LUOYANG FUDAU BIOTECH CO.LTD.--Cell Culture Consumables

  When culturing cells using a   cell factory , it is susceptible to various factors. Such as temperature, PH value, gas environment, etc., of which osmotic pressure is also a very important aspect. Cells in hypertonic or hypotonic solutions can shrink or swell and rupture immediately. Therefore, osmotic pressure is one of the important conditions for culturing cells in vitro. The maintenance of osmotic pressure of mammalian and other animal tissue cells in vitro is mainly related to NaCl, but the relationship of osmotic pressure of other dielectrics cannot be ignored. The osmotic pressure is proportional to the number of molecules and ions of the solute per unit volume of solvent. For this reason, it is very important to control the ion balance in the culture medium in a certain proportion and maintain normal osmotic pressure. This is not only to maintain cell tension, but also to regulate the metabolism of cells. Because extracellular ion transport and ion concentration change the tr

  Injection molding is a common processing method for various polymer materials, from plastic trinkets, toys, to auto parts, bottles, containers, and mobile phone casings. This processing method is used in all walks of life. The   cell factory   used in large-scale cell culture is also produced by the injection molding process. The injection molding process is to use an injection molding machine to heat, plasticize and melt the plastic, and then inject it into the cavity of the molding mold for molding. After cooling, the melt is solidified and then demolded. This processing method has the following advantages: 1. It is suitable for the production of complex precision products. The injection molding process can easily design complex plastic precision parts and assemblies. Compared to other techniques, injection molding will find features with very small tolerances. 2. Reduce waste: all the parisons made by injection can be blown into the blow mold, and no waste is generated during the

2. Materials and Methods 2.1. Maintenance of Huh7.5 Cells Huh7.5 cell cultures were maintained in  cell culture T flasks  in serum-containing medium (SCM): Dulbecco’s Modified Eagle Medium (DMEM) (Gibco, Paisley, UK) containing 4 mM GlutaMAX and supplemented with 10% fetal bovine serum (Sigma, St. Louis, MO, USA), 100 U/mL penicillin and 100 µg/mL streptomycin (Sigma). In T175 flasks (Nunc, Roskilde, Denmark) and triple layer culture flasks (Nunc), Huh7.5 cells were passaged every 2–3 days. All cell cultures were maintained at 37 °C and 5% CO2. 2.2. HCV Virus Stocks Two sequence-confirmed stocks of the genotype 5a recombinant SA13/JFH1core-NS5B [28], grown in monolayer Huh7.5 cell cultures in T flasks, were used in this study. A third passage virus stock with an infectivity titer of 5.8 log10 FFU/mL, grown under serum-containing (SC) conditions [39], was used to inoculate CelCradle™ cultures, shake flask cultures, and for virus characterization experiments. A fourth passage virus stock

  Cell factory   culture technology plays an important role in the fields of monoclonal antibody and vaccine preparation. When culturing cells, it will be affected by various factors, among which the cell inoculation density will also affect the cell culture process. When using cell factories for large-scale cell culture, the number of cell inoculations has a direct impact on the growth of cells. An appropriate inoculation density can promote better cell proliferation. Too high or too low inoculation density is not conducive to cell growth and proliferation. How to choose the appropriate inoculation concentration should be determined according to the cell metabolism, growth and reproduction speed and the needs of the work. Generally speaking, cells with strong metabolism and fast growth (such as tumor cells) should be inoculated at a lower concentration; while normal tissue cells grow slowly and their metabolism is not vigorous enough, the inoculation concentration may be higher; For e

  The   cell factory   is a multi-layered cell culture consumable. When culturing cells, different experimental operations are often performed with the help of various accessories. Platinum vulcanized silicone tube is a kind of supporting pipeline commonly used in the test process. Platinum vulcanized silicone tube is a silicone tube vulcanized with platinum catalyst. It is mainly used for the transfer of liquid in the cell factory. This kind of pipeline has the characteristics of odorless, non-toxic, acid and alkali resistance, ozone resistance and chemical corrosion resistance. Its processing technology is complex and the production cost is high. It is widely used in food and medical care, and belongs to the relatively high-grade level of silicone tubes. Platinum vulcanization is a two-component addition type vulcanizing agent. It has the characteristics of low vulcanization temperature and fast vulcanization speed. Compared with the traditional peroxide vulcanization system, it has

  A   cell factory   is a multi-layered cell culture consumable that is generally used for large-scale culture of adherent cells. The quality of consumables directly affects the growth state of cells, so how to control their quality? 1. Raw materials: The raw materials of the cell factory are generally polystyrene, and the raw materials must meet the USP Class VI standard. USPVI level medical equipment test is a relatively strict test for the application of plastic materials in the medical field and pipeline products in biopharmaceuticals. It is a non-clinical laboratory study that meets various experimental specifications. 2. Production environment: cells are particularly sensitive to the growth environment, so consumables are required to contain no substances harmful to cells such as endotoxin, which puts forward higher requirements for their production environment. Consumables must be produced in a dedicated 10,000-class clean room, and undergo rigorous verification (planktonic bact

  Materials and Methods 3.1. Shake Flask Cultivations Pre-cultures of MDCK.SUS2 suspension cells were grown in  shaker flasks  (125 mL polycarbonate Erlenmeyer flasks, #431143, Corning®, New York city, NY, USA) with 50 mL working volume (wv), in a Multitron Pro incubator (Infors HT, Bottmingen, Switzerland) at 37 °C and 5% CO2 atmosphere with a shaking frequency of 180 rpm. Cells were passaged every 3–4 days with a seeding density of 0.5–0.8 × 106 cells/mL. IAV A/Puerto Rico/8/34 (H1N1) seed virus was used for infection, generated in adherent MDCK cells (ECACC # 84121903). The seed virus had a titer of 1.1 × 109 TCID50/mL. Cells were cultivated in a chemically defined, protein-free and animal component-free medium, Smif8 (Smif8 PGD 2×, supplemented with 5 mM glutamine, and 8 mM pyruvate), specifically developed for the cultivation of suspension MDCK cells [95]. Growth and infection experiments for model validation were performed using 500 mL shaking flasks (#4113-0500, Nalgene™, Thermo

Cell culture technology is a commonly used technology in the fields of biopharmaceuticals, monoclonal antibodies, and cell therapy.   Cell factories   are mainly used for large-scale cell culture. The growth of cells is affected by many factors, of which temperature is a very important aspect. Under normal circumstances, the suitable temperature for in vitro culture of mammalian and avian cells is 37-38 °C. Too high or too low temperature will affect the growth of cells. The ability of cells to tolerate low temperature is stronger than that of heat resistance. At low temperature, the metabolic activity and nuclear division of cells are reduced. When the temperature is not lower than 0 °C, although it affects cell metabolism, it has no harmful effect; when the cells are placed at 25 to 35 °C, the cells can still survive and grow, but the speed is slowed down; Back to 37 ℃ culture cells can continue to grow. High temperature is not good for cell culture. Cells are cultured at 39-40°C for

  In large-scale cell culture,   cell factories   are commonly used consumables, which are mainly used for the culture of adherent cells. Cell growth requires a variety of nutrients, so what are they? 1. Culture medium The cell culture medium provides the cells in the cell factory with nutrients needed for growth, including carbohydrates, amino acids, inorganic salts, vitamins, etc. According to the nutritional needs of different cells, there are a variety of synthetic media to choose from, such as EBSS, Eagle, MEM, RPMll640, DMEM, etc. 2. Other additives In addition to the basic nutrients provided by various synthetic media, other components, such as serum and factors, need to be added according to different cells and different culture purposes. Serum provides important substances such as extracellular matrix, growth factors and transferrin, and fetal bovine serum is commonly used. The proportion of serum added should be determined according to different cells and different research p

The growth of cells has very strict requirements on the environment, temperature, pH value, etc., and the quality of cell consumables used in culturing cells will also affect the growth of cells.   Cell factories   are commonly used consumables for adherent cell culture, and there are four main factors that affect their quality. 1. Production raw materials: High-quality raw materials are the basis of high-quality products. The raw materials of the cell factory are polystyrene (PS) and must meet the requirements of USP Class VI. The word requirements are to test plastic materials in the medical field and pipeline products in biopharmaceuticals. The more stringent tests used in the field are non-clinical laboratory studies that comply with various experimental specifications. 2. Production environment: cells are particularly sensitive to the growth environment, so consumables are required to contain no substances harmful to cells such as endotoxin, which puts forward higher requirements

  In the large-scale cell culture,   cell factory   is an indispensable cell culture consumable. Cells have very strict requirements on the growth environment, so the consumables used need to be sterilized before they can be used. The commonly used sterilization method is irradiation sterilization. The use of this sterilization method has the following five advantages: 1. No pollution and no residue: Irradiation sterilization is different from chemical sterilization, which requires the addition of other components and does not produce radioactivity. 2. The sterilization effect is thorough: During the irradiation sterilization process of the cell factory, the gamma rays penetrate the goods in the irradiated cargo box, act on the microorganisms, directly or indirectly destroy the ribonucleic acid, proteins and enzymes of the microorganisms, thereby killing the microorganisms. Microorganisms, play the role of disinfection and sterilization. 3. Cold sterilization: Irradiation sterilization

  The   cell factory   is a cell culture consumable made of polystyrene (PS) raw materials. It adopts a multi-layer structure design. Common specifications include 1 layer, 2 layers, 5 layers, 10 layers, and 40 layers. Customize the number of layers as needed. So, what production process is used for this special structure of consumables? The injection molding process used in the cell factory mainly includes the following steps: 1. Pre-molding process: The pre-molding process is that the plastic material is heated, transported, compacted, sheared, mixed and homogenized in the barrel to transform the material from a glass state to a viscous fluid state, so as to meet the injection molding requirements. 2. Injection molding process: In the injection filling stage, the screw injects the melt in the storage chamber into the cavity through the nozzle, the mold runner and the gate under the thrust of the injection cylinder. 3. Compression and compression process: Continue injection molding to

Medium is an essential nutrient in the process of cell culture. When using a   cell factory   for cell culture, it usually involves a medium exchange operation, which can remove the metabolic waste produced by the cells during the growth process, and replenish the fresh medium to make the cells grow and reproduce better. To replace the medium in the cell factory, first prepare the required equipment: alcohol watering can filled with 75% medical sterilizing alcohol, PBS buffer, serum-containing medium, liquid transfer cover, transfer tube, waste tank, etc. Then put all the equipment into the ultra-clean bench and turn on the UV irradiation for sterilization. If the cultured cells are sensitive to temperature, you can put the medium bottle containing serum into a water bath at 37 °C, so that the temperature of the medium is at 37 °C, and then Transfer to the ultra-clean bench for UV sterilization, and UV irradiation is also required for about half an hour between cell cultures. After ult

  Cell factories   are mainly used for large-scale culture of adherent cells, such as hepatitis A vaccine, hepatitis B vaccine, chickenpox vaccine, etc. When culturing cells, sometimes we find that cells are difficult to adhere to the wall. What is the reason? First of all, there are many reasons for the phenomenon of non-adherence when the cells are cultured in the cell factory, which can be analyzed and solved from the following aspects: Excessive trypsin digestion: Trypsin helps cells to digest. If the digestion is excessive, the activity of cells will be greatly damaged, and cells will float. This can be controlled by shortening the digestion time or lowering the trypsin concentration. Mycoplasma contamination: cells are sensitive to the environment. If the operator does not pay attention to hygiene, or the working environment and experimental equipment are contaminated, it will lead to cross-contamination between cells and the phenomenon of non-adherence. If mycoplasma contaminati

  Cell factory   culture technology is a commonly used culture method for many large-scale cultures. When culturing cells, it will be affected by many factors, which hinder the normal growth and reproduction of cells. Radiation and ultrasonic waves are one of them. Visible light: The wavelength of visible light is 390~780nm. Various colored light can cause cell degeneration, prolong the interphase of nuclear division, and significantly reduce the ability of cells to attach to the wall. Therefore, when using the cell factory for in vitro culture of cells, direct sunlight should be avoided, and the culture should be carried out in the dark or stored for a short period of time as much as possible. Ultraviolet rays: Cells that are highly tolerant to weak UV rays do not change much, but sensitive cells are damaged. When the ultraviolet rays are strong, the isolated cells show that: complete mitosis cannot be performed; syneresis is increased during mitosis; cytoplasmic blebbing is reduced d

With the in-depth development of the field of life sciences, cell culture technology is widely used in many fields such as biopharmaceuticals, industrial vaccine production, and monoclonal antibodies. According to the different ways of cell growth, it is divided into two categories: adherent cells and suspension cells. So which cells can be cultured using   cell factories ? The cell factory is a multi-layer structure of cell culture consumables. Common specifications include 1 layer, 2 layers, 5 layers, 10 layers, 40 layers, etc. It is mainly used for the culture of adherent cells. The growth of adherent cells must have a support surface that can be attached, and cells can grow and reproduce on this surface by relying on the attachment factors secreted by themselves or provided in the culture medium. After the cells attach, they usually cover the culture surface within a few days and form a dense cell monolayer, such as Vero cells, HEK 293 cells, CAR-T cells, MRC5, CEF cells, porcine a

  Lentivirus is a kind of viral vector modified from human immunodeficiency virus (HIV), which is a kind of retrovirus. In in vitro experiments and in vivo experiments, lentivirus has become one of the commonly used forms of vectors for expressing exogenous genes or exogenous shRNA, and is being used more and more widely. In its mass production, the  cell factory   is a commonly used cell culture consumable. Lentiviral vectors can effectively integrate exogenous genes or exogenous shRNAs into the host chromosome, so as to achieve the effect of persistently expressing target sequences. In terms of infectivity, it can effectively infect neuronal cells, hepatocytes, cardiomyocytes, tumor cells, endothelial cells, stem cells and other types of cells, so as to achieve good gene therapy effects. For some difficult-to-transfect cells, such as primary cells, stem cells, undifferentiated cells, etc., the use of lentiviral vectors can greatly improve the transduction efficiency of the target gen

  With the continuous improvement and perfection of various processing techniques, cell culture consumables of various polymer materials are playing an increasingly important role in the field of life science research. A   cell factory   is a consumable made of polymer materials. According to its specifications, the application scenarios are also different. The cell factory is made of polystyrene (PS) raw materials. At present, the specifications on the market include 1 layer, 2 layers, 5 layers, 10 layers, and 40 layers. The products below 10 layers are mainly used for small laboratory cells. It is often used to replace cell culture flasks, which can save space, reduce pollution, and facilitate operation. Cell factories with more than 10 layers are generally used for large-scale cell culture, such as biopharmaceuticals, vaccine production and other fields, especially the industrial production of vaccines. Currently known vaccines that can be produced with this consumable include Japan

  The cell growth curve is a common method for determining the absolute growth number of cells, an important indicator for determining cell viability, and one of the basic parameters for the biological characteristics of cultured cells. The   cell factory   is mainly used for the culture of adherent cells, so how to draw the cell growth curve? Generally, after the cells are passaged, they are suspended for a short time and then adhere to the wall, and then spend different incubation periods of different lengths, that is, they enter the exponential growth phase of massive division. After the cells reach saturation density, they stop growing, enter the plateau phase, and then degenerate and die. In order to accurately describe the dynamic changes of cell numbers in the whole process, a typical growth curve can be divided into four parts: a slow-growing latent period, an exponential growth period with a large slope, a plateau-shaped flat-top period, and degeneration and decay. The growth