MICLAB-045 Environmental and Plant Hygiene Monitoring Procedure

Daily monitoring of sterile grade areas during production is to be conducted by trained production staff. The Microlab is to ensure that the necessary plates are delivered on a daily basis so monitoring can take place.

Once a test has been completed, the responsible operator is to initial the plate and make sure that the BPN of the batch running at the time of the test is written on the plate.  Plates will be labelled with prompts to ensure this isn’t forgotten.  If no batch is running at the time of the test N/A should be put on the plate instead of a BPN.

If an area of concern is noted during routine daily testing, inform Micro immediately so that further steps can be taken.

Once a week a Microlab technician will perform environmental monitoring and a housekeeping audit of the area.

If, in monitoring any of the following areas, any of the following events occur, the Microlab staff member responsible for conducting Environmental Monitoring in that area is to launch an Environmental Monitoring Investigation by IMMEDIATELY repeating the test and as follows:

Table A

An Environmental Monitoring Investigation form is to be the basis of the investigation but does not limit the scope of the investigation.

To facilitate final sign off of completed investigations the Investigation File is to be reviewed at 2 (two) monthly intervals by the Microbiology Manager.

OOL results will be reviewed in a two-phase approach.  Phase 1 is the review of the testing procedures by the microbiology technician to eliminate the possibility of lab introduced error or contamination.  Phase 2 investigations involve a thorough systematic analysis of the incident, to determine the root cause for the OOL result.  Phase 2 investigations are to be tailored for each incident to reflect the significance of meeting or exceeding either the ALERT or ACTION limits as outlined in Table A.

1. Micro-Organism Levels in Air

1.1.         Samples are to be taken using the MAS Air Sampling equipment, where organisms will become impinged onto Trypticase Soy Agar. When testing Laminar Flow make sure to turn on the L/F 10 minutes prior to sampling if not already on.  Charge the Air Sampling equipment as required.  The Air Sampling head must also be sterilized by autoclaving when the equipment is being recharged.  Record details in the autoclave log and maintenance record book.

1.2.         Prepared irradiated and non-irradiated Trypticase Soy Agar (TSA) plates purchased from an approved supplier are to be used in the monitoring of air inside the factory.  Use irradiated plates in sterile areas and non-irradiated plates in lower environmental graded areas. If plates are not available prepare plates with approximately 25mL/plate of Trypticase Soy Agar.  Ensure the petri dish has sufficient amount of agar to cover the surface of the entire plate and is not able to dry out.

1.3.         Prepared TSA plates will have an expiry date printed on the surface of the lid.  All other plates prepared within the microlab have an expiry date of 1 month from date of pouring.  Store plates in refrigerator.

1.4.         Each Agar plate should be labelled clearly with the date and the area sampled.

1.5.         Plates are to be incubated at 32°C (± 1.5 °C) for 48 hours (+ 24 hours).

1.6.         After each area is a sampled sign and date relevant form.

1.7.         Record any unusual activities/observations about the area being monitored in the Environmental Monitoring Comments book, include date and time.

1.8.         After incubation, the total count is recorded on a histogram in either the Environmental Results Surface Non- Sterile or the Sterile File.  Prepare separate graphs for each area sampled.  Alert and Action Limits need to be ruled onto the form in red pen and the location written on the top of the form next to Area.  (Once a Form is complete it must be signed by the Technician and reviewed by a Manager).  After each area is a recorded sign and date relevant forms.  Extended incubation is to be carried out for all areas sampled.

1.9.         After the total count is recorded, re incubate the samples for a further 72 hours (+ 48 hours) at 25°C (± 1.5°C).  Record the total count as well as number of molds present on the histogram graph shading them in with a Green pen.

1.10.      In the event that the result from the Air monitoring equals or exceeds either the ALERT or ACTION levels, follow the actions as listed under ‘Micro Organisms in Air” see section 9.

1.11.      Sampling Areas: Take one (1) sample from each area.

Summary: Table B for Incubation temperature, Incubation time and action.

Air sampling must also be conducted in all of the Filling rooms in the Sterile Area prior to, or as close as possible to the commencement of filling processes after any non‑aseptic procedure has been carried out in a Sterile Filling area, i.e. After “open day” maintenance, etc.  Any deviation from this SOP is to be recorded in the “Environmental Monitoring Comments” book and a DR raised, if necessary.

2. Micro-Organism levels on surfaces using Contact Plates

2.1.         Prepared irradiated and non-irradiated Count-Tact plates purchased from an approved supplier are to be used in the monitoring of surfaces inside the factory.  Use irradiated plates in sterile grade areas and non-irradiated plates in non sterile grades.  If plates are not available prepare contact plates with approximately 18mL/plate of Nutrient Agar + 3% w/v Tween 80.  Ensure plates are sufficiently full to allow a convex agar surface above the level of the plastic base.

2.2.         Prepared Count-Tact plates will have an expiry date printed on the surface of the lid.  All other plates prepared with in the microlab have an expiry date of 1 month from date of pouring.  Store plates on compactor shelves.

2.3.         Procedure for sampling – Operator to assess for “Worst Case” situation:

2.3.1.     Record details of date and area sampled on the contact plate.

2.3.2.     Touch agar surface of plate to surface to be sampled and hold plate down for 10 seconds.

2.3.3.     Replace the lid on the plate.

2.3.4.     Spray area sampled with 70% Isopropyl alcohol (IPA) and wipe with a clean wipe.

2.3.5.     Sign and date relevant form after each area is sampled

2.3.6.     Record any unusual activities/observations about the area being monitored in the Environmental Monitoring Comments book, include date and time.

2.3.7.     Incubate plates inverted at 32°C (± 1.5°C) for 48 hours (+ 24 hours).

2.3.8.     After incubation the total count is recorded on a histogram in either the Environmental Results Surface Non- Sterile or the Sterile File.  Prepare separate graphs for each area sampled.  Alert and Action Limits need to be ruled onto the form in red pen and the location written on the top of the form next to Area.  Once a Form is complete it must be signed off by the Technician and then reviewed by a Manager.

2.3.9.     Sign and date the relevant form after each area is recorded. Extended incubation is to be carried out for all areas sampled.

After recording the total count, re-incubate the plates for a further 72 hours (+ 48 hours) at 25°C (± 1.5°C).  Record the total count as well as number of molds present on the histogram graphs shading them in with a Green pen.

2.3.10.   In the event that the result from the Surface monitoring equals or exceeds either the ALERT or ACTION levels, follow the actions as listed under ‘Micro Organisms on Surfaces”, see section 9.

2.3.11.   Sampling Areas: Take one (1) sample from each area.

Summary: Table C for Incubation temperature, Incubation time and action.

2.3.12.   NOTE: Contact Plate monitoring must also be conducted in all Sterile Filling rooms prior to or as close as possible to, the commencement of filling processes after any non-aseptic procedure has been carried out in the Sterile Filling Area, i.e. After “Open Day” maintenance, etc.  On these occasions the floor and walls are to be examined.  Alert Level for walls in the Sterile Filling areas = 2 colonies/plate.

Any deviation from this SOP is to be recorded in the “Environmental Monitoring Comments” book and a DR raised, if necessary.

3. Personnel

(See MICLAB 095 and MICLAB 060)

3.1.         Operator’s Gloved Hands

The number of viable micro-organisms on the gloved hands of Sterile Area Operators is to be enumerated daily for every batch using Fingerprint Impressions.

During Sterility Testing the Microlab technician is to perform a gloved hand impression for each sterility session.  This is to be performed underneath the Laminar Flow of sterile filling areas. The method to be used is as follows:

3.1.1.     Micro. Lab. Preparation

90mm Prepared Letheen Agar plates purchased from an approved supplier are to be used in all Sterile Areas.  Prepared plates will have an expiry date printed on the surface of the lid.

If plates are not available prepare 150mm sterile petri dish with approx. 75mL/plate of Letheen Agar to use.  Ensure the petri dish has sufficient amount of agar as to cover the surface of the entire plate and is not able to dry out.  At time of use individuals will divide plate in half and label with the headings as outlined below:

All plates have an expiry date of 1 month from the date of pouring.  Store the prepared plates in clean designated buckets with lids on the compactor shelves with a label stating the expiry date.  This is to prevent a build up of condensation in the refrigerator.

3.1.2.     Sterile Area Operators

Each operator evaluates the sterile grade working background by conducting finger print impression testing.  This is done in the Sterile Scrub Preparation Room.
Sterile area limits are applied.

a) Place plate on bench with line to center of your body.

b) Raise lid with two fingers

c) Dab the four fingers of one hand on one side of the divided plate and then dab the thumb of the same hand in the center of that half of the plate.

d) Repeat with the other hand on the opposite side of the plate.

e) Label the plate against the corresponding headings. It is VERY important that all information is labelled on the plate as it corresponds to the batch and can affect batch release.

Full Name      ________________________
Date              ________________________
Area/Team     ________________________
BPN              ___________

f) Change gloves if returning to the Sterile work area.

Note:
This will have been demonstrated and taught to the Operator during Sterile Training.

Note:
After recording the fingerprint impression, the Operator must don a new pair of sterile gloves in the Sterile Scrub Preparation Room and disinfect their gloves with Hexifoam prior to re-entering Sterile Areas.  This is only valid if uniform monitoring has not been conducted.  If it has, then the operator must re-gown before re-entering the sterile area.

3.1.3.     When exiting the Sterile Scrub Preparation Room fill in Form 635 the Daily Personnel Monitoring Log for Sterile Areas.

3.1.4.     Collect the Letheen Agar plates from the factory after checking them against Form 635 and incubate in the Microbiology Laboratory.

3.1.5.     Incubate plates, inverted, at 32°C (± 1.5°C) for 48 hours (+24 hours).  After incubation, examine the plates and count the number of colonies present.

3.1.6.     Record the details of the number of colonies graphically in the Personnel Monitoring Reports files.  Prepare separate graphs for each Operator.  Record the total count as well as number of molds present on the histogram graphs shading them in with a Green pen.  For the purpose of determining if the count is over the specified limit, molds are included in the final result.

Record the details of the Microlab Technician results on the Monitoring results for sterility test sessions Finger, Fallout, Environmental and Personnel Results forms.

3.1.7.     Limits

Sterile Manufacturing.

Microlab Technician each sterility session.

On the Manufacturing graphs the Alert limit will be shown as 6 and Action limit will be 10 to account for both hands.  In the event that the result from the Finger dab monitoring equals or exceeds either the ALERT or ACTION levels, follow the actions as listed under ‘Micro Organisms on Personnel”, see Section 9.

3.2.         Operator’s Clothing

The number of viable microorganisms on the clothing of Sterile Area Operators is to be enumerated daily for every batch using contact plates.
At the completion of Sterility Testing the Microlab technician is to perform testing of their clothing.
Note: If a double consecutive session is being tested, glove print impressions should be taken after the first session with gloves then changed before the second session.  Personnel uniform monitoring should be performed after the second session only.

The method to be used is as follows:

3.2.1.     Prepared irradiated Count-Tact plates are purchased from an approved supplier.  If plates are not available prepare contact plates with approximately 18mL/plate of Nutrient Agar + 3% w/v Tween 80.  Ensure plates are sufficiently full to allow a convex agar surface above the level of the plastic base.

3.3.         Prepared Count-Tact plates will have an Expiry date printed on the surface of the lid. All other plates prepared within the Micro Lab have an Expiry date of 1 month from date of pouring.  Store plates on compactor shelves.

3.3.1.     Procedure for Sampling

a) Record details of the Operator’s name, clothing sampled, work area, BPN and date on the contact plate.  It is VERY important that all this information is written on the plates as they correspond to the batch and can affect batch release.

b) Decontaminate the plates by spraying with 70% IPA and place on the dividing bench in the Sterile Scrub Preparation Room.

c) At the conclusion of work, when exiting from Sterile the Operator is to remove the lid from the contact plate and touch the Agar surface to the area to be sampled.  The lid of the contact plate is to be IMMEDIATELY replaced.

d) When exiting the Sterile Scrub Preparation Room fill in
Form 635 the Daily Personnel Monitoring Log for Sterile Areas.

e) Microlab staff collect the contact plates from the factory after checking them against Form 635 and incubate in the Microbiology Laboratory.

f) Incubate plates, inverted, at 32°C (± 1.5°C) for 48 hours (+24 hours).
After incubation, examine the plates and count the number of colonies present.

g) Record details of the number of colonies graphically.  Prepare separate graphs for each Operator.  Record the total count as well as the number of molds present on the histogram graphs shading them with a Green   For the purpose of determining if the count is over the specified limit, molds are included in the final result.

Record the details of the Microlab Technician results on the Monitoring results for sterility test sessions Finger, Fallout, Environmental and Personnel Results form.

3.3.2.     Sampling Areas: Take one (1) sample from each area.

Microlab Technician also conducts monitoring:

In the event that the result from the Operator clothing monitoring equals or exceeds either the ALERT or ACTION levels, follow the actions as listed under ‘Micro Organisms on Personnel”, see section 9.

4. Settle Plates for each Batch

(See MICLAB 095)

4.1.         Prepared Nutrient Agar plates purchased from an approved supplier are to be used to monitor fallout during filling of each batch.
Each Filling Machine is required to have exposed at least one settle (fallout) plate for 4 hours/ per shift/ per batch/ per number of days of filling, (i.e. if filling over 3 shifts, then you are required to have at least 3 fallout plates).  The plates are to be placed as near as practical to the point of fill and exposed for a maximum of four (4) hours.  In the case of a fill that is less than 4 hours, for example during a media run, the fallout plates are to be exposed for the length of the run with start and finish times clearly written on the plate.

4.2.         Ensure that the plates have been correctly labelled.  The written details required on the plates are:

4.3.         Plates are incubated at 32°C (± 1.5°C) for 48 hours (+24 hours).

Each plate is to be subjected to a total count and this information is to form part of the batch record.  Results are to be recorded into the log book.  This information forms part of the decision to release a batch for sale.

When recording the results in log book, comment the details of the fallout plate and who entered the details.

If any colonies are present record the morphological description and Gram stain reaction (microscopic appearance).  Identify at least Genus Level.  Record this information in the log book.

Action LIMIT:

Terminally Sterilized Product

Sterile filling    <1 CFU/4 hours.
If any gpsr are present, D values must be determined.  Limit for D-value is 1 minute for terminally sterilized products, (see MICLAB 065 & MICLAB 075).

Terminally Sterilized Product

Sterile filling    5 CFU/4 hours.
If any gpsr are present, D values must be determined.  Limit for D-value is 1.5, (see SOP MICLAB 065 & MICLAB 075).

Action and Alert LIMIT

Aseptically Filled Product and Terminally Sterilized Product

Alert – 3 CFU/4 hours.

Action – 5 CFU/4 hours

In the event that the result from the Settle plate equals or exceeds either the ALERT or ACTION levels, follow the actions as listed under ‘Micro Organisms on Settle Plates”, see section 9.

5. Testing of Floor Disinfectant

5.1.         The disinfectant used to clean the floors of the Sterile Area and the Factory must be tested on a weekly basis to ensure it is not contaminated with microbes.  Sample into a sterile 20mL McCartney bottle at different sampling points.

5.2.         Test Method

Table D

Disinfectant Test Method:

A 1ml sample of the disinfectant solution is added to 9mL of a suitable diluent and mixed thoroughly.  Allow standing time of 10mins.  (This is to inactivate the disinfectant.)  From this solution transfer 1mL onto a 150mm Letheen Agar plate.  Using a sterile spreader, smear evenly over the agar surface.  This is to be done in duplicate.

5.3.         One plate is to be incubated at 32°C (± 1.5°C) for 72 hours (+24 hours) and the other at 25°C (± 1.5°C) for (7days) 168 hours (+ 48 hours).

5.4.         Limit: 10²cfu/ml of disinfectant solution.
(1 cfu plate is equivalent to 10 cfu in the original disinfectant sampled.)

5.5.         The counts must be recorded in the Disinfectant Results File.

5.6.         If the disinfectant is contaminated with 10 or more colonies on one plate, follow the action as outlined in section 9, ‘OOL Disinfectants’.

6. Test Method for Swabbing

6.1.         When conducting a swab of a surface area, use sterile swabs and sterile water for injection.  Using aseptic technique, moisten the swab with the sterile water then swab over the test surface 5×5 cm.  Spray swabbed surface with 70% IPA to remove any residue.

6.2.         If a limit test is required place the swab inside a bottle of sterile Peptone water and snap off the stick without touching that part of the stick inside the bottle.  Label the bottle with the location of the surface and the test date.

6.3.         Before filtering shake well, under the laminar flow filter the contents of the bottle through a 0.45-micron membrane filter. Place the whole filter onto a sterile petri dish containing TSA.  Incubate the dish at 32°C for 5 days.  After incubation examine the petri dish and count the number of colonies present.  Gram-stain and identify as per MICLAB 070 Record all results in the Environmental Monitoring Comments Book.

6.4.         If a Presence test is required place the swab directly into a 100ml bottle containing 50ml of TSB and break off the stick without touching that part of the stick inside the bottle.  Incubate the bottle at 32°C for 5 days.  Inspect daily.  After the 5 days if no growth is apparent leave at room temperature for 7 days.  If any growth is found, gram-stain and identify as per MICLAB 070 Record all results in the Environmental Monitoring Comments Book.

7. Waste Tank Sampling for Sterile Filling Machines

7.1.         It is the responsibility of the microbiology laboratory staff to organize sampling of waste tank waters by Sterile Operators or where are able themselves.  Please see MICLAB 055 for procedure.

7.2.       Results are to be recorded on appropriate files.

8. Recommissioning of an Environmental Area

When an environmental area is to be returned to its correct Environmental Grade conduct an inspection audit of the area.
This will also include conducting environmental monitoring of the air and surfaces.

9. Out of Levels/Limits “OOL”

Alert Limits

If a test gives a one off result of equal to or above the Alert limit but less than the Action limit, a Deviation Report DR is not required.  It must however be gram stained and future results for that location must be analyzed to make sure it is not a chance of becoming an Action Limit risk.  Record the investigation.  Refer to each subsection on Air, Contact, Personnel, Settle plates and Disinfectants for required actions to take when the Alert limit is breached.

How and When to raise an Environmental DR

As stated in Table A, a DR is to be raised in the event of a result being equal to or above the Action limit, if the result has equaled or been above the Alert limit in 3 consecutive tests, or the result of a trend analysis over a period of one month shows that the mean value of monitoring results for an area exceeds the Alert Level.  See also Tables B and C.

The person recording environmental results for the day is responsible for raising the DR (follow SOP QMS 035) Or, if unable to themselves, they are responsible for organizing the DR being raised the same day by another trained member of the microlab staff.

Make sure that all required information is entered, i.e. Batch number, Author, Description and Department responsible.

What to add into the extended text box of the DR form

• Area

• Sample site

• Batch running at time of test (if applicable)

• Time

• Date

• Result

• Reason for the DR

• Person who took the sample

• Answers to as many Phase 1 Investigation questions as possible (see Section 9.1)

Example of Environmental Result DR

Sterile filler left side Air plate taken at 12pm dd/mm/yy running batch number XXXXXX, has a count of 2 colonies.  This is above the Action Limit for this area as outlined in MICLAB 045.  Mrs Y was the technician responsible for this test.  She noted no unusual observations during the time the test took place.  Collection, storage, preparation, set up of test equipment, incubation, calculation and test method procedures were followed correctly.  Mrs. Y’s training records are complete and up to date.  The media used was within expiry (TSA irradiated Batch number XXXXXX, expiry date dd/mm/yy) and media stasis and sterility were satisfactory.  Gram Stain and ID work, if necessary, are pending.

Example of Personnel Monitoring DR

Mrs. Y had a count of 6 colonies on her hood plate from the dd/mm/yy at 12pm.  The batches running at the time of her being inside Sterile were XXXXXX, (add in all necessary BPN’s).  This is above the Action Limit for Uniform plates as outlined in MICLAB 045. She noted that there was maintenance being performed in a number of the rooms she entered.  Collection, storage, preparation, set up of test equipment, incubation, calculation and test method procedures were followed correctly.  Mrs. Y’s training records are complete and up to date.  The media used was within expiry (Letheen Agar Batch number aaaaaa, expiry date dd/mm/yy, Count-Tact plates with Batch number bbbbbbb, expiry dd/mm/yy) and media stasis and sterility were satisfactory. Gram Stain and ID work, if necessary, are pending.

Example of Settle Plate DR

Sterile filling Machine Left hand side Settle plate has a count of 6 cfu/4 hours from the dd/mm/yy, batch number XXXXXX.  Start time: 10.30, Finish time: 14.30.  This is above the Action level for this area according to MICLAB 045.  Items such as set up of test equipment, test method, up to date training records and any observations made at the time of test are awaiting interview with the sterile operator responsible for running the test.  Collection, storage, preparation, incubation and calculation procedures were followed correctly.  Media used is within expiry (Nutrient agar batch number xxxxxxx expiry dd/mm/yy) and media stasis and sterility were satisfactory. Gram Stain and ID work, if necessary, are pending.

Example of Disinfectant Sample Test DR

The Sterile filler disinfectant sampled on the dd/mm/yy by Mrs. Y and tested on the dd/mm/yy by Mr Z has a count of 21 colonies.  This is above the Action limit for this area as outlined in MICLAB 045.  Lisa noted no unusual observations during the time the test took place.  Collection, storage, preparation, set up of test equipment, incubation, calculation and test method procedures were followed correctly.  Mrs Y’s training records are complete and up to date.  The media used was within expiry (Letheen Agar Batch number xxxxxx, expiry date dd/mm/yy) and media stasis and sterility were satisfactory.  Gram Stain and ID work, if necessary, are pending.

9.1.         Phase 1 Investigation Responses

9.1.1.     Phase 1 investigations for Environmental OOL results must consider, (but are not limited to), the following details:

• Correct collection, storage and preparation of test sample.

• Correct set-up of test equipment.

• Correct method used for test.

• Completed Training records.

• Expiry of consumables and equipment observed.

• Media stasis and sterility satisfactory.

• Correct incubation conditions of test.

• Observations made during test procedure.

• All calculations checked.

• All test control results, where relevant, are satisfactory.

• Review of other test results for the test session, to determine a trend.

9.1.2.     Phase 2 investigations involve a thorough systematic analysis of the incident, to determine the root cause for the OOS result.  Phase 2 investigations are to be tailored for each incident to reflect the significance of meeting or exceeding either the ALERT or ACTION limits.

9.2.         Phase 2 Investigation Responses

9.2.1.     Determine if the alert excursion is to become an action level excursion that requires upgrading the notification to an action level based on the following criteria: See Table A under ‘Procedure’ at start of this SOP.

9.2.1.1.    Three alert/action level excursions or more of the in the same room on the same day, (e.g., Sterile filler floor surface, machine surface and airlock surface.) within the following time frames:

• Daily samples must not exceed the above criteria over a week period.

• Weekly samples must not exceed the above criteria over a month period.

• Monthly samples must not exceed the above criteria over 6 months.

• Annual samples must not exceed the above criteria over 2 years.

• Recovery of an undesirable organism in the critical grades clean rooms.

• Consistent alert level excursions, which do not show a decrease in magnitude or frequency of occurrence in a utility system – as per Microbiology group discretion.

9.2.2.     The minimum requirements for most situations are listed as below, this list is by no means exhaustive and specialist assistance from departments other than Microbiology should be utilized in the investigation.

Note: All affected departments will perform their part of the investigation, carry out corrective action and document all corrective actions and findings of investigation.

9.3.         OOL Micro organism Level in Air

9.3.1.     Alert Level

• Immediately re-sample.

• Report OOL result to Team Leader.

• The organisms present are to be Gram stained and a record of this and their morphological appearance kept in the relevant Environmental ID File.

• If any gram negative rods are found, identify to Genus level.

• Perform three (3) days follow-up testing on point.  Note: If follow-up results indicate that the point is not under control, the investigation should be elevated to an ACTION limit excursion.

9.3.2.     Action Level (in addition to Alert level requirements)

• Where required, take the environment out of use immediately, until corrective action has been taken.

• Identify all organisms to at least Genus level.

• Over room pressure data for the room is to be reviewed.

• Area to be re-cleaned immediately and review the cleaning records for the past week.

• Review data from area for past 3 months.

• Visual inspection of the area for contributing factors, i.e. general cleanness level / adherence to procedures

• List of activities/ unusual processes that occurred on day of high monitoring results

• Check for any maintenance performed or due on equipment / system.

• Liaise with equipment / system owner to determine possible causes of OOL/OOS result.

• Hold investigation meeting to determine corrective and preventative actions, (CAPA’s) and also determine impact on production activities.

9.4.         OOL Surface / Swab Sampling

9.4.1.     Alert Level

• Immediately re-sample.

• Report OOL result to manager.

• The organisms present are to be Gram stained and a record of this and their morphological appearance kept in the relevant forms.

• If any gram Negative rods are found, identify to Genus level.

• Perform three (3) days follow-up testing on point.  Note: If follow-up results indicate that the point is not under control the investigation should be elevated to an ACTION limit excursion.

9.4.2.     Action Level (in addition to Alert level requirements)

• Where required, take the environment out of use immediately, until corrective action has been taken.

• Identify all organisms to at least Genus level for sterile areas.

• Over pressure data for the room is to be reviewed

• Area to be re-cleaned immediately and review the cleaning records for the past week.

• Review data from area for past 3 months.

• Visual inspection of the area for contributing factors, i.e. general cleanness level /adherence to procedures.

• List of activities/unusual processes that occurred on day of high monitoring results.

• Check for any maintenance performed or due on equipment / system.

• Liaise with equipment / system owner to determine possible causes of OOL/OOS result.

• Hold investigation meeting to determine corrective and preventative actions, (CAPA’s) and also determine impact on production activities.

9.5.         OOL Personnel Monitoring

9.5.1.     Alert Level

• Report OOL result to manager.

• Each morphological group present on the plate is to be Gram Stained.  A record of this and their morphological appearance is to be kept in the relevant file.

• If any gram negative or positive rods are found, identify to Genus level.

• Ensure a review of the operator’s results for the following 2 results.

• Any colonies identified on the Microlab Technician results from a sterility test session, must be gram stained and identified to Genus level and kept until the results of sterility test session are completed.

• Review the bioburden levels for the 70% IPA used.  (Gloves.)

9.5.2.     Action Level (in addition to Alert level requirements)

When investigating an OOL result for a personnel plate linked to batch release these areas must be highlighted:

• Length of time in area

• Gram stain result

• Daily environmental monitoring results

• The Environmental Comments book

• Work orders raised by Micro for the area

• Operator behavior

• Settle plates

• Sterilization charts

• Operator must be counselled to determine their actions inside the sterile areas to determine the root cause.

• Operator to undertake re-validation.

• Determine what BPNs the individual had been working with on that day.  These BPNs are to be linked to the DR.

• Over pressure data for the room is to be reviewed.

• Area to be re-cleaned immediately and review the cleaning records for the past week.

• Review all environmental data from the areas in which the operator was working, (i.e. settle plates, air and surface).

• Visual inspection of the area for contributing factors, i.e. general cleanness level /adherence to procedures.

• List of activities/unusual processes that occurred on day of high monitoring results.

• Check for any maintenance performed or due on equipment/system.

• Hold investigation meeting to determine corrective and preventative actions, (CAPA’s) and also determine impact on production activities, batch disposition to be assessed.

9.6.         OOL Settle Plates (Fallout Plates)

9.6.1.     Alert Level

• Report OOL result to Team Leader, to discuss if extra samples are needed for testing.

• Each morphological group present on the plate is to be Gram Stained.  A record of this and their morphological appearance is to be kept in files.

• All micro-organisms are to be identified to at least Genus level.

• For any gpsr isolated, the D value level must be determined for terminally sterilized product.

• Major Stoppages, Sample Identification sheet and Run Time sheet.

• Review all other settle plate results for the BPN.

9.6.2.     Action Level (in addition to Alert level requirements)

• Operator must be counselled to determine their actions inside the sterile areas to determine the root cause.

• Over pressure data for the room is to be reviewed.

• Review Steaming records.

• Machine Cleaning records.

• Area to be re-cleaned immediately and review the cleaning records for the past week.

• Review all environmental data from the area, (i.e. personnel, air and surface).

• Visual inspection of the area for contributing factors, i.e. general cleanness level /adherence to procedures.

• List of activities/unusual processes that occurred on day of high monitoring results.

• Check for any maintenance performed or due on equipment/system.

• Hold investigation meeting to determine corrective and preventative actions, (CAPA’s) and also determine impact on production activities, batch disposition to be assessed.

9.7.         OOL Disinfectants

In the event that the level has been reached, Raise a Deviation Report and the following Actions are to be considered as part of the OOL investigation:

• Immediately re-sample and test the sample point.

• Report OOL result to manager.

• Liaise with the cleaning co-ordinate for the extent of the use of the disinfectant, when manufactured, changed, expiry, correct storage and preparation procedures for the past week.

• Check when buckets last sterilized and have the buckets re-sterilized.

• Identify organism to at least Genus level.  Results are to be recorded in the relevant files.

• If necessary, change the type of disinfectant being used in the area.

• Test the original concentrate.

• Perform a visual inspection of the storage area and preparation equipment and if necessary take swabs/samples.

• Review the microbiological water results used for the preparation of the disinfectant.

• Review Cleaning records for area.

• Hold investigation meeting to determine corrective and preventative actions, (CAPA’s) and also determine impact on production activities

10. Summary Sheet – Level of Identification

11. Summary Sheet – Recording Quick Reference

12. Summary of Changes