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    Add as Friendsop for sterilization

    by: ravikishore

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    1 : Standard operation procedure for sterilization _RAVIKISHORE.K.V
    2 : DEFINITION Sterilization is defined as the destruction or removal of all microorganisms and their spores. Sterilization is the part of daily routine of microbiological laboratories etc.,
    3 : Methods of sterilization Physical Methods Chemical Methods Sunlight Dry heat Moist heat UV heat Ionizing radiation Filtering through a bacteria – proof filter Liquid sterilizing agents Gaseous sterilizing agents
    4 : Moist heat sterilization by autoclave Materials which are not stable at high temperature can be sterilized by the autoclave using steam as medium.
    5 :         Standard operating procedures Sop NO#:1000.64 date issued: date revised: pages: Title: Autoclave sterilization Scope: Proper procedure for Sterilization,
    6 : 1.Purpose : To outline the proper procedure for sterilization, refers to the complete killing of all living Micro-organisms. 2.Responsibility: it is the responsibility of the facility manager / other appointed personnel to ensure all the sterilization equipment like autoclave, Hot air Oven is appropriately maintained in good condition and available for research personnel as requested.
    7 : Operation of Autoclave Remove the lid of Autoclave and clean the inside body with distilled water and benzalkonium chloride solution before operation. Clean carriage and container of sterilizing chamber with vacuum and then distilled water. Check working of autoclave thermograph. Ensure that there is sufficient water inside the chamber. contd..
    8 : Items to be sterilized should be placed into the autoclave . Small items to be sterilized should be bagged or wrapped with heat resistance plastic / paper / cloth In Autoclave sterilization time and temperatures are dependent on the composition and nature of items being sterilized. Minimum time and temperature are usually 15mins @ 250 F or 270 F for 5 Mins contd..
    9 : Autoclaving Of BIOHAZARDOUS WASTE Bags / containers should not be overfilled half to three quarter full is recommended. Add water prior to sealing the container to aid the generation of steam. Do not add water if there is a chance of bio-hazardous material may splash out of the container. Seal container.
    10 : Sterilized supplies must be labelled with date autoclaved / processed. Plug the power cord and the main switch to “START” Close the door and lock it by turning the handle clockwise. Set the temperature @ 250 F / 121 c with thermostat knob. This is the temperarure most often seed for sterilization.
    11 : Check on the unit after about 25 min to be ensure that it is reaching temp /pressure. Small burst of steam may be evident from the water reservoir during the run, that is normal and necessary. When the timer reaches zero the run is complete. Turn off the autoclave, let it cool. Slowly open the air and steam discharger taps after the pressure gauge has reached zero.
    12 : Before opening the door, the personnel should wear heat resistance gloves. After few minutes the autoclave material can be removed. Items will still be very hot for a while and are not used until cool to the touch.
    13 : Documentation
    14 : CALIBRATION OF INSTRUMENTS INSTALLED ON AUTOCLAVE Pressure/ vacuum gauge –calibration Calibration of thermo couple temperature measuring device for autoclave Calibration of time-temperature recorder
    15 : Maintenance Frequency of check Parts to be checked Once a week Door locking and clamping mechanisms chamber door seal & safety devices Indicating lights & controls chamber drain thermometer Air filter Air tightness of chamber
    16 : Once in three months Parts to be checked Once in six months Time temperature recorder and thermograph. Water and steam ejector Steam valves to chamber Vacuum & pressure gauges Electric motors & Electrical Connections.
    17 : DRY HEAT STERILIZATION HOT AIR OVEN: a hot air oven is electrically operated and should be equipped with fan to ensure the uniform temperature inside the oven. The required temperature is 160 c / hour. Equipment: Double door dry heat sterilizer with mounted HEPA filter module.
    18 : Hot air oven
    19 : Procedure Remove the lock and open the pressure lock door by turning the handle anticlockwise . Unload the carriage on the trolley provided. Check working of the alarm system when the door is opened, if not, report to the supervisor. Wipe and clean the carriage and the chamber of the dry heat sterilizer with distilled water. Place systematically the load to be sterilized as per the loading pattern on the carriage and put the carriage back into the chamber of the dry heat sterilizer.
    20 : Close the pressure lock by turning the hand lock clockwise till the door is tight and the put the display ‘’Do Not Open’’ board on the door. Fill the sterilization document provided stating date items, number of the units and operators Put on the main and check the working of the booster fan and HEPA module. Affix a thermograph duly labeled onto the thermograph recorder and put on the temperature recorder.
    21 : Set the temperature to 260 c on the temperature controller unit. Set the timer on the temperature controller unit to the validated set time depending upon the load to be sterilized. Start the heater switch and simultaneously start the timer switch the heater timer display and the HEPA timer display starts with zero reading. Temperature will gradually rise and will be indicated by the temperature indication and thermograph.
    22 : After sterilization cycle is completed the heater will stop and HEPA module will start to cool the load. Prior to the unloading the charge on the sterile side check the following Thermograph has recorded the desired time and temperature relationship. HEPA module is operating. Load is sufficiently cool to be unloaded on sterile side.
    23 : Sterilization of bulk solution is done by the sterile filtration using sterilized (10nm-10mic meter)membrane filter. These membranes are made up of variety of polymers such as: cellulose silica ester polyvinyl diene fluoride poly tetra fluoro ethylene MEMBRANE FILTRATION
    24 : SOP FOR MEMBRANE FILTRATION Cleaning of membrane filter holder: Dismantle the membrane filter holder immediately after use. First remove the membrane holder lid from the base by unscrewing wing nuts. Remove the membrane diaphragm and bleed valve from the lid unscrew the inlet nozzle from the lid Remove the gasket from the lid and screen. Rinse all the parts with hot distilled water twice and keep in a clean plastic bucket after draining of the rinse water.
    25 : Wash the all components top plate, base plate, gasket, supporting screen,nozzle,air bleed valve, and its holder twice with hot water with hot pyrogen free water directly from distillation still. A clean sponge for scrubbing hard to clean areas and stiff bristled brush for inside of nozzle can be used. An aqueous solution of 1% or 50% benzalkonium chloride can be used for washing a membrane filtrate holder which can used for an oily product. Rinse with filter free flowing dt water and then fresh free water till the rinse water is free from benzalkonium chloride.
    26 : Assembling of membrane filter holder After completing the cleaning allow to air dry all the components and do not wipe with cloth or tissue. Bring the all the items and accessories required for assembling the filter holder after washing under a laminar flow. Fix the gasket to lid and screen tightly into the slot. Put the screen supporting plate on the base . Take the correct size membrane wet it in distilled water and placed center it properly and not creaet any folds.
    27 : Place correct size pre filter on membrane with rough side on top. Put the diaphragm into inlet opening Hold the lid horizontally and slowly put on the base plate without disturbing the membrane and pre filter. Place the inlet nozzle and tight it Loosely cover the inlet and outlet nozzle with parchment paper and affix sterilization indicator. Holder with membrane filter now ready for sterilization.
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