culture media is pumped against and a non-porous deformable membrane inserted in the P3D chambers. Contact us  if you are interested in using the P3D chambers with a membrane susbtrate.

  • Can you supply P3D chambers for other scaffold types or geometries? +

    The P3D chambers have been designed to cover the most of the scaffolds types and dimensions which are frequently used in tissue engineering research. The P3D-6 model can hold scaffolds between 5 and 7 mm in diameter and up to 12 mm in length, whereas the P3D-10 model is designed for scaffolds between 9 and 11 mm indiameter and up to 12 mm length. The P3D cannot be modified to hold scaffold larger or smaller of those dimensions or to used with other non-cylindrical geometries.

    If you are interested in working with other scaffold geometries or dimensions, we strongly recommend you to contact our custom development service to explore the possibility of creating a bespoke chamber for your scaffold.

  • Can you supply P3D chambers with pre-inserted scaffolds? +

    The P3D samples are supplied disassebled without any scaffold. We do not manufacture scaffolds and, therefore, we are not including pre-inserted scaffolds in the chambers. However, we have had some experience with several scaffolds manufacturers and it could suggest specific suppliers if necessary. Contact us if you want to know more about scaffols suppliers!

    Read More
  • How many samples can be held in the Seeding Rack? +

    The seeding rack is designed to hold up to five seedin circuits two P3D chamber each. Thus, the rack can simultaneously hold up to ten P3D chambers.

    Read More
  • Is the Culture Rack necessary for the cultures? +

    It is not strictly necessary to perform a culture. However, using it is highly recommendable since it lets the user to keep the P3D chambers tidy and in a vertical position, which is very helpful in order to remove the influence of the chamber position when comparing two experiments. In addition, it becomes very helpful when simultaneously working with several samples to keep the set up manageable.

    Read More
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Seeding Circuits FAQ

  • Are the culture and seeding racks strictly necessary? +

    The culture rack is not indispensable, but is very helpful for having an organized experimental setup if several P3D chambers are being used simultaneously.
    The use of the seeding rack is strongly recommended to hold the seeding circuits upright in an adequate position.

  • Can the seeding circuits be reused? +

    Some of their components do not admit conventional ways of sterilization (autoclaving, ethylene oxide). Hence, reuse of the seeding circuits is strongly discouraged.

  • Do the seeding circuits include P3D chambers? +

    No. P3D chambers must be bought separately.

  • Why short- and long-term seeding circuits? +

    Short-term seeding circuits should never be used longer than 2-3 days. Thus, they are indicated only for seeding experiments, or for experiments with very short culture periods.

    If culture is being done after seeding with the same circuits, long-term circuits are strongly recommended.

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Static Tubular Chamber FAQ

  • Can the chamber be autoclaved? +

    Yes! The vascular chamber is made of autoclavable glass and plastics.

  • Can the chamber be used for tissue decellularization? +

    Yes! The Static Tubular Chamber can be directly used to decellularize a tubular piece of tissue or organ. Decellularizing medium can be both through the lumen and around the external surface of the organ in order to improve the efficiency of the process.

  • Can you adapt the chamber to my scaffold dimensions? +

    Yes. The standard chamber can hold scafolds between 7 and 10 mm in diameter and 20 to 80 mm in length. However, we can manufacture a bespoke chamber for other scaffold dimensions so it better adapts to the dimensions of your scaffold.

     

  • Can you repair my broken chamber? +

    The static tubular chamber is made of glass, which provides a lot of advantages regarding autoclaving, cleaning and sample visualization. However, glass is brittle and can accidentally break with use. We recommend you to contact our technical staff so they can evaluate whether your static chamber can be repaired or not. They will advice you about how to proceed with the broken chamber and indicate you the steps to take to get your chamber repaired, if possible.

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Rotating Tubular Chamber FAQ

General Brochure

 

Press here to download the brochure in PDF format.

Applications

Thanks to its versatile configuration, EBERS bioreactors may be applied to develop a huge amount of experiments with different types of cells and scaffolds.

Here you will find some experiments which have been carried out in our laboratory using the EBERS equipments.

 

 

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