Biotechnology is process that use live organism to produce biological product. The interesting product can be secretion or metabolites by most of them are protein. It has many type of cell depend on the interesting product such as bacteria, yeast, mammalian cell, hybridoma cell, and transgenic animal. In this paper will focus on mammalian cell due to its ability of complexity production of protein such as monoclonal antibody. The mammalian cell is more sensitive than bacterial cell and yeast that it needs eligible design to maintain system. (4, 5)
Now Biopharmaceutical product trend to expend the market that the growth will be raising of 10% in 2006 to 15% in 2015 of total pharmaceutical product revenue on the world market and also it take 50% of new drug product application. This kind of process cost a lot of money. The biological products are getting expiration starting 2009 then cost will be the point of concern for biosimilar product. Contamination is source of cost in production line so the elimination of contamination will reduce cost. On the other hand, regulator also take attention to contamination of product that it has been appealed that the contamination is top ten of filing warning letter to manufacturer in the past ten years. (1, 2, 3, 15)
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Biotechnology consists of many steps on difference process but all of them are required on the same fundamental aspect of good manner to avoid the contamination. From the point of view, we are focusing to minimize the opportunity of contamination that has some basic step will help to reduce the risk of contamination. The design of process has to be tight control on understanding the process. Starting with the raw materials, manufacturer has to find reliable vender that will provide good quality materials for the process. The absence of virus, mycoplasma, and endotoxin are key determine for material use. The equipments have to be sanitary design and reliable including appropriate size and maintenance. It also has capability to clean in place and steam in place because the cleaning of instrument is also the big concern in this area. After sterilization process, the instrument has to conduct sterile test. Operator that is the one of major source of contamination has to be trained to reduce the process failure. The proper of cloth will also prevent the contamination. Moreover the appropriate controlled environment including designing of plant, material flow, personnel flow, classification room and air handling system will be main way to prevent contamination. The sampling process has to conduct under designed condition to prevent accidental contamination. (5, 7, 15)
On the other hand, the risks of contaminations that they are introduced by different fundamental requirement are variable on each step. The Inoculation is process that produce sufficient cell to production plant. By the rule of thrum, it grow gradually from 20 liters to 200 liters and then to 2000 liters. Cells can grow in two difference type of culture system that they are surface and suspended system. The suspending system is more efficient than adhered system and wildly use in industrial. This step is potential to introduce risk from open transfer; such as adding nutrient, sampling, and changing the vessel, to the process that will be the major cause of contamination so it has to use appropriate behaved and technology. (3, 5, 17)
The adequate quantity of cell will transfer to bioreactor that it refer as growing mammalian cell tank to produce the product. It has two types of bioreactor process that they are batch and continuous process. Overall, the continuous process seem like have more efficient and advantages but one of its disadvantages is the process might have more risk of contamination than batch process due to the relative brief cultivation period. Nowadays, disposable is introduced to bioreactor system to minimize risk of contamination that it is a new choice for manufacturer to carry out the quality product. (5,6, 17)
The next step will be cell separation or harvesting that this process will separate the interesting product from cells. The interesting product can divide for two types that are intracellular and extracellular product that will affect the method and equipment. The intracellular product has to do cell disruption process to release the product to media. It has many methods to disrupt cells such as using of enzyme, osmotic pressure change, sonication, and freezing-thawing. The choosing of method will be influenced by many factors but industrial scale commonly uses mechanical technique such as high-pressure homogenizer and grinding in ball mill. The point of concern is the design of equipment that has to tightly seal to be close system and allow to do CIP and SIP. Extracellular product required the gentle process of cell separation to minimize the cell damage. Cell separation process can conduct via centrifugation or filtration method depends on type of cell, product, scale of production, etc. The centrifugation system has many models that will properly use in different property of product. The most common use in large scale is disk-stack centrifugation that has capability to process continuous mode. This mode is concerned about cleaning of machine. The filtration is old and acceptance process that its system consists of two types of filtration is dead end and tangential flow filtration (TFF). Harvesting step normally use 0.2 micrometer of diameter membrane size. In the production scale, TFF is more common use and has many advantages over dead end filtration. Before and after the process, both of filter type need to perform integrity test to make sure that membrane filter doesn't have any defect. The multiple use of membrane filter has to have proper clean and storage in preservative solution to avoid microbial growth that will depend on the type of membrane. Another concern is gel polarization that is concentration of component on surface membrane which will resist flux of the system. However, it can prevent by process parameters such as flow rate. After the separation, the product may conduct the ultrafiltration and diafiltration that it will prepare the product to purification process. Ultrafiltration is common use to concentrate the intermediate product by the normal pore size is 0.001-0.02 micron. The filter will retain the product and allow the salt or buffer pass thru membrane. Diafiltration is process that uses for buffer exchange by not changing the volume of intermediate product. The contamination concern of both ultrafitration and diafiltration will be the same as filtration on harvesting step. (4, 8, 9, 10, 11, 12, 13, 15, 16)
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Viral reduction is process that will eliminate the virus from the intermediate product before go to purification process. The source of virus can split into two types are the viral that come from the master cell bank (MCB) and adventitious virus that is introduce during the process. The viral that come from the MCB can be introduced from either deviation of cell line, using virus to create the cell line, using the contamination material, and contamination during processing. The adventitious virus can be brought to the process by using of contaminated biological reagents, reagent, excipient, using of a virus for the induction of expression of specific genes encoding a desired protein, using of a contaminated reagent, and contamination during cell and medium handling. The viral containing has to monitoring and controlled during the process. It has many ways to do viral clearance such as pH inactivation (e.g. pH 4.0), solvent/detergent inactivation (e.g. tri-n-butyl phosphate [TNBP]/Triton X-100), chemical inactivation (e.g. caprylic acid), ethanol precipitation (e.g. 40% ethanol precipitation), nano-filtration (e.g. dead end filtration and tangential flow filtration), affinity chromatography (e.g. Protein A), gel filtration chromatography, ion exchange chromatography, gamma-irradiation, Precipitation (e.g. ethylene glycol). The regulation requires the difference type of viral reduction method to conduct for one product. (18, 19)
Purification is process that gets rid of unwanted substance from product. This step can divide to three phases that are capture, intermediate purification, and polishing. The first step is capture that will perform to isolate, concentrate, and stabilize the product. The Intermediate purification is process that will remove most of bulk impurity such as other protein, endotoxin, virus, etc. Then the last step is polishing that will eliminate the trace of impurity from the product. To select the technique using for these steps, the manufacturer may define the purity requirement, and nature of contamination component and interesting protein. Chromatography is used to purify the product. It has four techniques that are ion exchange, gel filtration, hydrophobic interaction, and affinity technique. The points of contamination are the internal surface of column, packing process, cleaning process of column and resin used. The reuse of gel has to be relied on acknowledge with limitation. (4, 20)
After purification process, the bulk product has to have proper storage that will reduce degradation and microbial growth. Protein is unstable molecule that it has different conditions depend on shelf-life requirement of protein. It can store at 4°C, -20°C, and (-20°C)-(-80°C). At 4°C, it can stay for few week up to one month that this condition requires anti-microbial and some additive. At -20°C, it can stay up to one year that may require anti-microbial. This condition is the most suitable for antibody but it need some additive to extend the shelf-life. At 80°C, it can stay for several years but it will not allow to frequency freeze-thaw the protein that can cause protein damage. Each condition will require different additive depend on study and natural or product. The additive might be cryoprotectant, protease inhibitor, anti-microbial, reducing agent, and chelating agent. Moreover the dilute protein is found with more lost of degradation, adding the bovine serum albumin (BSA) 0.1-0.5% can reduce this influence. This step will concern of material use that has to be proper grade for bioprocess. (21)
Cell culture contamination:
Cell culture contamination continues to be a major problem at the basic research bench as well as for bioproduct manufacturers. Contamination is the only thing which endangers their use as reliable reagents.. Each cell culture system is unique. Usually contamination is chronic because it is unseen, unrecognized and often benign within a given cell culture system. Nearly every culture laboratory has experienced the loss of cell cultures at some point. Culture contamination can be happened in various types, but always has an adverse effect on the quality of the cell.
Cell culture contamination is any nonliving substance that causes undesirable and adverse effects. It is important to note that even essential nutrients can become contaminants in high doses, making it difficult to avoid unwanted changes. We can categorize cell culture contamination in two groups,
1) Chemical contamination
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2) Biological contamination
4.1) Chemical Contamination
Chemical contamination is best described as the presence of any nonliving substance that results in undesirable effects on the culture system. Sometime essential nutrients become toxic at high enough concentration not only its toxicity, even hormones and other growth factors found in serum can cause changes which are harmful to cultures. Here we discuss some of the contaminants which cause contamination in culture.
Media and Sera:
The majority of chemical contaminants found in cell culture media, come either from the reagents or water used to make culture or the additives like sera which is used to supplement them. Mistakes in media preparation, protocols, reading reagent bottle labels, or weighing reagents are other common sources of chemical contamination.
To prevent contamination by this reagents and sera they should always be of the highest quality and purity and must be properly stored to prevent deterioration. Ideally, they should be either certified for cell culture use by their manufacturer or evaluated by the researcher before use. Especially for sera, careful testing of sera before purchase, or switching to serum-free media can avoid these problems. Serum proteins have the ability to bind substantial quantities of chemical contaminants, especially heavy metals that may have entered the culture system from other sources, rendering them less toxic. As a result, switching from serum-containing medium to a serum-free system can unmask these toxic chemical contaminants, exposing the cells to their adverse effects.
Types and Sources of Potential Chemical Contaminants:
Metal ions, endotoxins, and other impurities in media, sera, and water
Plasticizers in plastic tubing and storage bottles
Free radicals generated in media by the photo activation of tryptophan, riboflavin or HEPES exposed to fluorescent light
Deposits on glassware, pipettes, instruments etc., left by disinfectants or detergents
Compounds in autoclave water, residues from aluminum foil or paper
Residues from germicides or pesticides used to disinfect incubators, equipment, and labs
Impurities in gases used in CO2 incubators
The water used for making media and washing glassware is a frequent source of chemical contamination and requires special care to ensure its quality.
For prevention of contamination by water, double or triple glass distillation was considered to be the best source of high quality water for cell culture media and solutions. Newer purification systems combining reverse osmosis, ion exchange and ultra filtration are capable of removing trace metals, dissolved organic compounds and endotoxins are increasingly popular.
Endotoxins, the lipopolysaccaride-containing by-products of gram negative bacteria, are another source of chemical contaminants in cell culture systems. Endotoxins are commonly found in water, sera and some culture additives and can be readily quantified using the Limulus Amebocyte Lysate assay (LAL).
These highly biologically reactive molecules have major influences in vivo on humoral and cellular systems. Studies of endotoxins using in vitro systems have shown that they may affect the growth or performance of cultures and are a significant source of experimental variability. In the past, sera have been a major source of endotoxins in cell cultures.
Efforts must be made to keep endotoxin levels in culture systems as low as possible.
As improved endotoxin assays (LAL) led to an increased awareness of the potential cell culture problems associated with endotoxins, most manufacturers have significantly reduced levels in sera by handling the raw products under aseptic conditions. Poorly maintained water systems, especially systems using ion exchange resins, can harbor significant levels of endotoxin-producing bacteria and may need to be tested if endotoxin problems are suspected or discovered in the cultures.
Media stored in glass or plastic bottles that have previously contained solutions of heavy metals or organic compounds, such as electron microscopy stains, solvents and pesticides, can be another source of contamination. The contaminants can be adsorbed onto the surface of the bottle or its cap during storage of the original solution. If during the washing process they are only partially removed, then once in contact with culture media they may slowly leach back into solution.
A fluorescent light is an overlooked source of chemical contamination. It produce contamination from the exposure of media containing HEPES (N-[2-hydroxylethyl] piperazine-N'-[2-ethanesul-fonic acid]) -which is an organic buffer used to supplement bicarbonate-based buffers, riboflavin or tryptophan to normal fluorescent lighting. These media components can be photo activated producing hydrogen peroxide and free radicals that are toxic to cells; if the exposure is longer, greater the toxicity. When fluorescent light exposure is more extensive will lead to a gradual deterioration in the quality of the media.
The incubator, often considered a major source of biological contamination, can also be a source of chemical contamination. The gas mixtures perfused through some incubators may contain toxic impurities, especially oils or other gases such as carbon monoxide that may have been previously used in the same storage cylinder or tank. This problem is very rare in medical grade gases, but more common in the less expensive industrial grade gas mixtures. Care must also be taken when installing new cylinders to make sure the correct gas cylinder is used. Other potential chemical contaminants are the toxic, volatile residues left behind after cleaning and disinfecting incubators.
Disinfectant odors should not be detectable in a freshly cleaned incubator when it is placed back into use.
4.2) Biological Contamination:
Biological contaminants can be subdivided into two groups based on the difficulty of detecting them in cultures: those that are usually easy to detect - bacteria, molds and yeast. Those are more difficult to detect, and as a result potentially more serious culture problems, - viruses, protozoa, insects, mycoplasmas and other cell lines. Ultimately, it is the length of time that a culture contaminant escapes detection that will determine the extent of damage it creates in a laboratory or research project.
Bacteria, Molds, and Yeasts:
(Contamination through bacteria) (Yeasts appear as round or ovoid particles.)
Bacteria, molds and yeasts are found virtually everywhere and are able to quickly colonize and flourish in the rich and relatively undefended environment provided by cell cultures. Because of their size and fast growth rates, these microbes are the most commonly encountered cell culture contaminants. In the absence of antibiotics, microbes can usually be readily detected in a culture within a few days of becoming contaminated, either by direct microscopic observation or by the effects they have on the culture like pH shifts, turbidity, and cell destruction. But if we used antibiotics routinely, organisms may develop into slow growing.
Viruses also causes the contamination and we can't detect them in a culture due to their extremely small .Their small size also makes them very difficult to remove from media, sera, and other solutions of biological origin. Although viruses may be more common in cell cultures than many researchers realize, they are usually not a serious problem unless they have cytopathic or other adverse effects on the cultures since cytopathic viruses usually destroy the cultures they infect, they tend to be self-limiting. Thus, when cultures self-destruct for no apparent reason and no evidence of common biological contaminants can be found, cryptic viruses are often blamed, they are perfect culprits, unseen and undetectable; guilty without direct evidence. This is unfortunate, since the real cause of this culture destruction may be something else, possibly mycoplasma or a chemical contaminant, and as a result will go undetected to become a more serious problem.
(Contamination through virus)
Both parasitic and free-living, single-celled protozoa, such as amoebas, have occasionally been identified as cell culture contaminants. Usually of soil origin, amoebas can form spores and are readily isolated from the air, occasionally from tissues, as well as throat and nose swabs of laboratory personnel. They can cause cytopathic effects resembling viral damage and completely destroy a culture within ten days. Because of their slow growth and morphological similarities to cultured cells, amoebas are somewhat difficult to detect in culture, unless already suspected as contaminants.
Insects and arachnids commonly found in laboratory areas, especially flies, ants, cockroaches and mites, can both be culture contaminants as well as important sources of microbial contamination. Warm rooms are common sites of infestation. By wandering in and out of culture vessels and sterile supplies as they search for food or shelter, they can randomly spread a variety of microbial contaminants. Occasionally they are detected by the trail of "footprints" (microbial colonies) they leave behind on agar plates, but usually they don't leave any visible signs of their visit other than random microbial contamination.
Mycoplasmas were first detected in cell cultures by Robinson and coworkers in 1956. In addition, they discovered that the other cell lines currently in use in their laboratory were also infected with mycoplasma, a common characteristic of mycoplasma contamination. Based on mycoplasma testing done by the FDA, ATCC, and two major cell culture testing companies, at least 11 to 15% of the cell cultures in the United States are currently infected by mycoplasmas. Since many of these cultures were from laboratories that test routinely for mycoplasmas, the actual rates are probably higher in the many laboratories that do not test at all. In Europe, mycoplasma contamination levels were found to be even higher: over 25% of 1949 cell cultures from the Netherlands and 37% of 327 cultures from former Czechoslovakia were positive (14). The Czechoslovakia study had an interesting, but typical finding: 100% of the cultures from labs without mycoplasma testing programs were contaminated, but only 2% of the cultures from labs that tested regularly. Other countries may be worse: 65% of the cultures in Argentina and 80% in Japan were reported to be contaminated by mycoplasma in other studies (11). Unfortunately, mycoplasmas are not relatively benign culture contaminants but have the ability to alter their host culture's cell function, growth, metabolism, morphology, attachment, membranes, virus propagation and yield, interferon induction and yield, cause chromosomal aberrations and damage, and cytopathic effects including plaque formation .
What gives mycoplasmas this ability to readily infect so many cultures?
Three basic characteristics:
a) These simple, bacteria-like microbes are the smallest self-replicating organism known (0.3 to 0.8 Âµm in diameter),
b) They lack a cell wall, and
c) They are fastidious in their growth requirements.
Their small size and lack of a cell wall allow mycoplasmas to grow to very high densities in cell culture (107 to 109 colony forming units/mL are common) often without any visible signs of contamination - no turbidity, pH changes or even cytopathic effects. Even careful microscopic observation of live cell cultures cannot detect their presence. These same two characteristics also make mycoplasmas, like viruses, very difficult to completely remove from sera by membrane filtration. In addition, their fastidious growth requirements) make them very difficult to grow and detect using standard microbiological cultivation methods. Thus, these three simple characteristics, combined with their ability to alter virtually every cellular function and parameter, make mycoplasmas the most serious, widespread, and devastating culture contaminants.
Mycoplasmas have been described as the "crabgrass" of cell cultures, but this is too benign a description for what are the most significant and widespread cell culture contaminants in the world.
4.3) How Can Cell Culture Contamination Be Controlled?
Cell cultures can be managed to reduce both the frequency and seriousness of culture related problems, especially contamination. Lack of basic culture management procedures, especially in larger laboratories, frequently leads to long term problems, making contamination more likely for everyone. One solution is to actively manage your cultures to reduce problems and if necessary set up a program for use in your laboratory. This program should be designed to meet the needs of your specific working conditions and be based on the nature of your past cell culture problems; it can be very simple and informal, or more structured if required.
The first step in managing cultures is to determine the extent and nature of the culture losses in your lab. Everyone in the laboratory should keep an accurate record for a month or more of all problems, no matter how minor or insignificant, that result in the loss of any cultures.
These problems may not only be contamination related but can also be from other causes such as incubator or equipment failures. Next, review the problems as a group to determine their nature, seriousness and frequency. The group's findings may be surprising: what were thought to be individual and minor random occurrences of contamination often turn out to have a pattern and be more extensive than any individual realized.
This problem sharing is often a painful process, but remembers the goal is not to place blame but to appreciate the extent and nature of the problems confronting the laboratory. A critical part of this process understands the seriousness and actual costs of culture loss; placing a dollar value on these losses is often required before the full extent of the losses can be appreciated.
Once the nature and consequences of the problems in the laboratory are better understood, the need for a management system, if necessary, can be determined. Basic problem solving tools can be used to help identify the source of problems; changes to minimize or prevent the problems from reoccurring can then be implemented.
The following suggestions, concepts and strategies, combined with basic management techniques, can be used to reduce and control contamination. These may require modification to fit your own needs and situation.
Steps for Reducing Contamination Problems:
Use good aseptic techniques
Keep the laboratory clean
Routinely monitor for contamination
Use frozen cell repository strategically
Use antibiotics sparingly if at all
The following suggestions are recommended to reduce the probability of contamination:
Make it more difficult for microorganisms to gain entry by using sealed culture vessels whenever possible, especially for long term cultures. The multiple well plates can be sealed with labeling tape or placed in sealable bags, 35 and 60 mm dishes can be placed inside 150 or 245 mm dishes. Use vented cap flasks (See Figure 7) whenever possible. These have hydrophobic filter membranes that allow sterile gas exchange but prevent the passage of microorganisms or liquids.
Avoid pouring media from cell culture flasks. or sterile bottles by using 50 or 100 mL pipettes or aseptic tubing sets to transfer larger volumes. Using a disposable aspirator tube and vacuum pump is an economical way to quickly and safely remove medium from cultures. A drop of medium remaining on the vessel's threads after pouring can form a liquid bridge when the cap is replaced providing a means of entry for bacteria yeasts and molds. If pouring cannot be avoided, carefully remove any traces of media from the neck of the vessel with a sterile gauze or alcohol pad.
Always carry unsealed cultures in trays or boxes to minimize contact with airborne contaminants. Square 245 mm dishes are excellent carriers for 384 and 96 well plates as well as for 35mm and 60 mm dishes.
Do not use the hood as a storage area. Storing unnecessary boxes, bottles, cans etc. in the hood, besides adding to the bioburden, disrupts the airflow patterns.
Never mouth pipette. Besides the risk of injury to laboratory personnel, mouth pipetting has been implicated as the likely source of human mycoplasma often found in cell cultures.
Use clean lab coats or other protective clothing to protect against shedding contaminants from skin or clothes. Their use should be restricted to the cell culture area to avoid exposure to dirt and dust from other areas.
Work with only one cell line at a time in the hood, and always use separate bottles of media, solutions, etc. for each cell line to avoid possible cross-contamination. Use disinfectant to wipe down the hood's work surfaces between cell lines.
Emerging technologies or procedure to improve mfg yield:
5.1) Clean in Place (CIP)
CIP system designed for automatic cleaning and disinfecting without major disassembly and assembly work. Basically, a well designed CIP system will enable the operator to clean one part of the plant while other areas continue to produce product. Furthermore, this system will not only save money in terms of higher plant utilization but also due to savings in CIP liquid (by recycling cleaning solutions), water (the system is designed to use the optimum quantity of water) and man-hours. The cleaning can be carried out with automated or manual. CIP is important to biopharmaceutical industry in which the processing must take place in a hygienic or aseptic environment.
Overall advantages of CIP
Reproductive washing result
Labor and time saving
Water and cleaning agent saving
Other benefits of a well designed CIP plant includes operator safety in which operators are not required to enter tanks and vessels to clean them and potent cleaning materials do not need to be handled by them.
Latest CIP System Design and Devices:
Multi-Tank (Detergent and Rinse Reuse): Sani-Matic CIP Systems
This system allows the reuse of wash solution and rinse water. Because of that, cost of water and waste-water can be reduced. It has three tanks which are detergent tank, rinse tank, and recovery tank.
Those three tanks are connected with simple piping design, the circulation velocity of cleaning agent and rinse water is operated using a supply pump located at the bottom of the skid.
Besides that, SIP piping design also is included in the system which probably works for steam sterilization and temperature control. The unique design will allow one additional tanks to be put in the skid (probably acid reuse application).
Dual- Operating (Multi-Circuit and Reuse): Sani-Matic CIP Systems
This system is like any other CIP system skid; it has three tanks which are detergent tank, rinse tank, and recovery tank. Those three tanks are connected with two different piping circuits (CIP supply #1 and CIP supply #2) and operated using two different pumps (Supply pump #1 and Supply pump #2).
Even though there are two different piping circuits, both of the circuits are operated with one centralized control panel. Besides that, due to its small size, it saves substantial cost and space saving advantages.
Like the previous system, this CIP skid also allows the reuse of wash solution and rinse water. This will help to reduce cost of water and waste-water. Furthermore, SIP piping circuit is includes in the system which probably works for steam sterilization and temperature control. Dual-Operating
CIP Device (Double Flush Mount Nozzle)
Another important device for CIP is double flush mount nozzle. Made using stainless steel, it is designed with fewer components than any other spray device. Fully retractable spray head allow the device to remain in the fermentor or bioreactor without disturbing inoculums growth.
It simple design allows very simple installation and maintenance (easy to assemble and removal). This helps to reduce CIP time. Most importantly, large spray range cleans large surface area (e.g. bioreactor).
Double flush mount nozzle
5.2) Transfer System:
Transfer system move raw materials (e.g. bulk material, powders, liquids, packaging material), parts and components, and everything from one place to another. This system is very important since raw materials need to be handled and operated in complete aseptic condition. Furthermore, it also helps to reduce contamination problem especially during media preparation process. Below are two equipments which are used in transfer system.
Â Rapid Transfer Port (RTP)
Rapid Transfer Ports (RTP) provides a safe, efficient and time saving method to put and remove material without contaminating the plant.Â RTP come in a variety of sizes ranging from small to very large cart. RTP is either made in stainless steel or plastic.Â Exterior of the sealed RTP being mated to the exterior of the BSC
Â The RTP has a beta flange lid exterior that mates with alpha flange door exterior and attached to the cabinet (e.g. Class III Biological Safety Cabinet (BSC)). The lid and door are rotated and sealed and must form an interlock before the door inside the BSC can be opened.
The exposed surface is only the interior of the RTP cylinder to air and materials inside the BSC.Â The process is reversed to uncouple the RTP.Â The contents can be stored or the RTP can be opened with an Allen key inside the cabinet where the materials are removed and the interior of the RTP is surface decontaminated for reuse. Rapid Transfer Port (RTP)
Butterfly Valve (Type 567 Butterfly Valve) type567.jpg
Type 567 Butterfly valve is designed with double eccentric operating principle. Its individual component is merely design to optimize particular function. With two O-ring and special bi-directional seal, the valve provides good protection against external leakage. Besides that, the profile seal is designed to match the double eccentric design. Quick opening disk allows the medium to flow freely between the disk and seal and flushing any accumulated particle out.
On top of that, the geometry of the disk gives the dirt particle nowhere to go. There is no deposit can form between seal and vault disk because there is no space contact between those components. Even with double eccentric design but still the valve maintain its bi-directional pressure rating of 10 bars. Pressure pulses are absorbed better due to seal profile design which increases surface life. Together With electric position indicator, the operator will always know which valves is open or closed and this give a clear overview of system at all time.
5.3) Disposable System:
A case study made by, Mark Fuller and Helene Pora suggest that the cause of product degradation in mainly due to formulation sensitivity to stainless steel to oxygen. Besides that, bio-molecules, which are highly sensitive to pH, temperature, and organic solvents, must be manufactured in processes that are quite 'gently' than the pharmaceutical. This gives the opportunity for the polymer equipments to replace stainless steel or glass in most of upstream and downstream process, material or product storage and handling.
The main reasons why manufacturer choose single-use system are:
Lower capital expenditure
Reduced validation costs
Shorter development time for new facilities
Specific Benefit Gains from Single-Use System:
Improved product quality
Substantially reduced process times
Lower operating costs
Below are two single-use models used in biopharmaceutical industries:
Mobius CellReady 3L Bioreator
Mobius CellReady 3L Bioreactor is a single-use stirred tank bioreactor which is suitable for bench-scale cell culture. It is pre-assembled and gamma irradiated, which significantly reduce turnaround times typically associated with glass bioreactors. It ensures maximum operational flexibility, with tubing and vent filter, two sparging options, and compatibility with most standard bioreactor controller configurations. Besides that, it has the additional function integrated side sampling, addition and drain ports.
Vessel Diameter (inner)
Vessel, shaft and base
Headplate, Thermowell and Impeller
O-rings and seals
PG 13.5 threaded (2)
MobiusÂ® Storage and Handling System
Mobius Handling and Storage Systems consist of a series of polyethylene drums, stainless steel bins and single-use assemblies, it is designed to reduce operators' error and improve process efficiency. This system will provide excellent flexibility, mobility and support for single-use technologies.
Mobius polyethylene drums and bins are designed for media and buffer solutions storage in biopharmaceutical process. The drums can accommodate top and bottom container port arrangements, including single-use filter assemblies for sterile applications.
Mobius Stainless Steel bins feature a single large door for an operator access and to position the process container inside the bin. It is constructed with a 304/304 L stainless steel-grade. The tray has a convenient slide-out capability, a positive pitch to make sure complete drainage, and space to hold disposable components. The bins are both pallet jack and fork lift compatible.
Mobius Standard Assemblies
(3-D, Drum-style Single-use Containers)
Mobius standard drum assemblies can be use for various such as storage and processing of sterile fluids or buffer. The assemblies are animal-derived component free and delivered pre-sterilized by gamma irradiation. Available in volumes from 50 L to 200 L, they are specifically designed to fit Mobius polyethylene drums but work just as well with other commercially available plastic storage drums.
Unique Registration System
The Mobius stainless steel bin design includes a patent-pending registration system. This system is comprised of a disposable plate and a corresponding opening at the bottom of the bin. Provided with every Mobius process container assembly, the plate's unique keying mechanism permits only one-way placement of the container inside the bin. Each registration plate can accommodate up to three bottom ports, allowing operators to use any process containers inside the bin.
Easy Process Container Filling
Disposable process containers are built with an easy-folding design that eliminates the need for operator or mechanical assistance during the filling process for bin sizes of up to 1,500 L.