Ultra-Fast and Optimized Method for the Preparation of Rodent Testicular Cells for Flow Cytometric Analysis 英文参考文献.docVIP

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Ultra-Fast and Optimized Method for the Preparation of Rodent Testicular Cells for Flow Cytometric Analysis 英文参考文献.doc

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Ultra-Fast and Optimized Method for the Preparation of Rodent Testicular Cells for Flow Cytometric Analysis 英文参考文献

Ultra-FastandOptimizedMethodforthePreparation ofRodentTesticularCellsforFlowCytometricAnalysis RosanaRodríguez-Casuriaga,AdrianaGeisinger,BeatrizLópez-Carro, ValentinaPorro,RodolfoWettstein,andGustavoA.Folle Abstract Homogeneity ofcellpopulations isaprerequisite fortheanalysis ofbiochemical andmolecular events duringmalegametedifferentiation.Giventhecomplexorganizationofthemammaliantesticulartissue, variousmethodshavebeenusedtoobtainenrichedorpurifiedcellpopulations,includingflowcellsort- ing.Currentprotocolsareusuallytime-consumingandmayimplylossofshort-livedRNAs,whichisun- desirable for expression profiling. We describe an optimized method to speed up the preparation of suitabletesticularcellsuspensionsforcytometricanalysisofdifferentspermatogenicstagesfromrodents. Theproceduretakesonly15minincludingtestisdissection,tissuecutting,andprocessingthroughthe MedimachineSystem(BectonDickinson).Thismethodcouldbeasubstituteforthemoretediousand time-consumingcellpreparationtechniquescurrentlyinuse. Keywords: Flowcytometry,Geneexpression,Spermatogenesis. 1.Introduction Spermatogenesisisacomplexdifferentiationprocessessentialforall thespecieswithsexualreproduction,whichleadstotheformationof malegametes.Inspiteofitsimportance,adeepercomprehensionof themolecularbasesisstillrequiredfortheunderstandingofthefun- damentalsofnormalsexualreproduction,aswellasforthetreatment oftesticularpathologies. Electronicsupplementarymaterial:Theonlineversionofthisarticle(DOI:10.1007/s12575-009-9003-2)containssupplementary material,whichisavailabletoauthorizedusers. ShulinLi(ed.),BiologicalProceduresOnline,Volume11,Number1 ?totheauthor(s)2009 DOI:10.1007/s12575-009-9003-2URL:; 184 Ultra-FastMethodforPreparationofTesticularCellSuspensions 185 Spermatogenesis can be divided into three phases: mitotic proliferation of spermatogonia, meiotic divisions of spermato- cytes, and spermiogenesis. Meiosis is a successful evolutionary widespreadmechanismpresentinalleukaryoticspeciesfromfungi tosuperi