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1、小鼠脾脏细胞培养PRIMARYCULTUREOFMOUSESPLEENCELLS&IDENTIFICATIONOFLIVEORDEADCELLSBYDIFFERENTIALSTAININGQianqianChen(118627140313)Teammembers:QingqingYu,QingZhangApr订7th2012AbstractCellsfrommuti-cellularorganisincanbeisolatedandculturedaliveinvitro,andkeepitsph
2、ysiologicalandbiochemicalproperties.Tovertifyit,ourteamculturedthemousespleencellsinvitroandfondthatthoughthecellsweremicrobio1ogicalcontaminated,thecelIssomehowsurvived,thusweprovedtheconclusion.Consideringourcellsweremicrobiologicalcontaminatedwecou
3、ntthecellsfromotherteamstolearnthemethcodofcountingcellstokeeptheexpcriment.completed・IntroductionCellsculturecanbedividedintoprimaryculture,subcultureandtheformationofcell1ines.CelIsdierctlyis1otedFromorganismwithoutproliferationinvitroarecalledprima
4、rycultures.Ifprimaryculturescontinuetodividandproliferatetoformmorecells,thenthecellswillbecalledsubculturedcells.Usuallythesubcultredcellswillgotodeathafterafinitenumbersofgenerationsinvitro,thisprcessiscalledcellsenescence,howeverifthesubcultredcell
5、saretransforedandbecomeimmortal,thentheywillbecomecelllines.Accordingtothedifferenceofcellmembranepernliabilityfromthedeathandthesurvives,somedyescanpermeatethemembianeofthelivecells(0.15%EosinY),butsomecannot(1%(w/v)Trypanblue).Allcellscanbecolouredb
6、y0.15%EosinY,butonlythedeadcellscanbecolouredby1%(w/v)Trypanblue.Thus1%(w/v)Trypanblueand0.15%EosinYcanbeusedtodistingwishthedeadcellsfromthelivecells.MaterialsandMethodsMethods1.Sacrificemousebyneck-breaking(cervicaldislocation).1.Soakthemousein70%et
7、hanolforafewminutes.exposethemusclelayer・Cutthemusclelayerexposeperitonealcavity・sideoftheabdomentoexposespleenwithscissorsandputit2.Laythemousefaceup,wettheabdomenwithwater,lifttheoutskinwithforcepsandcutahorizontaiopeningwithscissors,teartheoutskina
8、parttoopeninaVshapeandliftupto4.Pushasidethestomachintheupper-leftthesack-shaped,dark-redspleen.Cutofftheintotheglassdish・intothespleenalongthelonger5.Inject0.2mlPBSwiththeL-shapeneedleaxis.Spleenwillbloatandbecomepale.*Nottopenetratethespleen
