Protocol No. Sterility Test Validation Protocol AREA: Q.C - MICROBIOLOGY Page No.

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DEPARTMENT
PRO#$CT NAME E%%ECTIVE #ATE

QC -MICROBIO O!"

C&AN!E &ISTOR" #ate S'(ersede Nil CCR No. NA C)anges Made Original Issue Re*ision No. 00

TAB E O% CONTENTS S. No. 1.0 #.0 &.0 (.0 +.0 6.0 /.0 0.0 3.0 10.0 11.0 Proto ol a!!ro"al O$%e ti"e ' o!e )ali*ation *es ri!tion Res!onsi$ilities an* i*entifi ation of e,e ution tea.est !ro e*ures Re or*ing of o$ser"ations 1is re!an 2 an* orre ti"e a tion re!ort Co-!ilation4 re"ie5 an* su--ar2 re!ort A!!en*i, Re"ali*ation Criteria #escri(tion Page No. # & & & & & + + 6 6 6

Protocol No. Sterility Test Validation Protocol AREA: Q.C - MICROBIOLOGY Page No. # of 6

+., PROTOCO APPROVA 'igning of t6is a!!ro"al !age of !roto ol in*i ates agree-ent 5it6 t6e -et6o* "ali*ation a!!roa 6 *es ri$e* in t6is *o u-ent. If an2 -o*ifi ation in t6e -et6o* "ali*ation a!!roa 6 $e o-es ne essar24 a re"ision t6roug6 6ange ontrol s6all $e !re!are*4 6e 7e* an* a!!ro"e*. .6is !roto ol annot $e e,e ute* until a!!ro"e* $2 follo5ing !ersonnel. #e(art-ent Pre(ared /y Qualit2 Control Mi ro$iolog2 Re*ie0ed /y Qualit2 Control Mi ro$iolog2 A((ro*ed /y Na-e #esignation Signat're.#ate

Qualit2 Assuran e A't)orised /y

8O1 - QA

Protocol No. Sterility Test Validation Protocol AREA: Q.C - MICROBIOLOGY Page No. & of 6

1., OB2ECTIVE: .6e !roto ol is *esigne* so as to "ali*ate t6at an2 Ba teriostati an* 9ungistati a ti"it2 in6erent in t6e !ro*u ts to $e teste* *oes not a*"ersel2 affe t t6e relia$ilit2 of t6e test an* t6e test !ro e*ure to $e follo5e* is suita$le for testing t6e !ro*u ts as !er t6e !6ar-a oe!ial -et6o*s. 3., SCOPE .6is !roto ol is a!!li a$le to sterile ra5 -aterial an* finis6e* !ro*u ts4 56i 6 are teste* for sterilit2. 4., VA I#ATION #ESCRIPTION .6e !ur!ose of t6is !roto ol is to ensure t6at an2 Ba teriostati an* fungistati a ti"it2 in6erent in t6e Pro*u t *oes not a*"ersel2 affe ts t6e relia$ilit2 of t6e test !ro e*ure. 5., RESPONSIBI ITIES AN# I#ENTI%ICATION O% E6EC$TION TEAM +.1 Res(onsi/ilities: .6e grou! o-!rising of re!resentati"es fro- ea 6 of t6e follo5ing *e!art-ents an* t6e2 s6all $e res!onsi$le for t6e o"erall o-!lian e 5it6 t6is !roto ol. #e(art-ent Qualit2 Control Mi ro$iolog2 Res(onsi/ility :,e ute4 Parti i!ate an* !ro"i*e ne essar2 su!!ort for t6e "ali*ation a ti"it2. Pre!aration an* re"ie5 of t6e "ali*ation Re!ort of t6e *o u-ents an* its o-!lian e to -eet t6e acce(tance criteria of t6e !roto ol. re"ie5 an* o-!lian e to

Qualit2 Assuran e Monitoring t6e "ali*ation a ti"ities4 o-!ilation4 aut6ori;ation of t6e -et6o* "ali*ation re!ort an* its -eet t6e acce(tance criteria of t6e !roto ol. +.#

Identi7ication o7 E8ec'tors: All t6e i*entifie* e,e utors in"ol"e* 5it6 t6is Proto ol are to Re or* Na-e4 1esignation4 'ignature an* 1ate.

9., TEST PROCE#$RES Before !ro ee*ing for "ali*ations follo5ing -aterials are re<uire*. 'terile !ro*u t sa-!le. 'terile *istille* 5ater. 'terile Molten 'o2a$ean Casein 1igest Agar. Poure* 'C1A !lates. 'terile for e!s. 'terile -e-$rane filtration units. 'terile -e-$ranes. )orte, Mi,er. 0.1 = !e!tone 5ater. 'terile 9lui* .6iogl2 ollate Me*iu-. 'terile 'o2a$ean Casein 1igest Me*iu-.

Protocol No. Sterility Test Validation Protocol AREA: Q.C - MICROBIOLOGY Page No. ( of 6

Cultures

re<uire*

sus!ensions

of

Staphylococcus

aureus,

Pseudomonas

aeruginosa, Clostridium sporogenes, Bacillus subtilis, Candida albicans, Aspergillus Niger. .6e "ali*ation 5ill $e !erfor-e* un*er t6e follo5ing su$6ea*ings. 6.1 Pre(aration o7 +, :+,, cells . -l: Pre!are t6e 10 > 100 ells ? -l solution as !er t6e 'OP @@@. 6.#.1.1 6.#.1.# 6.#.1.& 6.#.1.( 6.#.1.+ 6.#.1.6 6.#.1./ 6.#.1.0 6.#.1.3 After sa-!ling4 t6e sterile !ro*u t sa-!le 5ill $ring to -i ro$iolog2 la$. Mi ro$iologist 5ill !la e t6e sterile !ro*u t sa-!le 5it6in t6e !ass $o, after *e onta-inating t6e outer surfa e of t6e !ou 6 5it6 *isinfe tant solution. :nter t6e sterilit2 testing roo- as !er t6e 'OP for sterile area entr2 an* e,it !ro e*ure for sterilit2 testing la$. After entering in t6e sterilit2 testing roo-4 filtere* IPA lean t6e LA9 surfa e 5it6 /0 =

9.1 To #eter-ine Sterility o7 t)e Prod'ct.

Pla e t6e 'terile !ro*u t 'a-!les 5it6in t6e LA9. Perfor- t6e sterilit2 test of t6e sa-!le $2 follo5ing !ro e*ure *es ri$e* in @@@. O!erate t6e sterilit2 test a!!aratus :<uino, as !er @@@@@. After !erfor-an e In u$ate 'o2a$ean Casein 1igest Me*iu- at #0 > #+ oC for 1( *a2s an* 9lui* .6iogl2 ollate Me*iu- at &0 > &+oC for 1( *a2s. O$ser"e t6e -e*iu- tu$es? anisters *ail2 for an2 gro5t6.

6.#.1.10 Re or* t6e o$ser"ations *ail2 in t6e for-at gi"en as anne,ure-1. 9.3 To #eter-ine t)e Bacteriostasis and %'ngistasis nat're o7 t)e Prod'ct. 6.&.1.1 6.&.1.# 6.&.1.& 6.&.1.( 6.&.1.+ 6.&.1.6 6.&.1./ 6.&.1.0 Perfor- t6e 56ole e,er ise un*er t6e LA9 in 'terilit2 testing area. .a7e t6e sterile sa-!le as !er t6e @@@@@ an* *issol"e t6e sa-!le in #00 -l of 0.1 = !e!tone 5ater. Perfor- t6e sterilit2 test for t6e a$o"e sa-!le $2 follo5ing !ro e*ure *es ri$e* in 'L.11#. Rinse t6e -e-$rane 5it6 # @ 100 -l !ortions of t6e sterile 0.1 = !e!tone 5ater. Ino ulate t6e final rinse 5it6 10 - 100 C9A? -L of t6e test -i roorganis-s. After filtration ase!ti all2 transfer t6e -e-$rane on to t6e 'C1M ? 9.GM *e!en*ing on t6e organis- $eing teste*. Perfor- t6e 56ole e,er ise 5it6 all t6e organis-s liste* $elo5. Note *o5n t6e results in Anne,ure-1. 9ilter # @ 100 -l !ortions of 0.1 = !e!tone 5ater t6roug6 0.(+ filter -e-$rane.

9.4 Positi*e Control: 6.4.1.1

Protocol No. Sterility Test Validation Protocol AREA: Q.C - MICROBIOLOGY Page No. + of 6

6.(.1.# 6.(.1.& 6.(.1.(

9inall2 rinse t6e -e-$rane 5it6 t6e t6ir* 100 -l 0.1 = sterile !e!tone 5ater ino ulate* 5it6 10 > 100 ells ? -l of an2 ulture liste* a$o"e. Re!eat t6e a$o"e e,er ise 5it6 all t6e ot6er organis-s. Ino ulate t6e -e-$rane into -e*iu-. S.No. 01 Medi'9lui* .6iogl2 ollate 'o2a$ean Casein 1igest Organis-s Staphylococcus aureus Pseudomonas aeruginosa Clostridium sporogenes Bacillus subtilis Candida albicans Aspergillus niger Strain N'-/er

0#

9.5 Negati*e Control: 6.5.1.1 6.+.1.# 9ilter & @ 100 -l !ortions of 0.1 = !e!tone 5ater t6roug6 0.(+ filter -e-$rane. Cut t6e -e-$ranes into t5o 6al"es an* ino ulate ea 6 6alf se!aratel2 into s!e ifie* -e*iu-. In u$ate test for sterilit2 sa-!le tu$es of 9lui* .6iogl2 ollate Me*iu- at &#.+ #.+BC an* for 'o2a$ean Caesin *igest Me*iu- in u$ate at ##.+ #.+BC for 1( *a2s. .6e negati"e re or*e*. ontrol s6oul* $e in u$ate* for 1( *a2s4 o$ser"e* *ail2 an* ontrol tu$es for & *a2s in ase of $a teria an* + *a2s

9.9 Inc'/ation o7 T'/es: 6.6.1.1

6.6.1.# 6.6.1.&

In u$ate t6e Positi"e in ase of fungi.

9.; Acce(tance Criteria: 6./.1.1 6./.1.# .6e gro5t6 of ea 6 test organis-s in t6e test sa-!le C5it6 t6e !ro*u tD is "isuall2 o-!ara$le to t6e gro5t6 of t6e !ositi"e ontrol. .6e negati"e in u$ation. ontrol s6oul* not s6o5 an2 gro5t6 *uring t6e 1( *a2s of

;., RECOR#IN! O% OBSERVATIONS Re or* t6e o$ser"ations after t6e e,e ution of ea 6 test !ro e*ures4 in t6e anne,ure>1. <., #ISCREPANC" AN# CORRECTIVE ACTION REPORT 1o u-ent an2 *is re!an ies o$ser"e* *uring t6e "ali*ation in anne,ure -1. In lu*e t6e orre ti"e a tions of t6e sa-e. E6en all t6e *is re!an ies are satisfa toril2 resol"e* or an a!!ro"e* a tion !lan is *e"elo!e* 56i 6 ensures t6at t6e *is re!an 2 5ill $e resol"e*. =., COMPI ATION> REVIE? AN# S$MMAR" REPORT Co-!ile an* re"ie5 t6at all test !ro e*ures 6a"e $een o-!lete*4 re on ile* an* atta 6e* to t6is !roto ol. )erif2 t6at t6e a!!ro"als for *e"iations 6a"e $een ta7en an* are resol"e* a!!ro!riatel2 to t6e satisfa tion. .esting -et6o* "ali*ation s6all $e onsi*ere* a in t6e test !ro e*ures 6a"e $een -et. e!ta$le 56en all t6e on*itions s!e ifie*

Protocol No. Sterility Test Validation Protocol AREA: Q.C - MICROBIOLOGY Page No. 6 of 6

Pre!are t6e su--ar2 re!ort in t6e anne,ure ># an* su$-it t6is for re"ie54 a!!ro"al an* aut6ori;ation to )ali*ation Core .ea-. +,., APPEN#I6 10.1 A$$re"iations an* *efinitions A//re*iation #e7initions

10.# Referen es. 10.#.# 10.#.& 10.#.( 10.#.+ 10.#.6 10.& Anne,ure 10.&.1.1 Anne,ure > 1 CRe or*ing Of O$ser"ations 9or )ali*ationD. 10.&.1.# Anne,ure > # C'terilit2 .est )ali*ation Re!ortD. ++., REVA I#ATION CRITERIA: Re"ali*ation s6all $e arrie* out in ase of 11.1 If t6e -anufa turing !ro ess is 6ange*. 11.# If t6ere is an2 6ange in testing !ro e*ure 56i 6 is $eing use* for sterilit2 testing. Pre!aration Of Culture 'us!ensions. 'terilit2 .esting. Gro5t6 Pro-otion test. 'terilit2 .esting Area4 :ntr2 An* :,it Pro e*ure 'OP. O!eration Of :<uino,F Ma7eG Milli!ore. 'OP.