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1、BoneMarrowAspirationsinMiceIwillbepreformingbonemarrowaspirationsonalineoftransgenicswiththeintentofculturingthecells.Mylabhashadpriorissueswiththis,notgettingthesameyieldofcellsthatothershaveobserved.Previously,wehaveextractedthefemurofthemice,cleanedtheb
2、oneandthenflushedthemarrowoutwithPBSinanaspiratingneedle. Doesanyonehaveanytipsforthis?Asweallknow,itsthesmallthingsthattendtocount.Thanks!TOPICS·BoneMarrowTransplantation·BoneMarrowExamination·BoneMarrowSep18,2012Share ····1 / 0 ALLANSWERS (10)·MartijnH.
3、Brugman ·LeidenUniversityMedicalCentreHiAshtyn,I'vetriedBMaspirationinmicebefore,butfounditratherdifficult.Acolleagueofmine,whoismoreskilledatit,usesthetechniquethatisdescribedinVerlinden,ExperimentalHematology1998.Perhapsthisisuseful.Fromyourquestion,i'mn
4、otquitesure,butareyoutalkingaboutbonemarrowisolationafteryoueuthanizedthemice?Sep19,2012·ChristophMetzendorf ·UniversitätHeidelbergHiAshtyn,weroutinelydothistomakebonemarrowderivedmacrophages.Iseesomefluctuationsincellcounts(reachingfrom2.25x10^6to3.5x10^6
5、cells/mLin10mLofbuffer)butgenerallytheyarenoproblemaswecountcellsandthenseedthesamenumberofcellsperwell.WeextractBMfrombothfemursandtibiasofeachmouse. Onethingthatseemstoaffectcellcountsareageofmiceandstrain.Ifyouareinterestedinamoredetailedprotocol,Iuploa
6、dedithere.oBMDMprepandculture.pdfSep19,2012·HendrikGremmels ·UniversityMedicalCenterUtrechtHi, TheprotocolsuppliedbyChristophMetzendorfisverysimilartowhatwearedoing.Therearesomesmallvariationsacrossprotocols:somepeopleuseRPMIforflushingtheboneswiththeideat
7、hatitleadstohigherviability.Itis(slightly)moreexpensivehoweverandyouhavelessvisualfeedbackofthemarrow'clearingup'ifit'sgotphenolredinit. Forcuttingupthebonesweuseasurgicalwirecutter,soyoucancutrightthroughtheheadsandwasteaslittleaspossible.Youcandomorebone
8、sifyouneedmorematerial,youshouldbegettingaround20Mcellsfromthetwofemurs,another10Mfromthetibiaandmaybe5Mfromthehumerialthoughmileagemayvary. Ofcoursethisisalterminalforthedonormouse.I'mnotsureiftherearemethod
