MycoplasmatestsaccordingtotheEuropeanPharmacopoeia(EP)Chapter2.6.7requireasensitivityof10colonyformingunits(CFU)permlsamplevolumeforNAT-basedmethodssuchasPCRinordertoreplacetrADItionalculturemethods.Thissensitivitymustbeaccomplishedaspartoftherobustnesstestingforanyparticularsamplematrix.
Usingvitalmycoplasmaisnotacceptableforthemajorityofcellculturelaboratoriesduetosafetyregulations.Our10CFU™sensitivitystandardscontainnon-vitalmaterialandallowsafeandreliablevalidation.
ThemycoplasmaweregrowninculturemediumasdescribedinEP2.6.7,subsequentlytitratedandplatedonHayflickandFreymediumforCFUdetermination.Themycoplasmawereharvestedintheearlylogarithmicgrowthphasetoensureahighratioofvitaltonon-vitalmycoplasmaandthusalowratioofGU*toCFU.AllstrainswereobtainedfromtheNationalCollectionofTypeCultures(NCTC),UK,andcultivatedinlowpassages.
10CFU™SensitivityStandardsareintendedforvalidatingtherobustnessandsensitivityofNAT-basedmycoplasmatests.
Unitpackage:3vialscontaining10cfuoftheparticularmycoplasmaaswellas3vialsnegativecontrol
Setpackage:2vialsofeachEP2.6.7-listedmycoplasmacontaining10cfu(18vialsintotal)aswellas2vialsnegativecontrol
Thecomponentsarelyophilized.
ApplicationofaPCR-basedmycoplasmatest,e.g.Venor®GeMqEPMycoplasmaDetectionkit
Sterile1.5mlreactiontubes,filtertips,appropriateDNAextractionkit
Microcentrifuge,microPipettes
Thedateofexpiryisgivenoneachpackage.Theunopenedkitshouldbestoredat+2to+8°C.
Figure–10CFUwerere-sUSPendedin1mlcellculturemedium,200µlwereusedforDNAextractionwiththeVenor®GeMSamplePreparationKit.ThemycoplasmaPCR-testwasperformedwiththeMicrosart®ATMPMycoplasmakitandaCorbettRotorGene®6000cycler.
SouthernBiotech/Goat Anti-Human Lambda-Alexa Fluor® 555/2070-32/1.0 mg
vectorlabs/Biotinylated Aleuria Aurantia Lectin (AAL)/B-1395/1 mg
vectorlabs/VECTASTAIN® Elite® ABC-HRP Kit (Peroxidase, Mouse IgG)/PK-6102/1 kit
vectorlabs/VECTASTAIN® Elite® ABC-HRP Kit (Peroxidase, Universal)/PK-6200/1 kit
vectorlabs/Unconjugated Musa Paradisiaca (Banana) Lectin (BanLec)/L-1410/5 mg
GiottoBiotech/Calmodulin N60D/5 mg/G02CLM60cn