ICS71.100.70

T/SHRH032-Anti-wrinklyandFirmingLiftingEfficacyTestofCosmetics-InvitroTestMethodofReactiveOxygenSpecies(ROS)Inhibitionwithkeratinocytes 上海日用化學(xué)品行業(yè)協(xié)會T/SHRHT/SHRH032-T/SHRHT/SHRH032-目前 1234567891011121314GB/T1.1—2020《標(biāo)準(zhǔn)化工作導(dǎo)則第1T/SHRHT/SHRH032-T/SHRHT/SHRH032-GB/T6682—2008SN/T3899—2014SN/T2328—2009化妝品急性毒性的角質(zhì)細(xì)胞試驗。humanreactiveoxygenfluorescence2',7'-二氯熒光黃雙乙酸鹽(2',7'-Dichlorodi-hydrofluoresceindiacetate，DCFH-DA)無自發(fā)平成正比。過氧化氫（H2O2）ROS精度0.1mg?CO2培養(yǎng)箱：375%CO2???可調(diào)節(jié)移液槍：1000μL、200μL、50μL、10μL???GB/T6682P7過氧化氫（H2O2）ROS250μM，具體刺激濃度也可根據(jù)試驗室培養(yǎng)細(xì)胞特點或選用的細(xì)胞系特性進(jìn)ROSROS0.05E（配置見附錄據(jù)歷史資料，試驗可接受標(biāo)準(zhǔn)為：陽性對照ROS30/EDTA[3]T251mL，T753mL。的規(guī)格可根據(jù)試驗需求進(jìn)行調(diào)整，1224662mL，12241mLCO224h±2h62mL，12241mLCO224h±2h30min250μM。PBS31mLDCFH-DACO230min，孵育結(jié)束后，PBS3T/SHRHT/SHRH032-T/SHRHT/SHRH032-3000r/min3min1mLPBS，3000r/min3min500μLPBSFITC參照9.1細(xì)胞消化步驟，進(jìn)行細(xì)胞消化及終止操作，終止結(jié)束后，將細(xì)胞收集至1.5mLEppendorf，3000r/min3min，棄掉上清液。FITC488nmImageJImageProPlusT?抑制率(%)=（1C?

）×100%…………………SD統(tǒng)計測得的各組平行孔間熒光強度的標(biāo)準(zhǔn)差（StandardDeviation，SD）ROS受試物與陰性對照相比，ROS（P<0.05），說明受ROST/SHRHT/SHRH032-T/SHRHT/SHRH032- ──1~4 ──30μg ──15 42.5μL325mLPBS1.5mmol/L250μmol/L?，F(xiàn)用現(xiàn)配。0.05%VEVE5%（m/v），1:1000.05%?，F(xiàn)用現(xiàn)T/SHRHT/SHRH032-RajindarS.Sohal,WilliamC.OrrTheredoxstresshypothesisofaging.FreeRadicalBiology&Medicine52(2012)539-555.MichelleVaz,NarsaMachireddy,AshleyIrvingOxidant-InducedCellDeathandNrf2-DependentAntioxidativeResponseAreControlledbyFra-1/AP-1.MolecularandCellularBiology.1694–1709[3]Coecke,S.Ballla,M.Bowe,G,Guidanceongoodcell