细胞培养规格及接种密度

™REFERENCEGUIDE–CELLCULTURER-10TrypsinVersene(mlof0.05%GrowthSurfaceAreaSeedingCellsat(mlof0.53mMtryspin,0.53mMMedium(mm2)DensityConfluency1EDTA)EDTA)(ml)Dishes35mm9620.331061.2310611260mm2,8270.831063.23106323100mm7,8542.231068.831065310150mm17,6715.0310620.0310610820ClusterPlates6-well9620.331061.23106223–512-well4010.131060.43106111–224-well2000.0531060.231060.50.50.5–1.0FlasksT-252,5000.731062.83106333–5T-757,5002.131068.43106558–15T-16016,0004.6310618.43106101015–30UsefulNumbersforCellCultureTherearevarioussizesofdishesandflasksusedforcellculture.Someusefulnumberssuchassurfaceareaandvolumesofdissociationsolutionsaregivenbelowforvarioussizeculturevessels.1Thenumberofcellsonaconfluentplate,dish,orflaskwillvarywithcelltype.Forthistable,HeLacellswereused.Bufferedsaltsolutionsforcellculturecanbedividedintotwoprincipaltypes:thosedesignedtoequilibratewithatmosphericconditions,andthosedesignedtoequilibratewithagasphasecontaining5%to10%carbondioxide.MediautilizingHanks’saltsaredesignedforatmosphericequilibration,baseduponthephysiologicallyrelevantpKaofphosphoricacid.Thesemediacontainbicarbonateionsforcarboxylationandsimilarbiologicalprocesses(1).UseofHanks’saltsinaCO2incubatorwillresultinarapiddropinpHoftheculturemedium.Thiswillcausethephenolredindicatortoturnyellow.MediabasedonEarle’ssaltsarebufferedwithabicarbonate/carbonicacidsystemandrelyuponthephysiologicallyrelevantpKaforcarbonicacidandtheequilibrationofgaseousandliquifiedcarbondioxidetomaintainthepHinaCO2equilibratedincubator(1).UseofEarle’ssaltsinatmosphericconditionswillresultinarapidriseinpHoftheculturemedium.Thiswillcausethephenolredindicatortoturnpurple.1.Jayme,D.W.andBlackman,K.E.(1985)Adv.inBiotechnol.Processes5,9.BufferSystemsforCellCulture