SensitiveandFastDNALAMPinanEasy-to-UseMasterMixFormat

  • Loop-mediatedisothermalamplification(LAMP):Facilitatesrunningamplificationreactionsindifficulttestenvironments,enablinguseoutsideofahigh-techlaboratory.

  • MasterMixFormat:Simplifiesreactionsetupandminimizesoptimizationrequiredtoproducethebestassayresults.

  • HighThermalStABIlity:HighlyThermostableenzyme(upto90°Cfor5min)enablespreheatingofreactionscontainingpurifiedtargetDNAwhich,dependingonthetarget,mayincreaseassaysensitivityanddecreasetimetoresults.

  • ElevatedLAMPReactionTemperature(68-74°C):ImprovesprimerspecificityandreducesbackgroundamplificationdependingontheDNAtarget.

  • Freeze-dryCompatIBLe:Enablesgenerationofroomtemperaturestabletestkitsthroughlyophilization.

  • Custom OEM ProductCustomizable:OnceprimersareoptimizedwiththeLavaLAMP™DNAMasterMix,wecanworkwithyoutogeneratecustommastermixesfurtheroptimizedforyourtargetand/ortestneeds–contactus

TheLavaLAMP™DNAMasterMixisintendedtosimplifydevelopmentandoptimizationofDNALAMP(loop-mediatedisothermalamplification)reactions.LAMPkitsarecommonlyavailableasmulti-componentkitsthatrequireoptimization(e.g.MgSO4,betaine,enzymeaswellastemperature,primerconcentration,etc.).TheLavaLAMP™DNAMasterMixgreatlysimplifiesreactionoptimizationbylimitingoptimizationtotargetspecificcomponents/conditionssuchasLAMPprimerdesign,targetconcentrationandreactiontemperature.Additionally,thismastermixisheatstableat90°Cfor≤5minutes,whichenablestheadditionofareactionpreheatingstepwhichmay,dependingonthetarget,increaseassaysensitivityanddecreasetimetoresults.Needmoreinformation?SeeourFAQs.

Loop-mediated Amplification Process (LAMP)

Figure1.Simplifiedoverviewoftheloop-mediatedisothermalamplification(LAMP)process.Forsimplicity,4/6potentialLAMPprimersareillustrated.Theinclusionoftwoadditionalprimers,F-Loop;ForwardLoopPrimerandB-Loop;BackwardLoopPrimer,oftensignificantlyenhancesamplification.Formoreinformation,pleasevisittheEikenwebsite.


LavaLAMP-loop-mediate-amplification-reaction-easy

Figure2.IllustrationofthedifferencesinLAMPreactionsetupandpotentialoptimizationparameterswhenusingamastermixvs.acomponentkitformat.PanelAillustratesthecomponentsaddedtoaLAMPreaction(left)whenusingtheLavaLAMPDNAMasterMixandthevariousparametersthancanbeoptimized(right).PanelBalsoilllustratesreactionsetup(left)andpotentialoptimizationparameters(right)whenusingastandardLAMPcomponentkit.


Loop-Mediated Isothermal Amplification enzyme faster detection.

Figure3.Loop-mediatedisothermalamplification(LAMP)withreal-timefluorescentdetectionofamplifiedproducts.LAMPreactionsweresetupusingtheindicatedkitsaccordingtomanufacturer’srecommendations.TargetDNA(C.difficile)atvaryinginputamounts,tcdAtargetLAMPprimers,andGreenFluorescentDye(LavaLAMPKit)wereincludedinallreactions.ReactionswererunonaCFX96ThermalCycler(Bio-Rad)atthefollowingtemperatures:LavaLAMP;68°C;otherkitsattherecommended65°Candfluorescencewasmeasuredover60minutestodeterminetheTTR.NTC=NoTargetControl.


Thermostable LAMP Enzymn

Figure4.TestingthethermostabilityoftheLavaLAMP™DNAMasterMix.TwosetsoftriplicateLAMPreactionsweresetupusingtheindicatedamountsoftargetDNA(C.difficile),tcdAtargetLAMPprimers,andGreenFluorescentDye.Onesetofreactionswaspreheatedto90°Cfor5minutes,andthenallreactionswereincubatedfor60minutesonaCFX96ThermalCycler(Bio-Rad)at68°CandfluorescencewasmeasuredduringthecourseofthereactionstodeterminetheTTR.NTC=NoTargetControl.


LAMP Enzyme can be lyophilized

Figure5.TestingtheperformanceoflyophilizedLavaLAMP™DNAMasterMix.TheLavaLAMPDNAMasterMix(12.5µL)wasaliquotedintotubesandlyophilizedusingaVirTisWizard2.0.Afteroneday,eachlyophilizedtubewasreconstitutedin12.5µLwater.ThensixreplicateLAMPreactionsperconditionweresetupusingthereconstitutedlyophilizedmastermixandthestandardLavaLAMPDNAMasterMix(Controls).TheindicatedamountsoftargetDNA(S.aureus),clfAtargetLAMPprimers,andGreenFluorescentDyewereaddedtothereactions.Allreactionswereincubatedfor60minutesinaCFX96ThermalCycler(Bio-Rad)at68°CandfluorescencewasmeasuredduringthereactionstodeterminetheTTR.NTC=NoTargetControl.

 


Figure6.Analysisofdaytodayvariability.Onthreedifferentdays,sixreplicateLAMPreactionsweresetupperconditionusingtheindicatedamountsoftargetDNA(M13mp18),M13mp18targetLAMPprimers,andGreenFluorescentDye.OneachdaythereactionswererunonaCFX96ThermalCycler(Bio-Rad),amplificationwasmonitoredinreal-time,andthenresultswereanalyzedtodeterminetheTTR.NTC=NoTargetControl.

ORDERINFORMATION

BothLavaLAMP™DNAMasterMixkitscontain:LavaLAMPDNAMasterMix,DNAPositiveControlLAMPPrimerMix,andDNAPositiveControl.TheLavaLAMPDNAMasterMixwithDyealsocontainsGreenFluorescentDyeforfluorescentdetectionofamplifiedDNA.

Licensinginformation:LucigenisafullylicensedproviderofLAMPreagentsforresearchuse.PatentsWO00/28082,WO01/34790,andWO01/77317regardingtheLAMPmethodareownedbytheEikenChemicalCo.Ltd.LavaLAMP™,OmniAmp®andBstPolymerase,ExonucleaseminusaresoldbyLucigenunderlicenseforuseinLAMPforresearchuseonly.TheproductsmaynotbeusedforLAMP-basedhumanordiagnosticpurposeswithoutobtainingalicensefromEiken.USPatent8093030forLavaLAMPandOmniAmpisownedbyLucigenCorp.

Itisthesoleresponsibilityofthebuyertoensurethatuseoftheproductdoesnotinfringethepatentrightsofthirdparties.Ifthepurchaserisnotwillingtoaccepttheseuselimitations,LucigenCorporationiswillingtoacceptreturnoftheproductforafullrefund.