GUIDE TO AUTOCLAVING

8/10/2019 GUIDE TO AUTOCLAVING

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GUIDE TO A U T O C L AV I N G

P I O N E E R S I N S T E R I L I Z AT I O N T E C H N O L O G Y


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Sterilization may be defined as the statistically completedestruction of all microorganisms including the most resistantbacteria and spores. This is a condition that is difficult toachieve and hard to prove. Whilst there are many chemicals,inorganic and organic, that kill microorganisms they may not betotally effective and can leave undesirable or toxic residues.Ultraviolet and Ionising radiations are also effective biocides,disrupting or modifying the DNA to prevent replication, butUltraviolet will not produce the effective results and easyvalidation that moist heat (steam) sterilization can provide.If sterility is an absolute requirement then todays scientists turn,as their predecessors did, to steam.Microorganisms tend to become more active as the

temperature of their surroundings rises, - most, but not all, dieat above 80 oC. In the case of Prions the temperature and timerequirements for deactivation are much higher.Steam molecules condense on cooler microorganisms, andtransfer 2500 joules per gram of steam, very efficiently heatingthe microorganisms to the temperature at which they aredestroyed. Other methods of heating suffer from the muchlower heat transfer of hot dry gases and boundary layer effects,

which can insulate and protect the microorganisms.For maximum effect the Steam must be saturated, and thiscondition, and the temperature and pressure of the steam areeasily monitored, facilitating proof that sterilization has occurred.By employing Steam Sterilization techniquesa high level of sterility can be achieved andthe most popular piece of equipment usedin laboratories and hospitals is the STEAMSTERILIZER or AUTOCLAVE.

The terms Autoclave and Sterilizer are interchangeable and mean

the same thing. Autoclave is a term used mainly in laboratorieswhilst Sterilizer is used more commonly for medical andpharmaceutical applications.

An effective Autoclave or Sterilizer must contain dry saturatedsteam. In order to achieve this air must be removed from boththe load and the chamber; this can be accomplished in a numberof ways. Air removal from high mass low load items (i.e.comprising mainly solid areas that contain little or no air pocketssuch as bottled media) will need little air removal and this can befacilitated by automatic air purging. Here air is allowed to leavethe chamber through a vent as steam enters the chamber eitherfrom an integral source (upward displacement) or an externalsupply (downward displacement); this vent only closes when all of the air is removed from the chamber. This method can be furtheradvanced by Freesteaming when the vent is allowed to stayopen for a set length of time. The turbulent steam then passesthrough the vent forcing any trapped air out of the autoclave. Formore stubborn loads that contain a number of air pockets suchas wrapped instruments or fabrics, a more effective method of airremoval is essential. By far the most effective way of air removalis a vacuum system, whereby a vacuum is achieved in thechamber before any introduction of steam, removing most of theair before freesteaming and/or vacuum pulsing.

Once all of the air is removed from the load and chamber thetemperature within the vessel will rise along with the pressure until

the pre-selected temperature is reached. To achieve a typicalsterilization temperature of 121 oC or more, requires the steam tobe pressurized to at least 1.1 Bar G. Since pressures used aregreater than 0.5 Bar G the Autoclave assembly is classified as apressure system and must be designed to a strict engineeringstandard such as PED97/23EC/PD5500/ASME etc.

The temperature within the chamber is reached in a number

of ways:

1) Steam can be injected into the chamber via an internalsteam source such as an integral stainless steel steamgenerator, which can be built within the Autoclave cabinetor can be supplied as a separate unit.

2) Some laboratories or hospitals have their own direct steamsource, which would similarly be injected, into the chamber.

3) In some Autoclaves heaters are built into the base of thechamber and the water is heated until it boils and producessteam.

The main advantage of using a steam generator (or the directsteam method) is that cycle times can be considerably faster asthe steam is immediately available. In basic autoclaves thewater is topped up manually by pouring water into the vesselwhilst some autoclaves are connected to a direct water supplyand the water level is maintained automatically. Some of Astellssmaller units are available with an integral water tank, whichonce filled, can run for up to 20 cycles repeatedly before beingreplenished. In CSSD applications it is a requirement that thesteam is dry saturated steam with a known dryness and non-condensable gas content e.g. in the UK, Steam with valueswithin the specification set out in HTM2010.

When the desired temperature is achieved for the required timethen the steam supply will cease either by shutting off the powerto the heaters or by cutting off the steam supply. Thus thetemperature and the pressure will gradually drop. In units with avacuum system fitted the vacuum pump can be used to removethe steam (i.e where drying is required).

How does an Autoclave work?

What is Sterilization?


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Typical temperature/time sterilization parameters are 115 oC for30 minutes, 121 oC for 15 minutes and 134 oC for 3 minutes.However, overall cycle times will vary depending on loadcharacteristics. Firstly it is important to realise that the time andthe amount of steam that is needed for the load to reachSterilization will vary greatly depending on the thermal capacityof the load in question. Two or Three 1 litre bottles of culturemedia will often need more steam than a dozen smallcontainers, and the heating time will take longer. Likewise thetime taken to achieve Sterilization once the temperature hasbeen attained will depend on the type of microbiologicalorganisms present. Bacteria and fungi are rapidly killed by moistheat but viruses and particular bacterial spores are very

persistent, and may need long Sterilization times. Thedevelopment and wide availability of reliable microprocessorsmeans that most modern autoclaves are fully userprogrammable, allowing the Sterilization temperature to be setto suit the requirements of the load.

The most accurate way of ensuring that Sterilization occurswhen the correct temperature is reached is by Load SensedProcess Timing. Here the cycle is controlled via the temperatureachieved in the centre of the load. A wandering thermocoupleprobe situated within the chamber is inserted into the load, or aload simulator and initiates the sterilization period once theprobe reaches the programmed threshold temperature. Thisensures that sterilization starts at the correct stage of the cycle.

Following Sterilization and depressurization of the chamber itmay be possible to open the autoclave immediately. However,when autoclaving bottled fluids, a cooling lock is normally fittedto comply with various safety regulations (e.g. in the UK: HSE

PM73). This prevents the autoclave from being opened until theload temperature has fallen to typically 80 oC, thus reducing therisk of both the boiling over of fluids (e.g media) and theexplosion of glass bottles.

Following Sterilization it is possible to have a variety of fastcooling options fitted which reduce the cool down period.

Deluge Cooling This can often cut cycle times down by up to 50% and is usedonly in conjunction with sealed fluid loads. Water at Sterilizationtemperature is sprayed onto the load after the completion of thesterilization stage. The water being sprayed is passed through aheat exchanger cooled by cold water thus producing acontrolled reduction in temperature throughout the load. This isa highly specialised method of cooling, and due to the volumeof water used during the deluge stage it is only suitable forsealed bottled fluids.

Water Fed Jacket A Jacket can be fitted to the autoclave chamber on bothcircular and square section units. Again cold water is introducedinto the jacket at the end of the sterilization stage of the cycleusing the chamber walls as a heat exchanger reducingtemperature and resulting in improved cooling. As the jacketacts as a very efficient heat exchanger pressure reduction canbe very fast using this method and therefore it is generallyrecommended that Air Ballast is used in conjunction with the

jacket to reduce the possibility of fluid loss.

Advanced Water Cooling This can only be fitted to machines that have been connected

to a mains water supply. A series of cooling coils are fitted tothe outside of the autoclave chamber. At the end of sterilizationthese coils are circulated with cold water and so results in arapid decrease of temperature and improved cooling time.

Assisted Air-Cooling

Fans are fitted within the cabinet and circulate ambient air overthe external chamber walls during the cooling stage of the cyclecreating a controlled improvement to cooling times.

Internal Convection Fan Assisted Cooling A fan or fans can be built within the chamber and ducted tocreate an efficient airflow. The fan is again activated during thecooling stage of the cycle. The fans create turbulence reducingcooling time greatly. Internal convection fan cooling is one of the most proficient ways of cooling the chamber. To ensureimproved cooling time the optional jacket and/or advancedwater-cooling is also needed. On loads using unsealed bottlesair ballast is recommended.

Typical Autoclave Cycles

Types of Cooling

Water Cooling

Fan Assisted/Air Cooling

The importance of drying!

It is important to take into account exactly what the autoclavewill be used for before running a cycle. Some items such asfabrics & wrapped instruments require special attention as theywill need a specific drying cycle to avoid them being left wet atthe end of the cycle. The best way of ensuring that this doesnot happen is by having a pre and post vacuum as well as asteam heated jacket fitted to the chamber. The pre-vacuum isrequired to remove air from the load and the post vacuum willremove moisture left on the load. The drying will be enhanced

with the addition of the steam heated jacket, which is filled withsteam throughout the cycle; this heat prevents excessivecondensation being formed on the chamber walls and maintainsthe temperature above the boiling point during drying so thatwater droplets boil off. The Heated jacket is essential foroptimum drying performance.


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Autoclaves can be used for numerous medical and laboratoryapplications in Market Sectors such as, Food and Dairy,Universities & Colleges, Agriculture & Horticulture Research,Healthcare (including Pathology), Industrial Research, Water & Environmental, Brewing, Biotechnology Industries andPharmaceuticals

Typical applications include:Media PreparationSterilization of FluidsInstruments (Wrapped and Unwrapped)Utensils and Lab EquipmentFabrics and Textiles

Media Preparation: Media is used to grow microorganismsunder test conditions and a wide variety of gel and liquid mediais used. Air removal from fluid/media loads is normallystraightforward; however some media cycles can benefit from ashort period of free steaming. Different types of media withstanddifferent sterilizing temperatures, therefore any autoclave usedfor the Sterilization of media must be capable of sterilizing at avariety of temperatures. Cycle times must be short and thespeed of processing is vital. A fast heat up and cool down isbeneficial in the maintenance of the quality of the finishedproduct and prevents the over cooking which leads to thedeterioration in the quality of the growth medium. Cycle lengthshould be set to as short as possible, and at the temperaturespecified by the media manufacturer. The cool down period

however, needs to be a controlled process in order to preventboil over and loss of media. The main method of controlling thesterilzation procedure of any cycle is to use a thermocoupleprobe in a dummy bottle within the chamber (Load SensedProcess Timing). Only when the probe reaches the correcttemperature will the sterilization process begin. Load SensedProcess timing is used in conjunction with Profiled OvershootBoost, this allows the temperature in the chamber to rise abovethe programmed level to allow the load to reach the correcttemperature faster. The risk of any boil over can be greatlyreduced by using Air Ballast which maintains chamber pressureduring the cooling phase.

Sealed Fluids: Some fluid loads may require to be sterilized in

special sealed containers. Air purging is normally sufficient toremove air, and extended freesteaming should be unnecessary.

As with media preparation the speed of processing is vital to thequality of the end product and often temperatures in excess of 121 oC will be used to speed up the Sterilization process. Againthe cooling should be as short as possible.

Instruments: The Sterilizationof instruments fallsinto two main categories:

a) Unwrapped Instruments b) Wrapped Instruments

Both types of instrument loads benefit from drying the end of the cycle. Unwrapped instruments can be dried by the heatingof the surrounding air at the end of the cycle, whilst wrappedinstruments require a more sophisticated drying system Postvacuum systems on small bench-top units will improve drying,whilst a jacketed pressure vessel is strongly recommended onlarger units.

Uses of Autoclaves

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VAC POSITIVE PULSE STERILIZE COOLING DRYING AIR BREAK

2100mbar

Ombar

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VAC POSITIVE PULSE STERILIZE COOLING DRYING AIR BREAK

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VAC NEGATIVE PULSE POSITIVE PULSE STERILIZE DRYING AIR BREAK

Sealed Fluids Cycle Graph (including assisted cooling)

Typical Glassware/Utensils (134C) Cycle Graph

Sealed Fluids Cycle Graph


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Fabrics and Textiles: A variety of fabrics and textiles can besterilized in an autoclave/sterilizer, e.g many hospitals will use anautoclave to sterilise gowns, instrument packs etc. As these aretrue porous loads they will require both pre and post vacuumsystems, negative and positive steam pulsing and a fully heated

jacket. Steam quality is important when operating Porous Load Autoclaves.

Discard Loads: There is an ever-increasing need for the safeSterilization of contaminated material before disposal to ensurethat it is microbiologically safe. The contaminated material canbe of many types and in many forms. For example, allmicrobiological laboratory waste should be processed prior todisposal, as must all clinical samples e.g. blood, tissue etc.Loads consisting of such a wide variety of materials in a broadrange of containers will present specific Sterilization problems.One of the main problems with such mixed discard loads is

that they invariably contain small pockets of trapped air, whichpresent particular air removal problems before steampenetration. Such loads are often contained in special plasticbags and comprise a variety of plastic containers; these areliable to melt and collapse before Sterilization , thus increasingthe likelihood of trapped air. A pre-vacuum and steam pulsingfeature is now a commonly requested option for autoclavesrunning frequent discard cycles. However an alternative tovacuum is a long period of freesteaming or PulsedFreesteaming PAT . There are special discard (Morrison) containersavailable for this application. In certain circumstances a double-

doored autoclave may be required. The loading (dirty or non-sterile) end is often separated from the unloading (clean orsterile) end by a wall, which often contains an SPF seal toprevent bacteriological migration. In category III laboratories aneffluent retention system would also be necessary.

Load Vacuum AssistedSensed Vacuum & Assisted CoolingProcess Pulsing Heated Cooling & AirTiming Jacket Ballast

x x ( 3 )

x x x

x x

x x x x

x ( 1 ) x x

x ( 1 ) x x

x ( 2 ) x x

Ombar

1000mbar

2000mbar

3000mbar

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VAC NEGATIVE PULSE POSITIVE PULSE STERILIZE DRYING AIR BREAK

Typical Fabrics Cycle Graph

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VAC POSITIVE PULSE STERILIZE DRYING AIR BREAK COOLING

Typical Fluid Discard Cycle Graph

Media

MixedDiscard

FluidDiscard

Glassware

UnwrappedInstruments

WrappedInstruments

Fabrics

Essential Recommended x Not Needed(1) If dry load is required

(2) Vacuum & Heated Jacket required to ensuredrying of Fabric Loads

(3) To prevent excessive cycle time


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Verification has become increasingly important, asthere is often a need to trace back through a series of results to ensure that a particular Sterilzation process

has been carried out. There are a variety of options thatare available for this; these can either be mounted externally i.eon an adjacent wall or shelf, or can be an integral part of theautoclave. Printers, Chart Recorders or Dataloggers are thepreferred options. These instruments can provide either a hardcopy record of exactly what the microprocessor is registering, orthey can be completely independent from the microprocessorand record independent results of chamber and load conditions.

Data Printers

Integral Data Printer Built as an integral part of an autoclave this is the most popularand cost effective way of monitoring the cycle. When the cyclebegins the printer records the starting t ime, cycle number,program number (or name) and machine type. The printer willprint a complete record of cycle conditions, freesteam, sterilizingstart and finish time and pressure readings at one-minuteintervals. There is also provision for operator signature andbatch number.

Secureprint Data Printer This is a Freestanding Data-Printer independent of themicroprocessor, which isdesigned to give an independentprinter verification of the sterilizingfunction. Chamber temperature,pressure, load and time isdisplayed and printed with a dualspeed papersave feature.(This printer will also operate withany non Astell Autoclave)

Chart recorders

Chart recorders are the traditional method of printing theSterilization process. These show a graphical display of pressure and/or temperature come with either one, two or threepens, single pen recorders register chamber temperature, twopen recorders register chamber temperature and loadtemperature and the three pen recorders register chambertemperature and load temperature via two wandering probes,options also include measurement and display of pressure.

The recorders can either come in strip or circular format.

Circular Chart

Recorders These are highperformancemicroprocessor basedrecorders with analphanumeric displayto provide moreinformation and easieroperation. Therecorders are availablewith up to 1, 2 or 3pens. Readingaccuracy is 1% of Span of theinstrument. Theprogramme system allows full control of ranging, plus control of the pen drive, lift and referencing, and display. The tactilemembrane switches are mounted at the front, alongside thedisplay, behind the door for security.

Strip ChartRecorders

These aremicroprocessorcontrolled strip chartrecorders availablewith 1,2 or 3 coloured

pens. Recordingaccuracy is 1% of span through the useof accurate steppermotors.

The recorder incorporatesa high definitionalphanumeric display, indicating temperatures and pressure.

Astell AutoSCRIBE Datalogger

The Astell datalogger is a purpose designed, wall mounteddatalogger with LCD display and keyboard.

The datalogger is specially configured to record temperatureand time parameters. The configuration of the AutoSCRIBEallows maximum storage by only recording during the operationcycle. The data is downloaded on demand to a laboratoryP.C. for analysis and archiving.

Monitoring the Sterilization process

Typical printout from integral or Secureprint data printer.


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Astell Scientific have been pioneers in the manufacture andvalidation of Scientific and Laboratory equipment for over 80years. In the last decade the company has concentrated onbecoming market leaders in Sterilization technology and is nowone of the premier manufacturers in this field in Europe.

A wide range of standard front and top loading microprocessorcontrolled Autoclaves are offered including circular section unitsfrom 23-330 Litres and Square Section Sterilizers from 100-490litres. In recent years Astell has successfully developed itsbespoke manufacturing facility and offers a wide range of nonstandard units in capacities up to 1000 litres. All of the unitsfeature the secure micro-computer controller, which is userfriendly and fully programmable. A variety of options and

accessories can be supplied with all models to ensureoptimum performance and user flexibility in the mostdemanding situations. These range from Data Printers,Load Sensed Process timing and a variety of cooling optionsto reduce cycle time.

The Astell range can be catogorised as follows:NB. For further information on Astells Autoclaves and Sterilizers

please refer to our separate brochure: The Autoclave Rangeor visit our Website at www.astell.com

Astell - Pioneers in Sterilization Technology

Swiftlock Compact Autoclaves,23, 40 & 60 Litres

This range of circular section autoclaves from Astell offerguaranteed precision and versatility for a wide range of applications. Featuring many of the benefits associated with

Astells larger units these stylish yet economical models meetthe demanding requirements of current Sterilization procedures.

These are now available in the Classic and Autofill versions.

Secure micro-computer control 316L stainless steel pressure vessels Temperature range 100-138C (0.2 2.4 Bar) Super-quick New Swiftlock door mechanism Compact ergonomic designs Automatic cycle control Built in drying system (Autofill version) Integral reservoir and/or condensate bottle 10 Year Pressure Vessel Integrity Warranty Media holdwarm feature (Classic version)


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Astell Scientific Limited, Powerscroft Road, Sidcup, Kent DA14 5DT United KingdomT (020) 8300 4311 F (020) 8300 2247 E [email protected] www. astell.com

Direct Sales: T (020) 8309 2024/3/2 F (020) 8309 2036 0594 Certificate No: Q536

GUIDE TO AUTOCLAVING

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