Monitoring of the personnel gowning

1.0 OBJECTIVE

To evaluate the aseptic technique followed by the clean room personnel while wearing the sterile garments and to monitor the bioburden on the gowns of personnel including the finger dabs in the aseptic processing area.

2.0 SCOPE

2.1 This evaluation and monitoring is applicable to personnel entering the aseptic processing area of Company Name.

3.0 RESPONSIBILITY

3.1 Microbiologist

4.0 ACCOUNTABILITY

4.1.1 Head – Quality control

4.1.2 Head – Quality Assurance

5.0 PROCEDURE

Introduction: The gown up practices of all persons entering the aseptic processing area should be routinely monitored for compliance with the procedure SOP. Improper gowning can contribute particles / microorganisms, which lead to contamination of sterile area.

5.1 Requirements

5.1.1 Sterile swabs

5.1.2 Sterile water for dipping swabs

5.1.3 Pre-incubated and sterile Petri plates filled with soyabean casein digest media containing 0.5% glycerol.

5.2 Media and Plate preparation

5.2.1 Prepare the media as per SOP

5.2.2 Prepare the plates as per SOP

5.3 Plates

5.3.1 Use sterile soyabean casein digest agar plates prepared as per SOP.

5.3.2 Collect the required number of plates that are preincubated at least for 48 hours.

5.3.2 Inspect each plate thoroughly for any sign of contamination such as, any colony, suspect colony, crack, condensate or dehydration in a black and white background prior to use.

5.3.3 If any such plate is observed record in the logbook and discard as per SOP.

5.3.1 Transfer the inspected good plates to Helmut block in a clean sanitized canister through IPQA pass box.

5.3.4 Follow the dress code and entry procedure for entry of personnel and material into the material pass box located adjacent to the component preparation area.

5.3.5 Sanitize the tray used for keeping plates in the material pass box with 0.2 micron filtered 70% IPA solution and sterilized lint free pads.

5.3.6 Ensure that all the disinfectant has evaporated from the surface of the tray prior to use.

5.3.7 Sanitize the plates on both the sides and periphery with 0.2µ filtered 70% IPA solution and sterilized lint free pads.

5.3.8 Leave the plates in the material pass box under U.V light for 15 min. on the inverted side by spreading on the tray.

5.3.9 Expose the other side of the plates also to U.V light for 15 min.

5.3.10 Enter into the aseptic processing area by following proper gowning procedure. SOP .

5.3.11 Again sanitize the plates from the aseptic processing side of the material entry pass box using sterile 70% IPA solution and sterile lint free pads.

5.3.12 Collect the plates and keep them in sterile storage area on a trolley dedicated for this purpose.

5.3 STERILE SWABS

5.3.2 Wrap the swabs with parchment paper in a clean and sanitized area.

5.3.3 The day before the swab test, pack the swabs in a beaker and autoclave the swabs and water collected in test tubes.

5.5 SAMPLING PROCEDURE

GOWNING EVALUATION

5.5.1 For a new person the gowning evaluation should be done three consecutive times. He should be allowed to enter the aseptic processing area only if the results comply for three consecutive times.

5.5.2 Gowning evaluation should be done immediately after gown up procedure in change room III, using sterile swabs. Test the uniforms at the following positions.

•Hood

•Uniform (a) Arm - R (b) Arm - L (c) Leg - R (d) Leg - L (e) zipper

•Booties

•Gloves (a) Fingers- R (b) Fingers - L

•Goggles

5.5.3 Take out the sterilized swab from the beaker and remove the parchment paper and hold the swab at one end.

5.5.4 Moisten the other end of the swab by dipping in sterile water.

5.5.5 Swab the sampling site by rolling the swab over 25cm2 area in a rolling fashion, vertically and horizontally.

5.5.6 Streak the sampled swab on the petri dish containing sterile Soybean Casein Digest Agar.

5.5.7 Plates must be identified as to location of garment sampled, name of the operator and date of sampling.

5.5.8 Additionally, in the microbiology laboratory, moisten one swab with sterile water and streak directly on the agar surface to serve as Negative control.

5.5.9 Additionally, in the microbiology laboratory, moisten one swab in a mixed culture of bacteria and fungi, and streak on the agar surface to serve as positive control. Record the numbers of colonies observed after incubation.

5.6 INCUBATION

5.6.1 Transfer the plates to the walk-in incubator in QC lab at 30 –35°C.

5.6.2 Incubate the plates in inverted position for 48 hrs at 30 –35°C and read the plates for bacterial colonies after completion of 48 hrs of incubation.

5.6.3 If there is a holiday then read the plates on the next working day.

5.6.4 While reading, record the results for each plate in the format – Annexure I.

5.6.5 Transfer the plates to the other walk-in incubator having 20-25°C and incubate for next 72 hrs. Read the plates for mold and yeast colonies on each day of incubation since mold may overgrow the entire surface of the plate.

5.4.1 If there is a holiday then read the plates on the next working day.

5.6.6 While reading, record the results for each plate in the format – Annexure I

5.6.7 After completion of incubation of the aseptic processing area plates, take the plates having counts for identification of the isolates obtained.

5.6.8 Give the reference number to each isolate as per the SOP and enter the reference number into the format – Annexure I.

5.6.9 Carry out the identification of isolates as per SOP

5.6.10 After completion of the incubation period, fill format, sign, check and submit to QA.

5.6 Sampling frequency:

5.6.1Gowning Evaluation to be done monthly to monitor gowning practices of all individuals who enter the aseptic processing area.

Limits:

Gloves: Less than 1 cfu

Other locations:

Alert limit: Not more than 2 cfu / plate.

Action limit: Not more than 3 cfu / plate.

5.7 Interpretation of Results:

5.7.1 If count is found to be more than the specified alert limit, inform QA, write in the corrective action report format

5.7.2 If contamination continues to be detected, further investigations should be carried out to eliminate the cause, as per SOP. The person concerned should be restrained from entering the aseptic area till retraining and evaluation shows the acceptable results.

5.7.3 The corrective action should be fully documented in the standard format

Note: Plates shall be exposed in change rooms while conducting gowning validation. This becomes part of evaluation report.

5. PERSONNEL MONITORING INCLUDING FINGER DABS

Frequency: Samples from the gowns and finger dabs of each operator at the end of the each shift.

Personnel monitoring

Personnel monitoring should be taken for each operator at the end of the shift.

Personnel swabs should be done in the sterile storage area. The personnel should stand outside the LAF and the sampling should be done on the person outside the LAF and transferred on the media plate under LAF.

Take the swabs at the following positions.

Finger dabs (both the hands in two different plates)

Arm pits (both the hands on the same plate that is divided to two halves by a marked line and each half identified for right and left)

Elbows (both the hands on the same plate that is divided to two halves by a marked line and each half identified for right and left)

Hand cuffs (both the hands on the same plate that is divided to two halves by a marked line and each half identified for right and left)

Shoulders (both the hands on the same plate that is divided to two halves by a marked line and each half identified for right and left)

Upper arms (both the hands on the same plate that is divided to two halves by a marked line and each half identified for right and left)

Upper Back (on one plate)

Hood (on one plate)

Negative control

By swabbing

5.7.1 Wet the sterilized swabs in sterile water for injection.

5.7.2 Swab the surface to be tested about 25 cm2 area with horizontal and vertical strokes and streak on plate containing Soybean casein digest agar in rolling fashion.

5.7.3 Identify the plates with location of garment sampled, name of the operator and date of sampling.

5.7.4 After the sampling, transfer the swabbed plates to the incubator through the material pass box in a tightly closed container.

By Finger Dabs

5.7.5 Take the glove prints before operator removes his gloves at the end of the operations in sterile storage prior to the operator carrying out any cleaning or tidying operations. Before performing the test, the operator should ensure that the gloves are dry and free from residual disinfectant.

5.7.6 First the four gloved fingers back and front should be imprinted on the agar surface of the plate for right and left hand separately. A firm and even pressure should be applied approximately for 5 to 10 seconds, taking care not to damage the agar surface. After that the gloved thumb should be imprinted on the agar surface and rolled a little in such a way that the entire thumb area is imprinted.

5.7.7 Replace the lid of the plates.

5.7.8 Identify the plates by the location sampled (right and left hand), name of the operator & date of sampling.

5.7.9 Operators must place the first pair of used gloves in a polythene cover and should be transferred immediately to QCD. Second pair of gloves shall be removed in the Anteroom.

5.7.10 Prior to destruction, sterilize the discarded glove by autoclaving at 121deg for not less than 30 min.

5.7.11 Transfer the plates to the incubator through IPQA pass box in a tightly closed container.

5.7.12 Record the details of activity, name of the personnel, sampled by, time of sampling etc.

Ensure that the gloves and surfaces are cleaned with a suitable disinfectant to remove any possible traces of residual agar before performing any other operations or before coming out.

5.8 Incubation

5.8.1 Transfer the plates to the walk-in incubator in QC lab at 30 –35°C.

5.8.2 Incubate at 30-35°C in inverted position for bacterial colonies after completion of 48 hrs.

EX: If the first shift plates were incubated at 13.00hrs on 12/04/22then the plates should be read after the completion of 48 hrs incubation, i.e. after 13.00 hrs on 14/04/22.

If the second shift plates were incubated at 21.00 hrs on 12/04/22then the plates should be read after the completion of 48 hrs incubation, i.e. after 21.00 hrs on 14/04/22.

Similarly if the third shift plates were incubated at 3.00hrs on 13/04/22then the plates should be read after the completion of 48 hrs incubation, i.e. after 3.00 hrs on 15/04/22.

5.8.3 If there is a holiday then read the plates on the next working day.

5.8.4 While reading, record the results for each plate in the Annexure II.

5.8.5 Transfer the plates to the other walk-in incubator having 20-25°C and incubate for next 72 hrs. Read the plates for mold and yeast colonies upon completion of each day of incubation as given in the example above.

5.8.6 If there is a holiday then read the plates on the next working day.

5.8.7 While reading, record the results for each plate in the Annexure II.

5.8.8 After completion of incubation, take the plates having counts for identification of the isolates obtained.

5.8.9 Give the reference number to each isolate as per the SOP and enter the reference number into the format

5.8.10 Carry out the identification of isolates as per SOP .

5.8.11 After completion of the incubation period, fill the format, sign, check and submit for review.

5.8.12 After completion of the review and signatures the original book copy is kept with the QC and the duplicate copy is given to QA.

Limits for personnel swabs

Alert limit: Not more than 2 cfu/plate

Action limit: Not more than 3 cfu/plate

Limits for Finger Dabs

Grade A (operational) Five fingers - less than 1 per glove