Tus kab mob-nyiam cov tshuaj tiv thaiv particle thiab nws cov txheej txheem hauv qab lim dej
Tus kab mob zoo li particle (vlp) yog cov tshuaj tiv thaiv tshuaj tiv thaiv kab mob uas yog nyob ntawm tus qauv kab mob tus kab mob tab sis tsis muaj cov khoom muaj caj ces. Lawv txhawb lub cev muaj zog teb los ntawm kev tiv thaiv morphology thiab cov kab mob ovigens ntawm cov kab mob ntuj, thiab muab kev nyab xeeb ntau dua vim lawv cov kev tsis muaj peev xwm yuav replicate. VLP cov tshuaj tiv thaiv yog ib qho zoo dua lwm txoj kev txhaj tshuaj VLP tshuaj tiv thaiv tsis tuaj yeem luam tawm hauv lub cev, yog li nws yog qhov haum rau txhua tus neeg, suav nrog cov poj niam cev xeeb tub, suav nrog cov poj niam cev xeeb tub, suav nrog cov poj niam cev xeeb tub, suav nrog cov poj niam cev xeeb tub lossis cov neeg uas muaj lub cev tsis xis nyob. Tsis tas li ntawd, capsid cov protein (suav nrog lwm cov kab mob uas tsis muaj kev txhawj xeeb tsis muaj kev txhawj xeeb txog cov tshuaj tiv thaiv caj ces zoo li kev txhaj tshuaj tiv thaiv kab mob Thib ob, VLPS muaj peev xwm qog cov kab mob ntawm cov kab mob uas tsis zoo, uas cov tshuaj proteins tsis zoo, uas tau hloov cov proteins ua ke yuav raug hloov pauv thaum lub sij hawm ua kom tsis taus, ua rau immunogenic tsis zoo.
Daim duab. 1. Tsim kev ua tshuab ntawm VLPS thiab Chimeric VLPS
Cov nta ntawm VLP txhaj tshuaj
1. Cov qauv zoo sib xws: VLPS yog tus kheej sib sau ua ke los ntawm kis tus qauv ib lossis ntau dua li capsid protein), thiab lawv qhov loj thiab cov duab zoo ib yam li tus kab mob sib txawv.
2. Tsis muaj kev kis tau: Tsis muaj tus mob genome, tsis tuaj yeem replicate lossis ua rau mob.
3. Kev immunogenicity: qauv granular tuaj yeem lees paub los ntawm kev tiv thaiv kab mob ntawm lub cev tau zoo thiab ua kom lub cell thiab t cell teb. Tuaj yeem ntxias cov tshuaj tiv thaiv nruab nrab thiab tshuaj tiv thaiv kab mob ntawm tes (piv txwv li, tshuaj txhaj tiv thaiv HIV, HPV).
4. Kev Nyab Xeeb Kev Nyab Xeeb: haum rau cov neeg muaj kev tiv thaiv tsis muaj zog (xws li Kab Mob Siab B VLP txhaj tshuaj).
Daim duab. 2. Muaj cov tshuaj tiv thaiv VLP
VLP Tshuaj Tiv Thaiv Ntau Lawm
VLPS tuaj yeem tsim los ntawm ntau cov ntsiab lus hais tawm, ib txwm muaj lub platform suav nrog:
1. Kab hlau-Baculovirus system:
Cov txiaj ntsig zoo: cov txiaj ntsig siab, tus nqi qis, haum rau cov protein nyuaj.
Daim ntawv thov: HPV tshuaj tiv thaiv, tshuaj tiv thaiv kabmob Ebola.
2. Mammalian hlwb (piv txwv li hek293 hlwb):
Qhov Zoo: Kev txhais lus hloov kho yog ze rau tib neeg thiab haum rau hauv VLPS (xws li tshuaj tiv thaiv khaub thuas npaws).
3. Cov ntsiab lus poov xab (piv txwv li Pichia pastoris):
Cov txiaj ntsig zoo: Ceev, tus nqi qis, tau siv nyob rau hauv cov tshuaj tiv thaiv kab mob siab B.
4. Cog tshuab (piv txwv li luam yeeb chloroplasts):
Tus phooj ywg ib puag ncig thiab cov pa taws, nyob rau hauv kev txhim kho (piv txwv li Norovirus VLP tshuaj).
Txheej txheem los ntawm kev siv tshuaj tiv thaiv VLP
Qhov sib txawv ntawm VLPs qhia cov qauv coj mus rau cov liv dej ntawm cov txheej txheem downstream.
Daim duab. 3 Ib txoj kev dav hlau ntws tawm
Sau thiab piav qhia:
Hauv txhua hom kev coj noj coj ua, kev tawm tswv yim los ntawm cov kab mob cell, muaj ntau cov kab lis kev cai me me, yog tias muaj cov kab lis kev cai me me, yog tias kev ua kom tiav thiab cov ultrafiltration membrane yog seem. Yog li, nws yuav tsum tau qhia meej ua ntej kev pom zoo, kom tshem tawm cov kab ke ntawm cov kab lis kev cai tuaj yeem ua rau cov txheej txheem kab lis kev cai tuaj yeem ua rau cov txheej txheem kev ua haujlwm ntawm lub neej ultrafiltration membrane. Kev Hais Qhia Ua Ntej Confrtration ntawm Kev daws teeb meem yog qhov txuas tseem ceeb txuas rau cov txheej txheem ntawm kev xav.
Cov txheej txheem piav qhia tau zoo yuav tsum tau muab cov khoom muaj txiaj ntsig zoo los tshem tawm cov khoom siv tau yooj yim, thiab kev tiv thaiv yooj yim, thiab kev tiv thaiv kom qis rau cov chav ua haujlwm.
Txojkev
Centrifugation: Cov colleial centrifugation tshem tawm cov ntawv loj.
Kev lim dej tob: Siv cov multistage lim (piv txwv li 1.2μm → 0. 45μm) kom paub meej tus supernatant.
Tangential Pom kev (TFF): Haum rau huab hwm coj ntau lawm, feeb thiab tshem cov khoom me me ntawm kev tsis sib xws.
Txawm li cas los xij, yog tias cov nplai ua haujlwm loj dhau, centrifugation siv sijhawm ntev, muaj ntau tus lej ntawm cov hmoov av lim dej ntev, yog li tus nqi ntawm cov neeg muaj txiaj ntsig zoo ntxiv. Nyob rau tib lub sijhawm, qhov siab ceev ntawm cov kab lis kev cai ntawm tes yuav txo tau cov kev thauj khoom ntawm lub siab lim ntawv membrane, uas ua rau cov nqi nce thiab ua haujlwm ntau dhau.
Muaj ob hom TFF membrane Cheebtsam: tiaj Cassette thiab Hollow Fiber. Tangential Pom kev lim (Tff) yog tsav los ntawm transmembrane siab txawv. Cov tshuaj thiab cov impurities uas me dua li tus membrane pore dhau los ntawm daim nyias nyias, thaum cov khoom siv hluav taws xob xws li cov ntsiab lus loj yog cuab. Lub pore loj ntawm daim nyias nyias siv rau microfiltration yog {0}. 45 / 0.22μm. Hollow Fiber tuaj yeem ua tiav cov txiaj ntsig siab cov ntsiab lus ua kua ncaj qha, nws tuaj yeem tshem tawm cov peev txheej ntawm centrifugation thiab cov nqi ua ntej ntawm Centrifugation
Ultrafiltration converration (Tff):
Hom Phiaj: Txhawm rau txo cov nyiaj ntawm Chromatographic, txhim kho cov chromatographic efficiency thiab tiv thaiv cov chromatographic kem.
Nyob rau hauv dav dav, ultrafiltration (Tff) muaj peev xwm ntau dua 100 lub sij hawm ntawm VLP, thiab kev tshem tawm tus feem pua ntawm cov heteroprotein tuaj yeem ncav cuag 99%. Ntawm lawv, hollow fiber membrane faltration technology muaj qhov zoo ntawm kev quab yuam me thiab qis, yog li nws raug pom zoo kom xaiv hollow fiber ntau rau cov kev xav thiab huv vlp.
Thaum ultrafiltration txoj kev yog siv rau concentration thiab purification, nws yog qhov tseem ceeb uas yuav tsum xaiv lub ntsej muag zoo, uas txiav txim siab ua tau zoo thiab zoo ntawm lub siab. Ntawm ib sab tes, nws yog ib qho tsim nyog los xaiv lub hom phiaj lwg me ntsis, thiab ntawm qhov kev tshem tawm ntawm cov herigroproteins yuav tsum tau txiav txim siab. Yog li ntawd, qhov zoo tshaj plaws yog los xaiv cov membrane nrog cov loj tshaj plaws loj tshaj plaws uas muaj peev xwm xaiv lub hom lim uas muaj cov uniform pore loj muab faib.
Thawj huv huv:
1. Cov Qauv Nrig Zej Zog
Polyethylene glycol (peg) nag lossis daus: Kev xaiv cov nag lossis cov nag lossis cov nag lossis daus
2. Chromatography
Affinity chromatography:
Heparin affinity: Siv VLPS thaj chaw ntawm kev them nqi ncaj ncees (piv txwv li, HPV VLP).
Tshuaj tiv thaiv cov tshuaj tiv thaiv cov chromatographaphy: qhov tshwj xeeb tab sis kim.
Ion txauv cov chromatography (ex): Xaiv anion txauv (piv txwv li, Q/ PDP) nyob ntawm VLP cov yam ntxwv sib txawv.
Hydrophobic sib cuam tshuam chrataphyraphy (hic): raws li VLPS saum hilrophobicity, haum rau qee qhov encapsulated vlps.
Zoo nkauj huv:
Molecular cais cov ntawv chromatogromaphy (sec, gel pom):
Kev tshem tawm ntawm cov khoom seem uas tseem tshuav ntawm cov neeg muaj protein, nucleic acids lossis agregates, nrog kev hloov pauv tsis sib haum. Cov kev daws teeb meem siab tab sis flux tsawg, feem ntau siv ua kauj ruam kawg.
Ntau hom chromatography:
Piv txwv li, Capto tub ntxhais series Resins, ua ke nrog ion txauv thiab molecular sab cib nyhuv, muaj peev xwm tshem tawm ntau yam impurities hauv ib kauj ruam.
Tus kab mob inactivation / nucleic acid tshem tawm (yog tias yuav tsum tau):
NucLease Kev Kho Mob: xws li Benzonase Demige Style DNA / RNA.
UF / DF: Kev sib koom tes Sff system kom tshem tawm cov me me ntawm nucleic acids thiab enzymes.
Concentration thiab npaj:
Tangential khiav pom kev (TFF):
Mloog rau cov hom phiaj rau cov hom phiaj thaum lub sijhawm tsis zoo (xws li PBS lossis kev tsim qauv tsis sib xws).
:
{{0 {0}}. 22Hμm MiBBrane Filtration ua kom muaj menyuam.
Cov ntaub ntawv raug
Cov hauv qab no yog cov txheej txheem ua kom huv rau cov tshuaj tiv thaiv vlp.
Hais Txog Guidling
Cov thev naus laus zis yog tsim kev tsim qauv thiab cov lag luam qib siab tsom mus rau hauv cov kab mob hauv qab, sib cais thiab purification ntawm biopharracuticals. Cov khoom siv dav hauv txheej txheem ntawm Mab, tshuaj tiv thaiv, kuaj ntshav, ntshav, ntshav qab zib, endoxin thiab lwm yam khoom lag luam; Cov tshuab lim dej uas muaj "Cassettes Lim thiab cov khoom lag luam lim dej ntau" thiab cov khoom siv sterrifugal "," Cov thev naus laus zis yog saib rau pem hauv ntej kom sib koom tes nrog koj!