FAQ | Esco VacciXcell

Are you offering training courses for your products?

Yes, the training courses will enable customers to acquire proper knowledge and skills to set up and operate the equipment. In addition, VacciXcell offers advanced courses that cover the topics such as preventive maintenance, inoculation/cell culture and sampling techniques, harvesting and primary product recovery. All bioreactors and fermenters are ready-to-use and delivered with complete FAT, installation, and SAT.

Is it possible to request a live demo of your bioreactors end fermenters?

Yes, interested customers can contact our technical support team for further information.

How does your company guarantee the continuous supply of the products?

We provide a continuous supply statement called "Material and Service Supply Guarantee". The content can be modified to meet all of your basic requirements.

How can I get price information of your products?

You can reach us through email at [email protected] or call us by phone at +65 6542 0833. We recommend you to provide us information by filling up the form of Preliminary Questions for Potential Customer in order for us to properly assist you on the experiments and budgets you require.

Do you provide warranty for your products?

Yes. We provide one year warranty for TideXcell system from the completion day of SAT against defects in material and workmanship. Consumables such as quick connectors, tubing, and cables are excluded. We also provide preventive maintenance for best performance during operation and process.

What are your capabilities for the successful promotion of your products?

We are knowledgeable on cGMP cell culture, virus production and biologics production. Our staff have basic cell culture knowledge with proper training to support sales activities.
We provide quick and good after-service and maintain engagement with our clients.
We actively participate in exhibitions to demonstrate our products and find the new leads.

Is it possible to culture hematopoietic cells?

How long does it take for the carrier to be saturated with cells?

Cell attachment is dependent on the cell line. It may vary from a few minutes to a few hours

Is there a chance of blockage in the system if there are cellular debris of 50%?

There is zero shear stress due to the gentle Tide Motion (only 2mm/s max). This means that there are almost no cell debris thus no blockage whatsoever.

Do cells grow in 2D or in 3D?

Cells grow in 3D

How to determine when to change the medium?

This is determined by glucose consumption of the culture. Generally, glucose level starts at 4.5 g/ml and media change is required when the glucose level goes down to 1 g/ml. Media glucose requirement may change depending on many factors and having a glucose curve would help determine the most efficient time for media change.

How can I prevent contamination during cell culture?

Possible sources of contamination during cell culture include improper sealing to the matrix vessel or connectors, non-sterile components in contact with the culture medium, and inappropriate aseptic connection techniques. It is important that filling reagents or connecting tubes must be handled under sterile condition such as a laminar flow cabinet whenever possible.

What are the causes of poor product yield?

If the setting of parameters for the bioreactor are out of the optimal range, the product yield of the cell culture may have been compromised. Over-fluctuation of pH or DO readings can be caused by the lack of nutrients, such as glucose, in the culture media and may result to poor yield of the product.

How long is the product harvest of secretory proteins?

Once the cell growth reaches plateau, the product harvest of secretory protein may last for a few weeks. Our records shows 4 months without breaking the bellow of CelCradle.

Can serum-free medium be used in your product?

Yes, the serum-free medium is compatible for cell culture in both CelCradle and TideXcell system. In an effort to eliminate potential risks associated with the introduction of adventitious agents from animal-derived components, VacciXcell also provides a medium supplement, Super Plus, to replace the use of serum in the medium.

Do I have to measure pH of the culture medium every day during cell culture?

It is necessary to replace the old medium three days after cell seeding; this is to ensure the attachment and propagation of cells. The appearance and turbidity of the medium can be inspected with the naked eye, while the pH can be measured when changing the medium. However, if you prefer not to change the medium, just add NaHCO3 to adjust the pH (till orange red).

How often should I change the medium during cell culture?

This highly depends on the customer experimental settings, such as the CO2 introduction, the components of the medium, the number of cells during cell seeding, the metabolic activity of the cells, growth factors' activity, and total working volume of your culture medium in CelCradle/TideXcell bioreactors. A rough visual indicator could be the change in the color of the culture medium (if phenol red is included) due to pH change, while a more accurate indicator is the measurement of glucose levels. The medium can be changed when the glucose level drops below the desired threshold.

Can I reduce the measurement of glucose concentration from triplicate to duplicate during cell culture?

Yes, you can. The purpose of triplicate measurement is to increase the precision of monitoring and controlling the process. The frequency of measurement can be decreased after the process has stabilized.

Are there any chances of virus clumping in TideXcell or CelCradle^® system as compared with the roller bottles?

Virus aggregation has not been reported in TideXcell or CelCradle^® system. The system functions by exposing and submerging the carriers periodically, in which the mechanism is similar with the roller bottles. The major difference is that the cell growth is in 3D pattern. However, higher virus concentration during down-stream purification process may cause virus aggregation and users can add emulsifying agents (Tween20 or Tween80) to alleviate the aggregation problems.

How can I maintain the integrity of the pH probe?

The probe must be washed thoroughly with appropriate cleaning agents before and after use.
Do not leave the end of the probe dry and keep it submerged in the electrolyte (e.g. 3 M KCl) when not in use to extend the lifespan. The electrolyte should be changed regularly.
Be careful not to introduce any bubbles to the electrode as this will make unstable measurements.
The pH probe can usually tolerate 20 - 50 autoclave cycles.

How can I maintain the integrity of the DO probe?

The probe must be washed thoroughly with appropriate cleaning agents (e.g. ddH₂O). Do not use alcohol as this can damage the sensor or lead to erroneous current.
The membrane foil should be cleaned and properly installed prior to calibration.
If you find fluctuation in the readings, inability to do calibration, or low DO probe current, the membrane body might need to be replaced.
For short term storage, keep the sensor submerged in O₂ electrolyte. For long term storage (at least 6 months), the probe should be stored dry.

After cell seeding, how long will it take to reach maximum cell count?

It will take approximately 10 days, when seeding 1x10⁸ Vero cells in one CelCradle bottle (100 ml matrix), to reach maximum cell density (4x10⁹). On the other hand, it will take approximately 6 days, when seeding 1.5x10¹¹ cells in TideXcell-100 (100 L matrix), to reach the maximum cell density.

How can we observe the Cytopathogenic Effect (CPE) after virus infection of cells?

CPE can be observed the same way as how you observe cells by fixing the carriers with alcohol and staining them with dyes. Different viruses will have different CPE with different cells. Some viruses will cause clear CPE, and some will not. For instance, rabies virus was observed under fluorescence microscopy and no CPE was detected; on the other hand, Sf-9 insect cells infected with baculoviruses were observed to have shrunken cells, while H5N1 influenza virus causes cell death and shape irregularity in MDCK cells.

For virus production process, we usually decide the end point by checking the glucose consumption rate, since CPE provides a rough evidence, while the glucose consumption rate is a more accurate determinant of harvesting time. Moreover, the end-point also depends on the stability of the virus during post-infection period.

How can we be certain that all the cells are infected by the virus after the inoculation period?

You can use a virus counter to determine the free total virus particles before and after infection. Alternatively, if you are not required to monitor real-time change, you can perform a validation study by measuring the virus titer against time after infection. You can analyze the medium using conventional method with intervals of 10 min., 30 min., 60 min., 2 hours, etc.

How much % CO₂ is required to maintain the pH after infection?

CO₂ concentration is adjusted dynamically according to the pH during cell culture. If cells continue to grow or multiply after the infection of virus, CO₂ requirement will be minimized. However, if the cells die, there will be an increase in pH and CO₂ requirement will increase as well.

What are important factors for optimization of virus production process?

Healthy cells
Infectious active virus
Post-Infection temperature
Multiplicity of Infection (MOI)
Harvest strategy

What cell lines can be grown on the CelCradle^® bioreactor?

All commonly-used adherent cells have been grown on the CelCradle^® bioreactor including VERO, HEK 293, CHO, and MDCK cells.

What are the advantages of CelCradle^® over other bioreactors available in the market?

The mechanism of CelCradle^® enables fast and complete diffusion of nutrients and oxygen in the medium to the matrix. All ingredients in the medium are properly mixed while the CO₂ produced by glucose hydrolysis can be expelled efficiently along the run. The CelCradle^® system also has a high productivity – 1 CelCradle^® bottle is equivalent to 20 x 850cm² roller bottles, 4 x 500ml Spinner Flask, 100 x 180cm² petri dishes, 2 x 6320cm² plates or 125 x 150cm² T-flasks. It also provides cells with an environment of low shear stress, zero foaming and bubbling, and no oxygen limitation.

Is CelCradle^® patented?

CelCradle^® are well patented and the patent covers US, Europe, Japan, China and Taiwan.

How many matrices are in CelCradle-500 family?

There are regularly 865 pieces of BioNOC II^® in each CelCradle bottle. The deviation is less than 5%.

How can I detach the cells growing on the BioNOC II^® carrier in the CelCradle?

There are several ways to solve the issue:

To use enzyme other than trypsin, for example, Accutase. Some other enzyme but not trypsin-based may also be helpful.
To disrupt the cells from the carriers through freeze-and-thaw, with lysis buffer, or other methods.

How many pieces of carriers in CelCradle are required for cell counting?

Two pieces of carriers are required for every cell counting procedure. However, it is suggested to perform the test thrice and then stain with CVD and then multiply averaged calculated cell number by 865 (piece/ CelCradle bottle). The result would be the total cell numbers. Before trypsinization, check the reference to make sure that cells can be detached. It is usually more difficult to detach bigger cells.

What is the cell productivity with respect to medium volume (# of cell/ ml of medium) of CelCradle?

Without changing the gene of cells, it is very unlikely to promote the cell propagation while limiting critical nutrients. With the same quantity of medium in a certain period of time, CelCradle’s maximum output may not be different from those of other commercial products. However, CelCradle does present higher cell density due to the cell growth pattern.

What is the maximum productivity of one CelCradle^® bottle? Is it easy to harvest the cells?

The output of one CelCradle^® bottle is approximately 5 × 10⁹ cells/bottle, which will vary depending on the cell type. Cell harvest is easy, you can follow the provided SOP and achieve more than 90% recovery.

Can I use CelCradle Stage for suspension culturing of the tumor cells?

CelCradle only works for adherent cell culture. However, if the specific suspension cells could attach in the CelCradle system, the secreted protein can be harvested for long periods with much lower cost than commercial suspension culture system.

Can I use CelCradle-500A or CelCradle-500AP in cell culture for production of secreted product?

Yes. CelCradle-500A or CelCradle-500AP is suitable for production of both secreted and non-secreted products. Since these two types of bottles are specifically designed to enhance the cell attachment efficiency, we suggest to use them with the up-side-down seeding protocol if your cell line has difficulties to adhere the carriers.

What is the level of the ingress protection of CelCradle Stage?

The main console of the CelCradle Stage meets IP54.

Can the parameters we set up in CelCradle-500 be applied in TideXcell-002?

CelCradle-500 is used mostly in the lab for research and tissue engineering whereas TideXcell-002 can be applied in both academy and small pilot tests. The final cell densities in both systems are quite related proportionally. However, the productivity and cell growth rate depend on several factors.

Since CelCradle-500 doesn’t have pH control and perfusion culture mode, the cell growth usually is slower than in TideXcell-002, under a manual operation. With respect to the linearity of scale-up, TideXcell-002 is better than CelCradle-500. The parameter setting in CelCradle-500 could be used in TideXcell-002, but some modifications are still necessary. Compared with other commercially-available systems, however, very few parameters are required to input in the TideXcell system. Therefore optimization is quite straight -forward.

We usually coat our cell culture surfaces with Matrigel (BD) for HEK cell line. Would that be OK?

The surface of the BioNOC II^® carriers in the CelCradle is negative charged and should be OK for the coating of Matrigel if it could be coated on petri dish or T-flask (those are all negative charge surface). Some sources of HEK293, for example, are difficult to attach, and therefore we would make coating or use CelCradle-500A to enhance the cell attachment efficiency.

Documentation related to the validation on the CelCradle system?

VACCIXCELL Bioengineering is an ISO 9001:2008 certificated company. The quality documents for CelCradle include the Certificate of Qualification (CoQ) and Certificate of Compliance (CoC). The CelCradle Stage includes the CE certificate and Certificate of Qualification (CoQ). Can the standard CelCradle bottle be frozen?

Can the standard CelCradle bottle be frozen?

Yes. The bottle could be placed in -80 °C freezer directly for free-and-thaw process to extract cellular components.

How can I maintain VERO cells in serum free medium for 10 days in CelCradle?

Once the nutrients of medium are consumed, you need to refresh culture medium. To maintain 10 days without exchanging culture medium, under high cell density, is very difficult unless the culture temperature can be reduced to below 10 C. Or supplement such as glucose and glutamine concentrate should be added to maintain the cell viability. Be sure to control pH within the proper range (7.0~7.4) by adding concentrated NaHCO3. However, the osmolarity will increase and affect cell viability if too much sodium bicarbonate is added.

How can we conclude that the cells have reached maximum density and may not further grow stage?

Cell growth test must be performed and the cell density must be monitored every other day until the cell growth reaches a plateau (no further growth of cells). The maximum cell density will vary depending on the cell line.

Is the virus infection procedure similar to the cell seeding procedure using the CelCradle^® 500A bottle? Should the bottle be placed upside down?

It depends on how the viruses infect cells. If the viruses could effectively infect the cells, he could do infection as the regular way as cell seeding.

For virus infection, how much volume do you suggest the users use?

Usually 100~120 ml is sufficient.

For trypsinization, is it recommended to place the bottle upside down, similar to seeding, for maximum cell detachment?

Yes. For the cell harvest, it is better to use CelCradle-500A since it requires less trypsin to detach cells, less wash buffer, and has higher cell recovery rate.

Can the parameters set up in CelCradle-500 be applied in TideXcell-002?

Since CelCradle-500 doesn’t have pH control and perfusion culture mode, the cell growth usualy is slower than in TideXcell-002, under a manual operation. With respect to the linearity of scale-up, TideXcell-002 is better than CelCradle-500. The parameter setting in CelCradle-500 could be used in commercially-available systems, however, very few parameters are required to input in the TideXcell System. Therefore optimization is quite straight-forward.

Are bottles sterile? What sterilization method?

Yes, the bottles are pre-sterilized using gamma irradiation process

What kind of proteins can be grown in the CelCradle?

As long as the desired protein is produced by adherent cell it can be grown in CelCradle

When harvesting the cells, the medium has to be stirred to the cells get released?

Yes. Mechanical agitation through rotating or shaking the bottle is required. Alternatively, clients can try the automated harvesting system for CelCradle for USD 5,000.

What is CelCradle?

CelCradle is a high yield tide motion cell culture device for the production of cells, proteins, monoclonal antibodies, viruses, and/or vaccine. CelCradle provides high aeration rate under the culture environment, allowing a high cell density culture to be achieved easily.

What is CelCradle composed of?

CelCradle is composed of CelCradle Stage, Controller, CelFeeder pump –depends on the application- and CelCradle Bottle.

CelCradle bottle is composed of 4 parts: Vented bottle Lid, Culture Container, BioNOC II^® (Carrier) Container, Media Container. Depends on Bottle type, some also comprise (500A and 500AP bottles) non-vented bottle lid.

Are CelCradle bottles autoclavable?

CelCradle bottle is ready to use bottle, designed for one time use only, and cannot be autoclaved. Reusing CelCradle bottle is not recommended.

What accessories are required to use CelCradle?

Users need a CelCradle Stage-CelCradle controller and 6-cube ft. CO₂ incubator. No tubing or pumps are required for batch system (CelCradle-500 or CelCradle-500A). Additional pump system, namely CelFeeder-1300 and tubing set will need to operate CelCradle-500P or CelCradle-500AP bottles.

What type of filter is using in CelCradle?

We are using PTFE hydrophobic filter with 0.22um pore size.

What is the material of CelCradle bottle?

CelCradle has 3 materials. Bottle parts is made of PET, Bellow part is made of EVA and Other blue parts are made of PP.

Can I culture cells by using serum-free medium in CelCradle?

Yes. Most serum-free medium can be applied in CelCradle-500 system.

How can we exchange the media?

If CelCradle-500/500A is used, then just to replace culture medium manually by opening the cap inside biosafety cabinet and discard the used culture medium and then fill in fresh culture medium.

If CelCradle-500P/500AP is used, then need to replenish the culture medium in the reservoir depends on how quick the glucose is consumed.

We usually coat our cell culture surfaces with Matrigel (BD) for HEK cell line. Would that be OK with BioNOC II^®?

Can cells in CelCradle be observed under a microscope?

Users have to sample the BioNOC II^® carriers inside the CelCradle before observing the cells. The disks are made of porous nonwoven fabrics. The common staining solution is hematoxylin, coomassie brilliant blue G, ... etc.

Can I take out carriers for sampling during culture in CelCradle?

Yes. CelCradle-500/500P equips a sampling port on the carrier basket. With sterilized forceps, users take out carriers during culture.

CelCradle-500A/500AP doesn’t have carrier basket, so with sterilized forceps, users can take out carriers, or harvest carriers.

Can I calculate cell population during culture in CelCradle?

There are two methods for users to observe the culture performance.

Way 1 : to measure the metabolite consumption in the cultured media. The pH and glucose are the bassic parameters to be monitored during culture.

Way 2 : to count cells. You can pick up carriers during culture, and count the cells by using dye.

Can I harvest cells after culture in CelCradle?

CelCradle-500 and CelCradle-500P are not possible to harvest the cells. Only CelCradle-500A and CelCradle-500AP are available.

How can I detach the cells growing on the BioNOC II^® carrier in the CelCradle with not using trypsin?

There are two ways to solve the issue:

Way 1 : To use enzyme other than trypsin, for example, Accutase. Some other enzyme but not trypsin-based may also be helpful.

Way 2 : To disrupt the cells from the carriers through freeze-and-thaw, with lysis buffer, or other methods.

How can we do seeding? How much volume is needed for seeding?

Usually we put 450 – 470ml fresh culture medium in CelCradle first and prepare cells in 50ml centrifuge tube after collection. Then we dispense cells with 30 – 50ml into the CelCradle bottle.

Are bottles sterile? What sterilization method?

Yes, the bottles are pre-sterilized using gamma irradiation process

What kind of proteins can be grown in the CelCradle?

As long as the desired protein is produced by adherent cell it can be grown in CelCradle

When harvesting the cells, the medium has to be stirred to the cells get released?

Yes. Mechanical agitation through rotating or shaking the bottle is required. Alternatively, clients can try the automated harvesting system for CelCradle for USD 5,000.

What is CelCradle?

What is CelCradle composed of?

CelCradle is composed of CelCradle Stage, Controller, CelFeeder pump –depends on the application- and CelCradle Bottle.

Are CelCradle bottles autoclavable?

CelCradle bottle is ready to use bottle, designed for one time use only, and cannot be autoclaved. Reusing CelCradle bottle is not recommended.

What accessories are required to use CelCradle?

Users need a CelCradle Stage-CelCradle controller and 6-cube ft. CO2 incubator. No tubing or pumps are required for batch system (CelCradle-500 or CelCradle-500A). Additional pump system, namely CelFeeder-1300 and tubing set will need to operate CelCradle-500P or CelCradle-500AP bottles.

What type of filter is using in CelCradle?

We are using PTFE hydrophobic filter with 0.22um pore size.

What is the material of CelCradle bottle?

CelCradle has 3 materials. Bottle parts is made of PET, Bellow part is made of EVA and Other blue parts are made of PP.

Can I culture cells by using serum-free medium in CelCradle?

Yes. Most serum-free medium can be applied in CelCradle-500 system.

How can we exchange the media?

If CelCradle-500/500A is used, then just to replace culture medium manually by opening the cap inside biosafety cabinet and discard the used culture medium and then fill in fresh culture medium.

If CelCradle-500P/500AP is used, then need to replenish the culture medium in the reservoir depends on how quick the glucose is consumed.

We usually coat our cell culture surfaces with Matrigel (BD) for HEK cell line. Would that be OK with BioNOC II^®?

Can cells in CelCradle be observed under a microscope?

Can I take out carriers for sampling during culture in CelCradle?

Yes. CelCradle-500/500P equips a sampling port on the carrier basket. With sterilized forceps, users take out carriers during culture.

CelCradle-500A/500AP doesn’t have carrier basket, so with sterilized forceps, users can take out carriers, or harvest carriers.

Can I calculate cell population during culture in CelCradle?

There are two methods for users to observe the culture performance.

Way 1 : to measure the metabolite consumption in the cultured media. The pH and glucose are the bassic parameters to be monitored during culture.

Way 2 : to count cells. You can pick up carriers during culture, and count the cells by using dye.

Can I harvest cells after culture in CelCradle?

CelCradle-500 and CelCradle-500P are not possible to harvest the cells. Only CelCradle-500A and CelCradle-500AP are available.

How can I detach the cells growing on the BioNOC II^® carrier in the CelCradle with not using trypsin?

There are two ways to solve the issue:

Way 1 : To use enzyme other than trypsin, for example, Accutase. Some other enzyme but not trypsin-based may also be helpful.

Way 2 : To disrupt the cells from the carriers through freeze-and-thaw, with lysis buffer, or other methods.

How can we do seeding? How much volume is needed for seeding?

Usually we put 450 – 470ml fresh culture medium in CelCradle first and prepare cells in 50ml centrifuge tube after collection. Then we dispense cells with 30 – 50ml into the CelCradle bottle.

Do I need programming skills in operating hybrid bioreactor?

No, the hybrid bioreactor contains PLC-based control structure and a touch screen HMI which makes it easier to use and understand.

How much carriers are required in 2 L, 10 L, 20 L, and 100 L matrix vessels of TideXcell^® system?

Model	Vol. of BioNOC II^®	Matrix volume	Equivalence to Stirred Tank Bioreactors
Model	Vol. of BioNOC II^®	Matrix volume	1:10 Mixing Ratio	1:20 Mixing Ratio
TideXcell-002	110g	2000 ml	20L	40L
TideXcell-010	550g	10,000 ml	100L	200L
TideXcell-020	1100g	20,000 ml	200L	400L
TideXcell-100	5500g	100,000 ml	1000L	2000L

What is the cell seeding requirement and productivity for TideXcell systems?

Model	Cell Seeding (Recommended)	Expected Resulting Cell Density
TideXcell-002	2x10⁹ cells	2x10¹⁰ cells
TideXcell-010	1x10¹⁰ cells	1x10¹¹ cells
TideXcell-020	2x10¹⁰ cells	2x10¹¹ cells
TideXcell-100	1x10¹¹ cells	1x10¹² cells

What's the ratio between the matrix vessel to the external medium tank?

This actually depends on the process. For rabies vaccine production, the proper volume ratio of matrix vessel to medium tank is 1:10.

What is the voltage of power supply for TideXcell system? Will be UPS necessary?

It should be 220 V, 50/60 Hz, single phase. The UPS is highly recommended.

What kind of special gas, utilities, or services are needed?

TideXcell system requires only electricity and compressed air. Three kinds of gases could be introduced in the system– air, CO2, and O2. For the area that electricity blackout occurs frequently, UPS is highly recommended.

How can I ensure the continuous running of the TideXcell system if the sudden power outage occurs?

All TideXcell systems can be equipped with on-line UPS (option) to avoid external interruption to the power supply.

Is the function of "Emergency Stop" available on TideXcell?

Yes, the button of Emergency Stop is on accessible area and independent for each major unit.

What are the major parameters controlled in TideXcell^® system?

The TideXcell^® system controls the up and down speed of the medium in the matrix vessel, duration of aeration and nutrition, temperature, level, duration of each FEED/HARVEST cycle, pH, DO, CO₂ concentration in the incubation chamber, valves for each gas, peristaltic pump speed, gas flow, and etc.

Do I need programming skills to automate my culture process?

All TideXcell^® systems are equipped with TideCommander and TideTracer. These user-friendly software offer a convenient way for parameter settings, profile check, and various feed/harvest strategies without any need of programming.

Does TideXcell system control DO?

TideXcell can only monitor the value of DO. We can supply pure oxygen from the mixing tank and from the data, the DO could rise to 80% with small amount of oxygen supply. That means the TideXcell could support most oxygen from the matrix side.

In TideXcell system, can I control pH by using CO₂ and NaHCO₃ as the other bioreactors?

TideXcell^® system controls the pH either by introducing CO2 to the matrix vessel in the culture chamber or by loading NaOH or HCl solution to the medium mixing vessel (bag). NaHCO₃ can be used as an alternative for NaOH, this will not become a problem in perfusion culture since the osmolarity will not accumulate.

Are pH and DO traceable? Is it possible to log the record on paper?

The data from pH and DO can be collected and saved automatically and are traceable. They can be retrieved depending on the time (Year-Month-Day-Minute-Second) but cannot be modified, in accordance to regulation standards. RS485 communication protocol/pin-out definition can be provided. Real-time data can be printed on paper with the use of RS485 interface.

What is the requirement of the floor load for TideXcell systems?

A floor load with 500 kg/m² is enough.

How are periodical medium filling and draining of the matrix vessel achieved in TideXcell system?

By air pressure. An air pump in the controller pump in and out of the air from the matrix vessel. When pumping filtered air into the matrix vessel, the medium will drain out to the external medium tank (bag); when pumping air out of the matrix vessel, the medium will fill(refill) in to the matrix vessel.

What flow rate should I use in TideXcell systems?

Each scale of TideXcell system has a different range of flow rate setting: TideXcell-002: 0.8~1.8 L/min; TideXcell-010: 1.8~3.5 L/min; TideXcell-020: 3.5~5.0 L/min; TideXcell-100: 20~30 L/min. At the initial stage of the cell culture, you need to apply somewhat lower flow rate in order to seed the cells on the BioNOC II^® matrices.

How can I sample cells directly from the matrix vessel?

The purpose for sampling cells is to have an overall image about the cell viability and density daily. In the TideXcell matrix vessel, the sampling port with the matrix basket allows the user to get pieces of BioNOC II^® matrices for cell observation and quantification. The glucose consumption rate is another index to judge the cell growth.

What are the building materials of TideXcell system for product and non-product contact surface?

The external surfaces of the TideXcell^® System is made of SS304L, while the internal surfaces are made of SS316L.

Are there any alarms in case of emergency situation or critical deviation?

Yes, besides the function of "Emergency Stop", event and alarming messages provide operators real-time monitoring of process variables and historical data. All deviations and user control actions (e.g. log-in/off, door open) are listed in the record.

An optional alarming notification is through the SCADA system or mobile communication such as cell phones via e-mail.

Is the monitor mount and keyboard tray available for TideXcell system?

Yes. We can provide special monitor mount and keyboard/mouse, which are ergonomic, for the customer.

How can TideXcell system match each customer's unique layout planning?

It is prerequisite for us to propose the layout as well as the 3D location and orientation of TideXcell systems on the customer site. It is part of URS.

What kind of validation of TideXcell system has been provided?

The validation of TideXcell system includes four major phases: DQ, IQ, OQ, and MQ. The validation may involve in inspection, functional testing, and documentation review, verifying that the system can run appropriately at the customer site.

DQ, Design Qualification, defines the functional and operational specification of an instrument or system

IQ, Installation Qualification, is the pre-installation detail that an instrument or system, its component parts and location, are fit for the purpose and satisfy the objectives or the user to carry out the intended function to expected standards.

OQ is Operation Qualification, demonstrating that an instrument or system will function according to its operational specification in the selected environment.

MQ is Maintenance Qualification, describing any maintenance required.

Does ESCO VACCIXCELL intend to offer stainless steel SIP/CIP bioreactors?

Yes, Esco VacciXcell offers Stainless Steel SIP stirred tank bioreactors and fermenters and Stainless Steel SIP solid state fermenters. Should the client require it, VacciXcell can also provide stainless steel SIP mixing systems for the TideXcell^® System.

My culture process requires up to 400 L of media. How does the autofeeder AF-1200 work? Will it automatically pump the media from the Feed tank once the mixer depletes? Or the operator has to be manually set the transfer of the remaining 100 L media in the feed tank to the mixer again?

The operator will define the feeding rate, and the autofeeder will pump in (the fresh medium) and pump out (the waste medium) automatically. For example, when we set the feeding rate is 2 L/hr, and 24 cycles/day (pump in and pump out 24 times a day, or once per hour), the autofeeder will pump out 2 L of waste medium first from the bioreactor to the waste line (or Harvest tank), and then pump in 2 L of fresh culture medium from the Feed tank to the bioreactor. The overall culture medium inside the bioreactor always keeps the same.

Because TideXcell system is based on the "Tide" movement. How can the system be homogeneous?

Cells are exposed to aeration during emerging and to nutrition during submerging and therefore vertically, cells may have different aeration and nutrition times. However, our findings show no disadvantage as long as the system can provide cells with sufficient oxygen and nutrient.

The real situation is that the cells are embedded inside the carriers and there is always a layer of culture medium absorbed the carriers. Therefore, whether the system is in aeration or nutrition phase, the cells are technically always in the liquid phase. The quantity of culture medium remaining on the carriers is 20% of the entire bed volume, which is sufficient to supply the nutrient to the cells. The tide movement is to maintain the supply of oxygen and nutrients and to remove waste, a very similar concept to roller bottles.

Because the TideXcell^® system needs a large seeding density, how is scale-up possible?

Most of our customers choose roller bottles as the seeding source. In addition to CelCradle^® System and smaller scale of TideXcell system, the customer can use any other bioreactors for seed preparation such as Cell Factory or microcarrier systems. Two CelCradle^® bottles can be used for the seed preparation for TideXcell-002 and ten CelCradle^® bottles can be used to seed TideXcell-010. VacciXcell also provides automatic Cell Harvest System to harvest cells from TideXcell matrix vessel.

What is the disadvantage of the microcarrier system?

The commercial microcarrier system is a very difficult process. Though it is relatively easy in the small scale, scaling up to production scale is difficult and complex. The microcarrier system also incurs shear stress, foaming, and bubbling, which may all lead to cell death. Cell death not only decreases productivity but also increases downstream processing costs, due to increased cell debris.

VacciXcell’s tide motion system is simple and applies a similar principle as that of the roller bottles. Most of our customers use roller bottles and transfer to our technology directly without any problems. Those that tried to use the microcarrier system failed and eventually switched to the tide motion system.

How to seed in TideXcell?

Inoculation

Clamp the tube between Matrix Vessel and Mixing Vessel.
Prepare at least 3x109 cells in pre-warmed 800 ml culture medium per L matrices.
Disconnect the air tube inside TideXcell Culture Chamber with the Matrix Vessel. Move the Matrix Vessel to Biosafety cabinet.
Load the cells into the Matrix Vessel. Mix the cell containing culture medium together with the Matrixes by gently shaking the matrix vessel in a rotational manner by hand for 3 minutes.
Place the Matrix Vessel back to the Culture Chamber. Reconnect the air tube with the Matrix Vessel.
Remove the clamp on the tube between the Matrix Vessel and Mixing Vessel.

How to do BioNOC II^® sampling in pilot and industrial systems?

Currently, we only have closed sampling for single use system but we are developing a sampling mechanism for multiple use system. There is no need to open the matrix vessel during sampling. If you can imagine a “crane game” in arcades, the concept is similar to that one.

Report of system reject (failure or contamination rate)

For contamination issues, it highly depends on how the customer controls the culture environment and operation. Customers seldom report contamination issues to us which we think is due to the fact that most contamination issues are due to their process and practices and is rarely due to the equipment itself. Using our single-use system would make contamination risk almost 0.

In terms of component failure, we have had some cases but it is quite rare, roughly 1% and all these cases are after warranty periods. Strictly following the maintenance schedule is very important to minimize or reduce the risk of system failure.

What is BioNOC II^® made of?

BioNOC II^® matrix is made of 100% non-woven polyethylene terephthalate (PET) . It is resistant to chemicals and autoclavable.

Is BioNOC II^® applicable to all cell type?

BioNOC II^® is applicable to anchorage-dependent cells, and we have widespread, successful experiences on the cell culture.

The shelf life of BioNOC II^®?

The shelf life is four years without decay of hydrophilicity.

What is the surface area of BioNOC II^®?

There is approximately 2400 CM² of effective surface area. There are 5.5 g of BioNOC II^® per 100 ml of bed volume.

What brand of the serum-free medium can I use in CelCradle system?

CHO: CHO-S-SFM II (Gibco), CHO III A (Gibco), CHO-A-SFM (Gibco) , ExCELL 301 (JRH), HyQ PF-CHO (HyClone), HyQ-CCM 5 (HyClone), HyQ SFX-CHO (HyClone), IS-CHO-CD (IRVINE), proCHO4cdm
HEK-293: EX-CELL 293 (JRH), Pro293a-CDM (CAMBREX), HyQ SFM4 HEK293 (HyClone)
VERO: VP-SFM (Gibco), Plus VERO (VACCIXCELL), ICN-VERO (ICN), PEEK-1 (Biochrom),
Sf-9: SF-900 II (Gibco), EX-CELL 420 (JRH), EX-CELL 400

What does mean that BioNOC II^® is low bioburden?

The essence of a low bioburden in BioNOC II^® is that it does not encourage the growth of microorganism on the surface of the carriers, which may be a source of contamination.BioNOC II^® also has low lint content making it ideal for 3D cell culture.

How much media does one BioNOC II^® absorb?

25uL

What kind of microscope is needed to be able to observe cells inside BioNOC II^® carriers directly after staining?

Phase-contrast inverted microscope with a good depth of field function is recommended

Are all the bags sterile before shipment?

Yes. All products shipped to the customers are sterilized by gamma irradiation. An irradiation certificate is provided with the product.

What is the thickness of the customized bag and its related documentation?

The thickness depends on the material and purpose of the mixing bag. All bags are qualified in accordance with cGMP, USP Class VI Plastics, ASTM, and ISO.

Can I customize the medium bag?

Yes, you can. The medium bag is manufactured from US with ISO 13485;2003 certification and FDA registration, which has accepted a host of OEM from leaders of the medium bag in the market for years. Besides the comprehensive product line and validation documentation, we are confident to provide customization of bag products including film, tubing, connection, and filters, etc., to meet your specific needs and lean budget.

For your medium product, what would be the quantities available for order?

For cGMP grade, the minimum quantity is 25 kg. For the lab grade, the minimum order is 1 kg.

What is the advantage of SuperPlus compared with serum?

The purpose of SuperPlus is to reduce the use of serum in order to eliminate lot-to-lot variation and cost.

Are GlucCell glucose meter and test strips for the medical or diagnostic purpose?

No. The meter and strips are not for medical or diagnostic use, and are only for lab and bioprocess application.

We intend to use it for monitoring glucose consumption from a culture media containing bacteria.

GlucCell Glucose Monitoring System is designed specifically for animal cell culture in which the embedded calibration curves are made. Currently we don’t have sufficient data showing that it could be used for microorganism culture with these existing calibration curves. However, following steps can be done to validate your application:

Perform calibration with your existed culture medium with known glucose concentration by spiking known amount of glucose. If the initial glucose concentration of the medium is not adjustable, you have to prepare the medium by yourself by adding the component one by one and making several groups with different glucose concentration that could cover your test range.
Spin-down the culture medium to remove the solid and measure the glucose only from the supernatant.

Does GlucCell measure only glucose? If the medium contains other sugars, does GlucCell measure those as well?

By the enzyme reaction, GlucCell only measures specifically glucose but not other sugars such as fructose or sucrose.

What does error code E53 mean on the GlucCell glucose meter?

The reading E53 means that the detected glucose concentration is below the detectable range (20 mg/dL).

What is the difference between GlucCell glucose meter and other conventional monitors?

Generally, "glucose meter" is usually used by diabetic patients to monitor their blood sugar concentration. Therefore, it is difficult to obtain accurate data from cell culture medium by using this kind meter. On the contrary, GlucCell^® glucose meter is specifically designed to measure glucose concentration in cell culture medium. The accuracy is not affected by the type of media, whether with serum or serum-free, with cell or cell-free. GlucCell^® glucose meter is a palm size meter for glucose measurement particularly for animal cell culture.

Is it suggested to check that the meter is working correctly before each measurement?

The meter is designed to be calibration-free with our special technology to save customers’ time and money. However, especially in cGMP environment, customers have to do validation at the beginning to ensure the validity of the system. We encourage customers to do the validation by themselves and using the standard solution that they trust.

Why is the GlucCell^® system considered to be “calibration free”?

The system has been pre-calibrated and generates a CODE along with the Test Strips (The CODE is used as the calibration curve). Whenever users change to a new test strip batch/lot that has a code different from the code in your meter, the CODE needs to be reset to match the calibration curve.

How to do measurement verification/validation for GlucCell^®?

Customer can do validation by measuring the glucose from standard glucose solution and confirm the accuracy matches their specifications. VacciXcell does not provide standard glucose solution but but we can provide a protocol on how to prepare a standard glucose solution.

Will the measurement be accurate if the code number on the GlucCell^® is different from the one indicated in the test strip?

No, the measurement will be invalid. If the meter’s CODE is different from the test strips’ CODE, it means that the system will use the wrong calibration curve thus resulting in a wrong result.

Can the GlucCell^® be used to measure sugar as in the food and beverage industry? How about corn syrup?

GlucCell^® can only measure glucose and will not be able to measure other types of sugar

Which Tide Motion bioreactor products are cGMP compliant and/or FDA listed?

The CelCradle X® and TideXcell® systems are cGMP compliant. CelCradle, CelCradle X, CCX-SAH, CelShaker, CelXrocker, MinTide, TideXcell, TXLHS, and VXL Hybrid are ALL FDA-Listed products.

cGMP (FDA cGMP standard): CelCradle^® can comply to cGMP but it relies on the clinical sponsor or the appointed CMO through manual recording as part of their Process Qualification (PQ) and quality management release. The CMO or the clinical sponsor defines their own lot release for e.g. Esco Aster have a long list of quality control and quality assurance test, including cell-based assays and downstream purification methods to purify the product from our bioreactors. Used materials in all steps are fully traceable and uses serum/xeno-free chemically defined GMP media and cryopreservation.

See link: http://www.escoaster.com/white-paper-and-protocols/optimizing-large-scale-expansion-of-mesenchymal-stem-cells-in-3D-tide-motion-bioreactors/#content

https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfRL/rl.cfm?lid=618434&lpcd=KJF

How different is TideXcell^® and CelCradle^® in terms of GMP compliance?

GMP compliance: In terms of computer software, TideXcell^® and CelCradle^® X uses Siemens HMI/PLC that is FDA 21 CFR part 11 Compliant. CelCradle relies on external CO₂ incubator where it may not be cGMP/GMP. See information below for further elaboration.

Why is CelCradle^® not suitable for GMP? Is it because the system doesn’t have a recording device?

CelCradle^® can be GMP-compliant if it is applied under cGMP (FDA cGMP standard) provided that they use a cGMP CO₂ incubator that is connected to a Scada or Dcs system with eBatch records and eSignature.

If you are not using a cGMP incubator, manual logging is needed as part of Quality Management system and SOP or protocol. They should manually record in their production records what was adjusted in the incubator or cell manipulation, which person and at which time. This should also indicate which supervisor verified and who did the process QC to manually sample the BioNOC II during culture.

Disclaimer: cGMP/GMP (Iso 13485) may differ depending on your local authority who is final regulatory body, these suggestions from Esco as equipment manufacturer should not be mis used or mis represented. Your country local authority has final say in all cGMP/GMP matters.

Does the TideXcell equipment detail depend on compliance with the GMP standards?

Yes, the unit is controlled via Siemens HMI & PLC based monitoring and control system with simple, intuitive touchscreen. 21 cfr part 11 compliance, GAMP 5. VacciXcell Platform runs on WonderWare SCADA by default.

Note:

Dual redundant systems for critical components such as PLC and pumps are catered at optional costs.

The PLC-based monitoring and control system can be connected and configured to SCADA systems that have DeltaV and Pcs7 controls