Duringthepastfivedecades,varioustypesofchemistrieshavebeenusedforconjugationofmoleculessuchasantibodiestothesurfaceoftheliposomes.Ingeneral,theconjugationcanbeachievedthroughtheN-terminus,theC-terminusortheavailablesulfur(e.g.Fab’fractionorthiolatedAb).NotallchemistrieshavethesameyieldandefficiencyofconjugationandoftenreproducingbiocompatIBLebatchescanbeachallenge.Theliposomescontainingpyridyldithiopropionate(PDP)lipidsareusedtoconjugateproteins,antibodiesandothermoleculescontainingthereactivemoiety.PDPlipidsarenotaswidelyusedasmaleimidelipids,buttheydohavetheirownnicheapplication.ThePDPgroupcontainsdisulfide,whichcanreactwithsulfhydrylorthiolatedproteins/antibodies.Therefore,PDP-functionalizedliposomescanbeusedintwoways:
MethodA.Inthisapproach,thepyridyldithiogrouponthedistalendsofthePEGchainscontainsPDPisforestreducedbyareducingagent(dithiothreitol,DTT).Maleimide-containingantibodiesarethenefficientlycoupledtothesurfaceofliposomes.Thethiol-maleimideprocedureisoneofthemostdesirablereactionsinbioconjugatechemistryduetoitssimplicityandhighcouplingefficiencyinaqueoussolution.Thereaction,whichisbasedonthestablethioetherlinkagebetweenathiolgroup(reducedformofPDP-liposome)andthecorrespondingmaleimidegroup,occursselectivelyandirreversiblyatneutralpH(6.5-7.5),andtheformedbondsarenotcleavedbyreducingagents.Inaddition,duetothepresenceoftwodifferentoxidationstatesofsulfurresidues(oxidizedandreducedstatesasadisulfidebondandsulfhydrylgroup,respectively)onthetwoconjugatingcomponents(i.e.,theliposomeandprotein/antibody),theprobABIlityofthecrosslinkingofthehomologousagentsislow.Therefore,protein-proteinandliposome-liposomecrosslinkingdoesnotusuallyhappen.
MethodB.Alternatively,thePDPgroupcanparticipateindisulfideexchangereactionswiththiolspresentontargetingproteins/antibodies.Thecouplingreactionisfastandconductedundermildconditions.However,theformeddisulfidebondshavebeenreportedtobelessstablethanthioetherbonds.Moreover,eveninanalkalinemedium(pH8.0),thiolgroupsareoxidized.Thedisulfidebondformedbetweentheprotein/antibodyandliposomescanalsobebrokeninthepresenceofareducingagentandtherefore,theconjugationreactionisreversible.
MethodA. Conjugationofmaleimide-modifiedantibodytoaPDP-modifiedliposome.
MethodB.Conjugationofathiol-modifiedantibodytoaPDP-modifiedliposome.
ImmunoFluor™-PDPisaPEGylatedproduct.Forotherreactive(PEGylatedandnon-PEGyalatedproducts)ImmunosFluor™productssuitableforothertypesofconjugationmethodsee here.
