{"id":708,"date":"2025-06-14T06:10:24","date_gmt":"2025-06-14T06:10:24","guid":{"rendered":"https:\/\/pharmacoguide.com\/?p=708"},"modified":"2025-06-14T06:10:24","modified_gmt":"2025-06-14T06:10:24","slug":"sop-title-validation-of-laboratory-autoclave-located-in-microbiology-laboratory","status":"publish","type":"post","link":"https:\/\/pharmacoguide.com\/?p=708","title":{"rendered":"SOP Title: Validation of Laboratory Autoclave located in Microbiology Laboratory"},"content":{"rendered":"

SOP Title:<\/u><\/strong> Validation of Laboratory Autoclave located in Microbiology Laboratory<\/u><\/strong><\/p>\n

    \n
  1. Objective:<\/strong><\/li>\n<\/ol>\n

    To lay down the procedure for Validation of Laboratory Autoclave in Microbiology section of Quality Control Department.<\/p>\n

      \n
    1. Scope<\/strong><\/li>\n<\/ol>\n

      This procedure is applicable for Validation of Laboratory Autoclave section of Quality Control Department.<\/p>\n

        \n
      1. Responsibility:<\/strong><\/li>\n<\/ol>\n

        3.1 Quality Control: To prepare and review the SOP. To follow the procedures for Validation of Laboratory Autoclave in Microbiology section as per this SOP.<\/p>\n

          \n
        • Quality Assurance Department: To review and approve the SOP and Annexure.<\/li>\n<\/ul>\n
            \n
          1. Accountability:<\/strong><\/li>\n<\/ol>\n

            Head Quality Control Department, Head Quality Assurance Department.<\/p>\n

             <\/p>\n

              \n
            1. Procedure:<\/strong><\/li>\n<\/ol>\n\n\n\n\n\n\n\n\n\n\n\n\n\n
              Sr. No.<\/strong><\/td>\nProcedure<\/strong><\/td>\n\u00a0<\/strong><\/td>\n<\/tr>\n<\/thead>\n
              \u00a0\u00a0 5.1<\/strong><\/td>\nSafety Precautions:<\/strong><\/td>\n<\/tr>\n
              5.1.1<\/td>\nEnsure that the appropriate protective equipments are used during handling of Autoclave.<\/td>\n <\/td>\n<\/tr>\n
              5.1.2<\/td>\nCheck the water level before each cycle in the Autoclave.<\/td>\n <\/td>\n<\/tr>\n
              5.1.3<\/td>\nEnsure that the instrument is cleaned and free from dust and is disconnected from the mains after use.<\/td>\n <\/td>\n<\/tr>\n
              \u00a0 \u00a05.2<\/strong><\/td>\nProcedure to carry out validation of Autoclave :<\/strong><\/td>\n<\/tr>\n
              5.2.1<\/td>\nProtocol for Autoclave validation should be prepared.<\/td>\n <\/td>\n<\/tr>\n
              5.2.2<\/td>\nValidation of Autoclave is carried out for<\/p>\n

              1)\u00a0\u00a0\u00a0 Heat distribution\/penetration study with temperature Mapping.<\/p>\n

              2)\u00a0\u00a0\u00a0 Sterilization study of Biological Indicators.<\/td>\n

              		 <\/td>\n<\/tr>\n
              \u00a0 \u00a05.3<\/strong><\/td>\nHeat distribution study with temperature Mapping:<\/strong><\/td>\n<\/tr>\n
               <\/td>\nAcceptance criteria: <\/strong>The load penetration study carried out in Autoclave should comply with the acceptance criteria of respective heat distribution study for Empty load, half load and full load.<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

               <\/p>\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n
              <\/td>\n<\/td>\n\u00a0<\/strong><\/td>\n<\/tr>\n<\/thead>\n
               <\/td>\nFrequency<\/strong>:\u00a0 Initial once after installation and then once in a Year.<\/td>\n<\/tr>\n
              5.3.1<\/td>\nTo ensure that the autoclave is capable to attain the temperature of 121\u00b0C during the sterilization hold at a specified hold period with the steam pressure on the pressure gauge is 15 psi.<\/td>\n <\/td>\n<\/tr>\n
              5.3.2<\/td>\nEnsure that the autoclave temperature and pressure gauges are calibrated. Check the calibration status of Data logger and sensors used for calibration.<\/td>\n <\/td>\n<\/tr>\n
              5.3.3<\/td>\nProtocol for validation and revalidation of autoclave to be prepared as per SOP.<\/td>\n <\/td>\n<\/tr>\n
              \u00a0 A.<\/strong><\/td>\nHeat Distribution study at 121\u00b0C \/ 15 psi \u00a0in Empty chamber<\/strong><\/td>\n<\/tr>\n
              5.3.3<\/td>\nEmpty cycle load heat distribution study shall be performed.<\/td>\n <\/td>\n<\/tr>\n
              5.3.4<\/td>\nFill purified water in autoclave upto the mark before starting.<\/td>\n <\/td>\n<\/tr>\n
              5.3.5<\/td>\nSet the parameters for the sterilisation cycle set to be operated.<\/td>\n <\/td>\n<\/tr>\n
              5.3.6<\/td>\nInsert minimum 8 to 12 nos. of sensors or suitable numbers depending upon the volume of autoclave into the chamber through the port provided for Autoclave.<\/td>\n <\/td>\n<\/tr>\n
              5.3.7<\/td>\nPlace one sensor near the autoclave controller probe, one sensor near the drain and one sensor in the middle position, distribute the remaining probes uniformly in the chamber. Place a chemical indicator (Autoclave sterilization indicator) tape\/strip and a biological indicator near each of the first three sensors mentioned above. Define Annexure for sensor locations.<\/td>\n <\/td>\n<\/tr>\n
              5.3.8<\/td>\nSeal the logger port with silicon sealant so that steam leakage does not take place.<\/td>\n <\/td>\n<\/tr>\n
              5.3.9<\/td>\nConnect the sensors to a suitable calibrated data logger which can scan and print the actual temperature with time.<\/td>\n <\/td>\n<\/tr>\n
              5.3.10<\/td>\nSet the Autoclave to attain the desired temperature of 121\u00b0C at 15 psi during the complete sterilisation hold period.<\/td>\n <\/td>\n<\/tr>\n
              5.3.11<\/td>\nStart the autoclave cycle after ensuring all the necessary precautions are taken. In Empty load cycle the chamber must be totally empty except for data logger sensors and chemical and Biological indicators.<\/td>\n <\/td>\n<\/tr>\n
              5.3.12<\/td>\nStart the data logger recording once the cycle is started; record the actual temperature at an interval of 1 minute during sterilization cycle of 15 minutes or 30 minutes by using a data logger. After completion of sterilization, record the data in datalogger until temperature drops down to 100 \u00b0C.<\/td>\n <\/td>\n<\/tr>\n
              5.3.13<\/td>\nThe temperature spread should be within the range of 121\u00b0C to 124\u00b0C during sterilisation for set temperature of 121\u00b0C at 15 psi pressure for 15 minutes and 30 minutes.<\/td>\n <\/td>\n<\/tr>\n
              5.3.14<\/p>\n

               <\/td>\n

              		Review the temperature data for all sensors printed on datalogger print, check and compare the temperature observed at different location.<\/td>\n <\/td>\n<\/tr>\n
              5.3.15<\/p>\n

               <\/td>\n

              		For Sterilization study of Biological Indicators refer point No. 5.4.<\/td>\n <\/td>\n<\/tr>\n
              5.3.16<\/td>\nFor Heat Distribution study details record in Annexure.<\/td>\n <\/td>\n<\/tr>\n
              5.3.17<\/td>\nDefine Annexure for the list of cycles and for sterilization parameters.<\/td>\n <\/td>\n<\/tr>\n
              \u00a0B.<\/strong><\/td>\nHeat Distribution\/Penetration study at 121\u00b0C \/ 15 psi for Media(Half Load\/Full Load)<\/strong><\/td>\n<\/tr>\n
              5.3.18<\/td>\nMedia Sterilization cycle is validated for 121\u00b0C at 15 psi pressure for 15 minutes.<\/td>\n <\/td>\n<\/tr>\n
              5.3.19<\/td>\nPlace one sensor near the autoclave controller sensor, one sensor near the drain and one sensor in the middle position, distribute the remaining sensors uniformly in the chamber. Place a chemical indicator (Autoclave sterilization indicator) tape\/strip and a biological indicator near each of the first three sensors mentioned above. Define sensor locations.<\/td>\n <\/td>\n<\/tr>\n
              5.3.20<\/td>\nUse same numbers of sensors as used in the Empty load distribution cycle.<\/td>\n <\/td>\n<\/tr>\n
              5.3.21<\/td>\nCarry out the autoclave cycles as per Empty load method. Refer Point No.5.3.4 to 5.3.17.<\/td>\n <\/td>\n<\/tr>\n
              5.3.22<\/td>\nPrepare Solid and Liquid media for carrying out the media cycles as per SOP. Prepare preferably SCDA and R2A agar medium as solid media. SCDM and MacConkeys broth shall be prepared as liquid media. If these media are not available, any other media can also be used. Enter Media prepared in Annexure.<\/td>\n <\/td>\n<\/tr>\n
              5.3.23<\/td>\nHalf load<\/strong> refers to the the media load which occupies half or minimum space in the Autoclave during sterilization cycle. Record in Annexure.<\/td>\n <\/td>\n<\/tr>\n
              5.3.24<\/td>\nFull load<\/strong> refers to the Media load which occupies the maximum loadable space in autoclave during sterilization cycle. Record contents of cycle in Annexure.<\/td>\n <\/td>\n<\/tr>\n
              5.3.25<\/td>\nPlace sensors in between the media glasswares and in the media at different locations to map the temperature distribution and penetration of steam.<\/td>\n <\/td>\n<\/tr>\n
              5.3.26<\/td>\nFor Steam sterilization study of Biological Indicators refer point No. 5.4<\/td>\n <\/td>\n<\/tr>\n
              5.3.27<\/td>\nFor Heat Distribution study record in Annexure.<\/td>\n <\/td>\n<\/tr>\n
              C.<\/strong><\/td>\nHeat Distribution\/Penetration study at 121\u00b0C \/ 15 psi for Garments\/ Glasswares<\/strong>\/Accessories.(Half Load\/Full Load)<\/strong><\/td>\n<\/tr>\n
              5.3.28<\/td>\nGarments\/ Glassware\/Accessories Sterilization cycle is validated for 121\u00b0C at 15 psi for 30 minutes as to sterilize the garments and other items for maximum duration.<\/td>\n <\/td>\n<\/tr>\n
              5.3.29<\/td>\nPlace one probe near the autoclave controller probe, one probe near the drain and one probe in the middle position, distribute the remaining sensors uniformly in the chamber. Place a chemical indicator (Autoclave sterilization indicator) tape\/strip and a biological indicator near each of the first three sensors mentioned above.<\/td>\n <\/td>\n<\/tr>\n
              5.3.28<\/td>\nUse same numbers of sensors as used in the Empty load distribution cycle.<\/td>\n <\/td>\n<\/tr>\n
              5.3.30<\/td>\nCarry out the autoclave cycles as per Empty load method. Refer point No. 5.3.4 to 5.3.17.<\/td>\n <\/td>\n<\/tr>\n
              5.3.31<\/td>\nHalf load<\/strong> refers to the the Garment\/Glassware\/ Accessories load which occupies half or minimum space in the Autoclave.<\/td>\n <\/td>\n<\/tr>\n
              5.3.32<\/td>\nFull load<\/strong> refers to the Glasswares\/ Garments\/ Accessories which occupies the maximum loadable space in autoclave. Record in Annexure for loading pattern and contents of cycle.<\/td>\n <\/td>\n<\/tr>\n
              5.3.33<\/td>\nPlace sensors in between the Garments and glasswares at different locations to map the temperature distribution and penetration of steam.<\/td>\n <\/td>\n<\/tr>\n
              5.3.34<\/td>\nFor Analysis of Biological Indicators refer point No. 5.4<\/td>\n <\/td>\n<\/tr>\n
              5.3.35<\/td>\nFor Heat Distribution study record in Annexure.<\/td>\n <\/td>\n<\/tr>\n
              D.<\/strong><\/td>\nHeat Distribution\/Penetration study at 115\u00b0C \/ 10 psi for Media.(Full Load)<\/strong><\/td>\n<\/tr>\n
              5.3.36<\/td>\nMedia Sterilization cycle for some media like Rappaport is to be validated for 115\u00b0C \/ 10 psi pressure for 15 minutes.<\/td>\n <\/td>\n<\/tr>\n
              5.3.37<\/td>\nFollow the same procedure as media cycle point No. 5.3.19 to 5.3.25<\/td>\n <\/td>\n<\/tr>\n
              E.<\/strong><\/td>\nHeat Distribution study at 121\u00b0C at 15 psi for Media Discard autoclave.(Full Load)<\/strong><\/td>\n<\/tr>\n
              5.3.38<\/td>\nMedia Decontamination\/Sterilization cycle is validated for 121\u00b0C at 15 psi for 30 minutes.<\/td>\n <\/td>\n<\/tr>\n
              5.3.39<\/td>\nPrepare Solid and Liquid media (Refer point No. 5.3.22) for carrying out the media decontamination cycles. Glassware should be included for this type of cycle. Record media prepared in Annexure.<\/td>\n <\/td>\n<\/tr>\n
              5.3.40<\/td>\nPerform the validation as per Empty cycle method point No. No.5.3.4 to 5.3.17.<\/td>\n\u00a0<\/strong><\/td>\n<\/tr>\n
              \u00a0\u00a0 5.4<\/strong><\/td>\nB) Steam Sterilization study of Biological Indicators<\/strong><\/td>\n<\/tr>\n
               <\/td>\nFrequency:<\/strong>\u00a0 Once a year during validation, every six months as to check sterilization efficiency or as per validation requirements.<\/strong><\/td>\n<\/tr>\n
              5.4.1<\/td>\nPlace 3 spore strips\/ampoules of Biological indicator Geobacillus stearothermophilus<\/em> (Bacillus Stearothermophilus<\/em>) ATCC 7953 in the most difficult to sterilize positions in the chamber.<\/td>\n <\/td>\n<\/tr>\n
              5.4.2<\/td>\nSterilize in cycles as per procedure for individual cycle.<\/td>\n <\/td>\n<\/tr>\n
              5.4.3<\/td>\nAfter sterilisation, incubate each spore strip in a individual 10 ml Soyabean Casein Digest Medium test tube for 7 days at 55\u00b0C to 60\u00b0<\/sup>C. For media preparations refer SOP QC 034. Keep positive and Negative control with the test strips. Positive control (Unexposed) should be incubated in SCDM along with test strips for 7 days at 55\u00b0C to 60\u00b0<\/sup>C. Negative control (Unexposed) Indicators to be incubated at 30\u00b0C to 35o<\/sup>C for 7 days for checking the spore growth if any in normal conditions. Check the Biological Indicator tubes daily for 7 consecutive days and record in Annexure.<\/td>\n <\/td>\n<\/tr>\n
              5.4.3<\/td>\nThe unexposed spore strip (positive control) shows growth (turbidity), indicates that spores are viable.<\/td>\n <\/td>\n<\/tr>\n
              5.4.4<\/td>\nNegative control shows no growth (turbidity) means spores are not activated.<\/td>\n <\/td>\n<\/tr>\n
              5.4.5<\/td>\nIf\u00a0 the\u00a0 positive\u00a0 control\u00a0 shows\u00a0 turbidity\u00a0 and\u00a0 all three\u00a0 test\u00a0 samples\u00a0 are\u00a0 not showing\u00a0 turbidity after 7 days of\u00a0\u00a0 incubation\u00a0 period,\u00a0 then\u00a0 the Autoclave\u00a0 is\u00a0 working\u00a0 satisfactory\u00a0 as\u00a0 per\u00a0 the\u00a0 requirements.<\/td>\n <\/td>\n<\/tr>\n
              5.4.6<\/td>\nIn case of ampoules, incubate the ampoules after sterilization at 55\u00b0C to 60\u00b0<\/sup>C for 48 hours. Keep positive and Negative control with the test ampoules. Incubate positive control (Unexposed) along with test ampoules. Incubate the negative control (Unexposed) at 30\u00b0<\/sup>C -35\u00b0<\/sup>C for 48 hours.<\/td>\n <\/td>\n<\/tr>\n
              5.4.7<\/td>\nExposed ampoules should not show any colour changes.<\/td>\n <\/td>\n<\/tr>\n
              5.4.8<\/td>\nThe unexposed ampoule (positive control) shows colour change, indicates that spores are viable.<\/td>\n <\/td>\n<\/tr>\n
              5.4.9<\/td>\nNegative control should show no colour change.<\/td>\n <\/td>\n<\/tr>\n
              5.4.10<\/td>\nRecord the data on Annexure<\/td>\n <\/td>\n<\/tr>\n
              5.4.11<\/td>\n\u00a0If\u00a0 validation fails then do not use the Autoclave. Inform the Head of Quality control and the Head of Quality assurance,\u00a0 raise an incident and investigate\u00a0 the\u00a0 reasons\u00a0 &\u00a0 take\u00a0 immediate\u00a0 action\u00a0 to\u00a0 rectify\u00a0 the\u00a0 same\u00a0 and \u00a0perform\u00a0 Revalidation.<\/td>\n <\/td>\n<\/tr>\n
              5.4.12<\/td>\nDecontaminate the spore strips in autoclave before disposing off. Refer SOP for Decontamination of culture media and Microbial waste.<\/td>\n <\/td>\n<\/tr>\n
              <\/td>\n<\/td>\n<\/td>\n<\/tr>\n
              <\/td>\n<\/td>\n <\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

              \u00a0<\/strong><\/p>\n

                \n
              1. Definitions \/ Abbreviations: <\/strong><\/li>\n<\/ol>\n

                \u00a0<\/strong><\/p>\n

                  \n
                • Definitions<\/strong><\/li>\n
                • Abbreviations :<\/strong><\/li>\n<\/ul>\n\n\n\n\n
                  Abbreviation<\/strong><\/td>\nExpansion<\/strong><\/td>\n<\/tr>\n
                  psi<\/td>\nPressure square inch<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

                   <\/p>\n

                  *Note – Ready to use SOP available in \u201cDOWNLOAD\u201d Section.<\/u><\/em><\/strong><\/p>\n

                   <\/p>\n","protected":false},"excerpt":{"rendered":"

                  SOP Title: Validation of Laboratory Autoclave located in Microbiology Laboratory Objective: To lay down the procedure for Validation of Laboratory Autoclave in Microbiology section of Quality Control Department. Scope This procedure is applicable for Validation of Laboratory Autoclave section of Quality Control Department. Responsibility: 3.1 Quality Control: To prepare and review the SOP. To follow […]<\/p>\n","protected":false},"author":1,"featured_media":0,"comment_status":"open","ping_status":"closed","sticky":false,"template":"templates\/template-full-width.php","format":"standard","meta":{"footnotes":""},"categories":[7],"tags":[],"class_list":["post-708","post","type-post","status-publish","format-standard","hentry","category-microbiology"],"_links":{"self":[{"href":"https:\/\/pharmacoguide.com\/index.php?rest_route=\/wp\/v2\/posts\/708","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/pharmacoguide.com\/index.php?rest_route=\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/pharmacoguide.com\/index.php?rest_route=\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/pharmacoguide.com\/index.php?rest_route=\/wp\/v2\/users\/1"}],"replies":[{"embeddable":true,"href":"https:\/\/pharmacoguide.com\/index.php?rest_route=%2Fwp%2Fv2%2Fcomments&post=708"}],"version-history":[{"count":1,"href":"https:\/\/pharmacoguide.com\/index.php?rest_route=\/wp\/v2\/posts\/708\/revisions"}],"predecessor-version":[{"id":709,"href":"https:\/\/pharmacoguide.com\/index.php?rest_route=\/wp\/v2\/posts\/708\/revisions\/709"}],"wp:attachment":[{"href":"https:\/\/pharmacoguide.com\/index.php?rest_route=%2Fwp%2Fv2%2Fmedia&parent=708"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/pharmacoguide.com\/index.php?rest_route=%2Fwp%2Fv2%2Fcategories&post=708"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/pharmacoguide.com\/index.php?rest_route=%2Fwp%2Fv2%2Ftags&post=708"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}