Description:
Basedonourinnovativeandproprietarylipid-conjugationtechnology,LipoJet™TransfectionKit,formulatedfromnovelfluorinatedcationiclipids,exhibitssignificantdifferencefromotherlipidstransfectionreagentsinthemarket.LipoJet™TransfectionKitisthemostpowerfulyetverygentlegenedeliverytoolforavarietyofapplicationsincludingplasmidDNAand/or siRNAformostofmammaliancelltypes. ComparedwithleADIngproductsinthemarket, LipoJet™ismore cost-effectiveandalways provideshighertransfectionefficiencywithlesscytotoxicity.
Kitcontent:
-LipoJet™Reagent,1.0mlsufficientfor1000DNAtransfectionsand1000siRNAtransfections (24-wellplate).
-LipoJet™TransfectionBuffer(5x),8.0mLtomake40mLofworkingsolution(1x).
Features:
-Versatile:DNAtransfection,siRNAtransfectionandDNA/siRNAco-transfection.
-Powerful: Outstandingefficiencyonadherentcells.
-Extremelylowtoxicity: ImproveCellviABIlityaftertransfection
-Economical:Moretransfectionswithlessreagent
-Easytouse:OnetubereactionandcompatIBLewithserum/antibiotics.
-Suitableforhigh-throughputapplication
Storage:
Storeat4°CforLipoJetReagentandRTforLipoJetTransfectionBuffer(5x). Ifstoredproperly,theproductisstablefor12monthsorlonger.
BroadTransfectionSpectrumforMammalianCellTypes
CellLine | DNATransfection | siRNATransfection |
293,293T | 85% | - |
ExamplesShowingExcellentDNAandsiRNATransfectionEfficiencyofLipoJet™InVitroTransfectionKit
AefficiencycomparisonofLipoJet™reagentvs.brandnameproductstotransfectHela,Cos-7,NIH-3T3,CHOandHEK293(leftpanel)andCos-7(rightpanel). pEGFP-N3CDNA(0.5µg/wellof24-wellplate)wastransfectedintodifferentmammaliancellsperthestandardtransfectionprotocolsinpresenceofserum(10%FBS)andantibioticsasrecommendedbymanufacturers. ThetransfectionefficiencywereanalyzedviaFACS48hoursposttransfection.
AtoxicitycomparisonofLipoJet™reagentvs.brandnameproductstotransfectHelacells. pEGFP-N3cDNA(1.0µg/wellof24-wellplate)wastransfectedtoHelacellsperthestandardtransfectionprotocolsinpresenceofserum(10%FBS)andantibioticsasrecommendedbymanufacturers. TheMTTassay(rightpanel)andphasecontrastimaging(leftpanel)wereusedtoanalyzethecellviability48hoursposttransfection.
AcomparisonofLipoJet™reagentvs.Lipofectamine2000(L2K)andPolyJet™transfectionreagentonHEK293Tcell.pEGFP-N3cDNA(0.125µg/well,0.25µg/welland0.5µg/perwellof24-wellplate)wastransfectedinto293Tcellsusingthestandardtransfectionprotocolsinpresenceofserum(10%FBS)withLipoJet™(upperpanelatLipoJet™/DNA(µl/µg)ratio=2),L2K(middlepanelatL2K/DNA(µl/µg)ratio=3)andPolyJet™(lowerpanelatatPolyJet™/DNA(µl/µg)ratio=3)respectively.ThecellswerevisualizedbyNikonEclipseFluorescencemicroscope48hoursposttransfection.
AcomparisonofLipoJet™reagentvs.Lipofectamine2000(L2K)andPolyJet™transfectionreagentonHelacell. pEGFP-N3cDNA(0.125µg/well,0.25µg/welland0.5µg/perwellof24-wellplate)wastransfectedintoHelacellsusingthestandardtransfectionprotocolsinpresenceofserum(10%FBS)withLipoJet™(upperpanelatLipoJet™/DNA(µl/µg)ratio=2),L2K(middlepanelatL2K/DNA(µl/µg)ratio=3)andPolyJet™(lowerpanelatatPolyJet™/DNA(µl/µg)ratio=3)respectively.ThecellswerevisualizedbyNikonEclipseFluorescencemicroscope48hoursposttransfection.
ExceptionalgenesilencingofLipoJet™reagentduringDNA/siRNAco-transfectiononmammaliancells.TransfectionofGFPcDNA(leftpanel,0.25µgperwellof24-wellplate)andco-transfectionofGFPcDNA(0.25µgperwellof24-wellplate)andGFPtargetedsiRNA(final10nM,rightpanel)withLipoJet™reagentleadstoexceptionalgenesilencingonHEK293(upperpanel),Hela(middlepanel)andSaoS-2(lowerpanel)cellsrespectively.ThecellswerevisualizedbyNikonEclipseFluorescencemicroscope48hoursposttransfection.
ExceptionalDNA/siRNAco-transfectionefficiencyonHUVEC.Co-transfectionofmCherrycDNA(0.10µgperwellof24-wellplate)andFITCconjugatedsiRNA(final30nMperwellof24-wellplate)toHUVECwithLipoJet™reagentgaveriseto60%mCherry+(phasecontrastoverlappedwithmCherryimaging,leftpanel)andnearly100%FITC-siRNA+(phasecontrastoverlappedwithFITCimaging,rightpanel)HUVEC24hoursaftertransfection. ThepicturesweregivenfromDr.PanKongofUSCascourtesy. 
ExcellentsilencingofendogenouslyexpressedKIF11(alsoknownasEG5)inHEK293(upperpanel)andHela(lowerpanel)cellswithLipoJet™reagentat10nMEG5siRNA.KIF11(alsoknownasEG5)encodesamotorproteinthatbelongstothekinesin-likeproteinfamilyinvolvedinchromosomepositioningandbipolarspindleformationduringcellmitosis.AreductioninKIF11levelscausesmitoticarrest.LipoJet™reagenteffectivelydeliversEG5siRNA(final10nM)toHEK293andHelacells,leadingtomorethan80%of"round-up"phenotypeofHEK293andHelacells24hposttransfectionovernegativecontrol(final10nMwithshamEG5siRNA).Thephenotypeof"rounded-up"HEK293andHelacellswerevisualized24hposttransfectionwithaNikonmicroscope.
AimageshowingexceptionaltransfectionefficiencyofLipoJetTransfectionKitonHumanembryonicstemcells (hESCs). ThehESCsgrowninE8mediumonGeltrexvs(leftpanel,DICimaging)wastransfectedwithpEF1α-GFP.TheGFPexpression(rightpanel)wasvisualizedbyNikonEclipseFluorescencemicroscope24hoursposttransfection.TheabovepictureswereprovidedbyDr.MarinaPryzhkovaofJohnsHopkinsUniversityascourtesy
DataSheet&Protocol:
-AProtocolforDNAandsiRNATransfection
-AShortProtocolforDNATransfection
-AShortProtocolforsiRNATransfection
-AProtocolforDNA/siRNACo-transfection
- TechnicalNote&TransfectionTips
Weareoffering"WirelessPowerPointPresenter"tocustomerswhopurchase5x1.0mLLipoJet™reagentbyDecember2014.Takeadvantageofitanddon"tletthischancego...
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FG-3:WirelessPowerPointPresenterwithCaseforpurchaseof5X1.0mlormore.Thispresenterisofferedforlimitedtimeonly! |
Torequestafreetrialsample,pleaseCreateAnAccountwithustoenteryourshippingaddressandemailusatorder@Signagen.com
Testimonials:
Sorryforsooobigdelay,butnowIamtotallyinlovewithbothPolyJetandLipoJet.RecentlywediscoveredthatLipoJethassupergoodtransfectionefficiencywithmESCsandhESCswithi muchlesstoxicity.LastweekItransfectedovernighthumanESClineH1usingLipoJetwithsameamazinglyhighefficiency-upto50~70%.DNAconstructincludesGFPunderEF1apromoter(CMVpromoterdoesnotworkinhESCs).ThisisthebesttransfectionefficiencyIeversawwithESCs.Lipofectamin3000didn"tgivemetheseresults(usedonmouseESCs).
--------MarinaPryzhkova,Ph.D.,JHU
Itriedtheplasmid/siRNAco-transfectionusingtheLipoJetsampleinEAhy926cellwhichisaHUVECcellline.Theresultsareprettygood,atleastforthetransfectionefficiency.Fortheplasmidtransfection,48hourslater,around60%cellaretransduced.ForsiRNA,theefficiencyisalmost100%.WeareorderingmoreLipoJetreagenttousefromnow.
-----Dr.PanKong,UniversityofSouthernCalifornia
Wegot>90%efficiencywithLipoJetvs.80%withX-tremeGENE9on293Tcell.Definitelywillordermore....
-----Dr.NuoYangfromRoswellParkCancerInstitute
HerearetheresultsfromourpreliminaryexperimentswithLipoJet.Weareverysatisfiedwithitsefficiency.Unfortunately,wewereoutofFugene6/HDandwereunabletotestthemsidebyside,butbasedonpreviousexperience,wedobelievethatyourproductworkedwithbetterefficiency.
------Dr.AlisonMcKelveyfromUniversityofPittsburgh
IamhappytosaythatIhadgreatsuccesswithyourtransfectionreagentsonOKF6/TERT2humankeratinocytes.IwillputtogetherthedataonceIhavecompletedmyanalysis.IreallyliketheLipoJet.MycellofinterestdidnotdoverywellintheGenMutealthoughmycontrolcellsdid.WewanttogoaheadwithorderingASAP.Ialsohaveacouponfor10%foraPOorder.
-----Dr.MichelleSimpson-AbelsonfromUPMC
WedidindeedtestthemsidebysidewithourhomemadeCaPhos.Theresultsaregreat, LipoJetandCalFectinbothdidextremelywell.LipoJetbeingthebetterofthetwo.Iwillsendyoutheresultswhen Igetthem, Iamcurrentlyoutofthelabbutwewillbeorderingmoreforcertain.Thankyouforsendingthosesamples.
------Dr.AhmedHassib,CornellUniversity
FortheLipoJet,IonlycompareditwithLipoLTXinHelacells.TheLipoJetgaveamuchhigherefficiency(~35%betterthanLipoLTX)24hoursaftertransfection.WewillordersomeLipoJet.
-----AbetatesterfromUSC
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但是有在转染前，将细胞进行重新传代的情况，这样做的目的在于保持细胞的活性状态。
“转染前将细胞以1.5-4.5X104 cells/well 的量(具体接细胞数请参考表1）接种在孔板中，于37℃， 5%CO2 的条件下进行培养，18-24小时（ sf9细胞为3-4小时）后转染。”
其次要看下你选择单位的规模如何，上海这边的，你可以看下基尔顿生物，原代细胞培养，动物造模，整体课题外包。
我刚开始做转染，悬浮细胞，分别做过表达和敲减，看了很多文献，大都没有提及转染后是用转染的这同一批细胞同时做pcr，wb，cck8，凋亡，细胞周期；还是说这次转染只做pcr或wb，再转染一次做cck8或细胞周期。剩下的功能试验均同前，转染一次做一次？我养的是悬浮细胞，转染后做cck8这些功能试验前需要离心换液吗？跪谢解答！
目标蛋白对细胞有毒性，导致细胞死亡；
转染试剂以及DNA用量信息需要优化，否则对细胞具有伤害；
细胞贴壁转染之后没有正常换液。
建议：考虑对目标蛋白进行截短构建、尝试其他细胞系统；摸索转染试剂以及DNA用量信息，如果转染试剂毒性太大，可以考虑尝试义翘转染试剂sinofection；对转染后的细胞进行换液处理，如果细胞状态感觉不够理想，可以考虑添加一些血清来帮助细胞恢复健康。
以上所有分析、建议的前提是，细胞培养、无菌操作等等都没有问题。祝顺利，加油~
转染分2种，一种是瞬时转染，即转染后让细胞表达目的蛋白后即提取蛋白，提一次蛋白，转染一次，这种方式一般不传代；
另一种转染为稳定转染，转染后加入一定选择压力进行筛选，没有转染的细胞不能存活，只留下转染的细胞，这种情况下可以筛选单个转染细胞，构建稳定表达某一特定蛋白或基因的细胞系。
各位版友求助，
我使用Hek293构建转染模型，瞬转5质粒，用lipo2000做转染体系。
转染48h，发现荧光较强的细胞都在爬片的边缘，比例十分少。是因为我添加试剂的手法不对吗。
同时也发现加入转染体系后细胞状态特别差。想问一下用lipo2000时可以用无双抗的10%FBSDMEM吗。我转染前6h现在用的是纯DMEM，不含FBS。
请各位大神帮忙
不建议你用MDCK细胞，非常难转染。
做的比较好的，一般都是上海地区的，你可以看下基尔顿生物。
