Autoclave, equipment sterilization

In order to prevent cross contamination when performing PCR it is important to separate your PCR reactions from previous DNA preparations. Try to perform PCR reactions in a bench area apart from other laboratory work, or if this is not possible use a tray to keep work isolated. It is also good practice to keep a separate set of pipettes for pre-PCR work. Keep your own stocks of PCR reagents, and in small aliquots frozen at -20°C. Use sterile techniques and autoclaved equipment. Exposure to UV irradiation in a UV DNA crosslinker (e.g., Strata-gene) can sterilize pipettes and other equipment from contaminating DNA. 

In a closed container such as a pressure cooker, a pressure greater than 1 atm can be obtained, which means that water boils at a temperature higher than 100 °C. Laboratories and hospitals use closed containers called autoclaves to sterilize laboratory and surgical equipment. Table 11.5 shows how the boihng point of water increases as pressure increases. 

Other heating equipment. Autoclave used for sterilization can be used to achieve superheating condition at 120°C. For higher temperature heating, a domestic pressure cooker, or a plastic steamer may be used. Some commercial laboratory pressure cookers have been designed for AR-IHC with controlled temperature. A water bath can be used to achieve lower temperature heating condition. 

The final product containers must also be pre-sterilized. This may be achieved by autoclaving or passage through special equipment that subjects the vials to a hot WFI rinse, followed by sterilizing dry heat and UV treatment. 

Polyarylsulfones offer materials with good thermal-oxidative stability, solvent resistance, creep resistance, and good hydrolytic stability. Their low flammability and smoke evolution encourage their use in aircraft and transportation applications. They hold up to repeated steam sterilization cycles and are used in a wide variety of medical applications such as life support parts, autoclavable tray systems, and surgical and laboratory equipment. Blow-molded products include suction bottles, surgical hollow shapes, and tissue culture bottles. PPS has a number of automotive uses including as an injection-molded fuel line coimector and as part of the fuel filter system. 

Three vacuum systems are commonly used in modern aseptic manufacturing facilities (1) house vacuum systems, (2) vacuum systems dedicated to lyophilization equipment, and (3) vacuum systems dedicated to autoclaves or other sterilization equipment. 

Component sterilizers steam sterilization process. A steam autoclave is used at ABC Pharmaceutical Industries to sterilize equipment and components used in aseptic processing. The following system description... 

Autoclave. (ABC 009) serial no. (provide number) autoclave is utilized for sterilization of solution buffer vessels, filters, filter housings, closures, filling machine dosing system, and support equipment. The unit has a chamber size of (width x length x height) mm and (provide volume) liter capacity with dual interlocking doors. 

Steam sterilization validation. All autoclave operations conform to the Master Validation Plan employed at ABC Pharmaceutical Industries. The autoclave has undergone installation qualification, operational qualification, and performance qualification. The autoclave is revalidated on an annual basis. The equipment steam sterilization cycle revalidation data for the autoclave are provided in validated archives. 

Medium for use in a process simulation test can be rendered sterile using either moist heat (autoclaving) or filtration. The method chosen depends on the availability of suitable equipment and the information desired from the study. 

At industrial scale, careful consideration of the materials of construction for the bioseparation equipment is vital to ensure that the product does not become contaminated, by rust, for example, and also to assure long plant life with good reliability to maximize throughput. Materials that were suitable on a laboratory or pilot scale may no longer be appropriate, where the process and mechanical demands on the equipment may be greater. For example, the plant could be located outside where there are greater extremes of temperature in summer and winter, or equipment may need to be steam sterilized in situ rather than being autoclaved. 

Select steam traps for the following five types of equipment (1) where the steam directly heats solid materials, as in autoclaves, retorts, and sterilizers (2) where the steam indirectly heats a liquid through a metallic surface, as in heat exchangers and kettles where the quantity of liquid heated is known and unknown (3) where the steam indirectly heats a solid through a metallic surface, as in dryers using cylinders or chambers and platen presses and (4) where the steam indirectly heats air through metallic surfaces, as in unit heaters, pipe coils, and radiators. 

Determine the condensate load. The first step in selecting a steam trap for any type of equipment is determination of the condensate load. Use the following general procedure. a. Solid materials in autoclaves, retorts, and sterilizers. How much condensate is formed when 2000 lb of solid material with a specific heat of 1.0 is processed in 15 min at 240°F by 25-psig steam from an initial temperature of 60°F in an insulated steel retort ... 

Glass equipment will normally be sterilized by dry-heat sterilization. Smaller equipment, like plastic tubes and rubber stoppers, can be sterilized by autoclaving. 

Aseptic Assembly. The first interventions performed are those that prepare the equipment for the aseptic process. This entails the removal of sterilized materials and equipment items from the autoclave and transfer to the location where the aseptic processing activities will be performed. This is ordinarily followed by the assembly/preparation of the equipment for the process. Aseptic assembly in which sterilized parts are removed from protective materials, installed and adjusted in preparation for the aseptic process are perhaps the most potentially invasive of all of the activities which must be performed. The operator must be meticulous in their execution of these tasks to prevent the inadvertent contamination of product contact surfaces. Strict adherence to the principles of aseptic technique described earlier is essential. These interventions are a necessary part of every aseptic activity, and it is common to identify the first containers filled as they may be more indicative of potential problems with the aseptic assembly. For this reason, the validation program should include process simulations that include containers filled immediately after the set-up of the equipment. 

Air removal is particularly important in porous and equipment loads, but is usually of little importance in the sterilization of aqueous pharmaceutical products. Air removal can be important to the specification of new autoclaves—those which are to be designated only for aqueous product sterilization have no need for the pumps and ancillary equipment required to pull deep vacuums. 

Data showing survival on some but not all of the Bis may be valuable in process development (particularly in the development of sterilization specifications and sterilizer parameters for porous and equipment loads), but would likely raise issues at regulatory inspection. The inference taken from having some survival could be that each item in the autoclave load is not being exposed to the same treatment. 

The experimental scheme for anaerobic decomposition is shown in Fig. 5-4. Exactly 1.5 g of each substrate was added to a modified 1 litre Sohngen flask and autoclaved at 120°C and 15 psi to ensure sterility, after which each flask was filled to capacity with a sterile inorganic nutrient medium and pH adjusted. Next 50 ml of a heterogeneous innoculum prepared from muds from a local lake was injected into each flask as "inoculates", while 50 ml. of sterile nutrient medium was used for control samples. Headspace C1-C4 hydrocarbons were measured prior to incubation to provide baseline concentrations. Minimum detection limits were 3 ppb on a volume basis using a high-sensitivity gas chromatograph equipped with a flame ionisation detector. Samples were incubated at 25°C and 36°C over a five week period. 

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