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Department: Quality Assurance

DOC. No. : AV/MB/SOP/001 Page No. : Page 1 of 3 00

Effective Date: 01.06.06


!u"#ect

evision No. :

!OP for isolation $ i%entification of !tap&. Aureus.

E)*!*ON! evision + 00 evise% Pages Description Original issue Date 01.06.06

Prepare% 'y Microbiologist

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Aut&ori(e% "y Factory Manager

Department: Quality Assurance

DOC. No. : AV/MB/SOP/001 Page No. : Page ( of 3 00

Effective Date: 01.06.06


!u"#ect

evision No. :

!OP for isolation $ i%entification of !tap&. Aureus.

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Purpose )o *efine proce*+re for Stap,ylococc+s a+re+s-isolation an* i*entification. !cope ),is proce*+re is applicable to isolate an* i*entify Stap,ylococc+s a+re+s. esponsi"ilities Microbiologist AM-/+ality Ass+rance Definition : SOP& Stan*ar* operating proce*+re. Proce%ure /.1 #noc+late sample on Bair*-Par0er me*i+m an* in salt me*i+m. /., #nc+bate t,e Bair*-Par0er me*i+m at 312! for at least 30 ,o+rs. /.- From t,e salt me*i+m3 ma0e s+bc+lt+res on t,e Bair*-Par0er me*i+m an* inc+bate at 312! for 30 ,o+rs. /.. $4amine not less t,an 5 of t,e s+spect colonies of Stap,ylococc+s on t,e soli* me*ia an* mar0 o+t as many s+spect colonies as possible to investigate. /./ 0est for i%entification /./.1 1ram2s stain ),e stain consists of a3 0.5 6 met,yl violet or crystal violet in 7ater. "3 #o*ine sol+tion816 io*ine an* ( 6 potassi+m io*i*e in 7ater9. c3 !o+nter stain 80.1g ne+tral re*3 0.(ml of 16 acetic aci* an* 100 ml of 7ater9. On a clean grease-free sli*e3 very lig,t an* t,in smear covering a small area is ma*e *irectly from li:+i* c+lt+re an* in clean tap 7ater if from soli* me*ia. ),e smear is fi4e* by passing to an* fro over a flame an* coole*. !over t,e smear 7it, t,e stain 8a9 for 30 secon*s. Approve% 'y Management Representative
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/./., /./././.. Prepare% 'y Microbiologist

Aut&ori(e% "y Factory Manager

Department: Quality Assurance

DOC. No. : AV/MB/SOP/001 Page No. : Page 3 of 3 00

Effective Date: 01.06.06


!u"#ect

evision No. :

!OP for isolation $ i%entification of !tap&. Aureus. /././ /./.6 Po+r off t,e stain an* 7as, 7it, 8b9 an* t,en cover 7it, 8b9 an* allo7 remaining for 30 secon*s. ;as, off 7it, et,anol +ntil t,e *ye ceases to stream o+t. ;as, in r+nning tap 7ater an* apply8c9 for abo+t one min+te. ;as, in tap 7ater an* *ry for e4amination.

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Coagulate 0est& ),is test may be carrie* o+t by t,e follo7ing met,o*s& a3 !li%e met&o% < $m+lsify a portion of t,e s+spect colony in normal saline or 7ater. Mi4 t,is 7it, a straig,t 7ire *ippe* in ,+man or rabbit plasma. !oag+lase-positive stap,ylococci pro*+ce visible cl+mping imme*iately. "3 0u"e met&o% < $m+lsify a single s+spect colony from a (= ,o+r gro7t, of bloo* agar me*i+m in 1 ml citrate* rabbit plasma *il+te* 1 in 5 in 0.>5 6 saline. ),e rest is +s+ally carrie* o+t in narro7 t+bes. Place in an inc+bator or preferably in 7ater-bat, at 312!. Observe every ,o+r to note clotting of plasma. Rea*ing s,o+l* be carrie* o+t for as long as possible3 preferably avoi*ing overnig,t inc+bation. Positive control 7it, a 0no7n coag+lase-positive strain of Stap,ylococc+s an* a control of t,e *il+te* plasma 7it,o+t inoc+l+ms s,o+l* be incl+*e* in t,e test

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Documentation: Microbiological )est Report

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eferences: #S& 5>>1 8Part ##9 < 1?1@

Prepare% 'y Microbiologist

Approve% 'y Management Representative


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Aut&ori(e% "y Factory Manager