Muaj -Scale Downstream Purification Process For Bovine Infectious Rhinotracheitis (IBR) Vaccine — "Ultrafiltration" Section
Bovine infectious rhinotracheitis (IBR) yog tshwm sim los ntawm kev kis tus kab mob bovine infectious rhinotracheitis virus (IBRV), tseem hu ua bovine herpesvirus type 1 (BHV-1). Tus kab mob no feem ntau tshwm sim los ntawm cov tsos mob ua pa thiab rho menyuam. Ntxiv nrog rau cov kev kuaj mob no, IBR tuaj yeem ua rau txo qis cov kua mis tawm hauv cov nyuj thiab txo qhov hnyav nce hauv nyuj nyuj, ua rau muaj kev lag luam loj rau cov tsiaj txhu ua liaj ua teb.
Tus kab mob yog immunosuppressive nyob rau hauv cov xwm. Thaum tshwm sim raws li ib tug kab mob, nws pathogenicity kuj tsawg; Txawm li cas los xij, thaum sib xyaw ua ke nrog lwm cov kab mob kis los yog kab mob tshwm sim, qhov hnyav thiab kev puas tsuaj nce ntxiv. Kev txhaj tshuaj tiv thaiv yog txoj hauv kev zoo tshaj plaws rau kev tiv thaiv thiab kev tswj hwm, muaj ob hom tshuaj tiv thaiv tseem ceeb: cov tshuaj tiv thaiv nyob qis thiab cov tshuaj tiv thaiv tsis muaj zog. Tam sim no, cov tshuaj tiv thaiv kab mob rhinotracheitis bovine siv rau hauv cov liaj teb feem ntau yog cov tshuaj tiv thaiv tsis muaj zog.
Cov tshuaj tiv thaiv kab mob muaj sia nyob muaj qhov muaj zog tiv thaiv kab mob, pib tiv thaiv kab mob sai, thiab kev tiv thaiv ntev ntev (feem ntau ntau dua rau lub hlis). Lawv feem ntau siv rau kev txhaj tshuaj tiv thaiv xwm txheej ceev thaum muaj kab mob. Txawm li cas los xij, lawv muaj peev xwm ua rau muaj kev pheej hmoo ntawm tus kab mob tso tawm, ua rau muaj kev pheej hmoo rau cov nyuj cev xeeb tub, thiab tsis tuaj yeem siv rau hauv cov kab mob latently tab sis cov nyuj asymptomatic.
Cov tshuaj tiv thaiv tsis muaj zog yog tus cwj pwm los ntawm kev nyab xeeb siab, tsis muaj kev pheej hmoo ntawm tus kab mob los yog rov qab mus rau kev ua phem, thiab suav tias muaj kev nyab xeeb kiag li. Lawv tuaj yeem siv rau hauv cov nyuj ntawm txhua theem, suav nrog nyuj cev xeeb tub, nyuj, thiab yug menyuam nyuj. Txawm li cas los xij, qhov pib ntawm kev tiv thaiv yog qhov qeeb thiab lub sijhawm tiv thaiv luv dua, yog li kev txhaj tshuaj tiv thaiv kab mob feem ntau yuav tsum tau ua. Qee qhov xwm txheej, kev tiv thaiv kev ua tau zoo yuav tsis muaj zog dua li cov tshuaj tiv thaiv muaj sia nyob.
Txawm hais tias siv cov tshuaj tiv thaiv kab mob los yog cov tshuaj tiv thaiv tsis muaj zog, cov txheej txheem purification hauv qab tuaj yeem muab faib ua plaub theem: sau thiab qhia meej → concentration thiab thawj purification → polishing purification → inactivation / sterile filtration thiab formulation.
Kev sib xyaw ua ke yog ib kauj ruam tseem ceeb hauv cov txheej txheem ua kom huv hauv qab ntawm cov tshuaj tiv thaiv, tam sim ntawd tom qab kev qhia meej. Nws lub hom phiaj tseem ceeb yog kom txo tau cov qoob loo loj heev - ntim, qis - cov kab mob sib xyaw kom pom tseeb cov tshuaj rau ib qho me me - ntim, siab- daim ntawv concentration thaum tswj cov kab mob bioactivity. Qhov no tsim cov xwm txheej tsim nyog rau qib siab tom ntej - kev daws teeb meem tab sis qis - muaj peev xwm ua kom huv huv cov kauj ruam, xws li chromatography.
Cov kauj ruam no feem ntau yog ua los ntawm kev siv tangential flow ultrafiltration (TFF). Lub hauv paus ntsiab lus yog raws li nram no: cov tshuaj tua kab mob kis mus rau qhov chaw ntawm ib qho ultrafiltration membrane nrog ib qho pore loj. Nyob rau hauv lub siab, me me molecules xws li dej, ntsev, thiab tej yam impurities kis perpendicularly los ntawm daim nyias nyias thiab raug tshem tawm, thaum cov kab mob tsis zoo, uas loj npaum li cas ntawm daim nyias nyias pores, yog khaws cia, tsis tu ncua recirculated, thiab concentrated. Piv nrog rau cov tsoos siab -ceev centrifugation, txoj kev no yog softer rau cov kab mob tsis yooj yim xws li IBRV, uas muaj lub hnab ntawv lipid. Nws ua tau zoo txo tus kab mob cov qauv kev puas tsuaj thiab kev ua haujlwm poob los ntawm kev ua haujlwm siab shear thiab muaj peev xwm ua tau zoo dua rau cov nplais linear - rau kev tsim khoom lag luam.
Kev ua tiav kev ua haujlwm concentration yog ntau tshaj qhov tsuas yog txo qhov ntim. Cov ntsiab lus tseem ceeb rau kev txhim kho cov txheej txheem muaj xws li: tswj cov kev hloov pauv ntawm lub siab thiab cov khoom noj txaus kom sib npaug pom kev ua haujlwm thaum txo qis concentration polarization thiab membrane fouling; xaiv cov khoom siv membrane uas tsim nyog thiab qhov pore loj kom ntseeg tau tias muaj tus kab mob zoo khaws cia thiab permeate flux; thiab nrhiav qhov zoo tshaj qhov sib npaug ntawm cov kab mob rov qab, qhov tseem ceeb concentration, thiab lub sijhawm ua haujlwm. Cov tshuaj tiv thaiv kab mob siab tsis yog tsuas yog ua tiav qhov titer siab dua xwb tab sis kuj tseem ua tiav qhov kev ua kom huv ua ntej los ntawm kev tshem tawm ib feem ntawm cov dej ntau -soluble impurities. Qhov no muab qhov tsim nyog ntim thiab concentration lub hauv paus rau cov kauj ruam tseem ceeb tom ntej, xws li chromatography thiab nuclease kev kho mob, ua kom concentration ib lub hauv paus efficiency hub nyob rau hauv tag nrho cov downstream txheej txheem.
Secondary diafiltration yog ib qho tseem ceeb hauv cov tshuaj tiv thaiv downstream purification, positioned tom qab zoo purification thiab ua ntej formulation. Nws feem ntau yog ua tom qab chromatography thiab kho nuclease. Nws lub hom phiaj tseem ceeb tsis yog thawj zaug concentration, tab sis kev sib pauv hloov pauv thiab kev hloov pauv ntawm qhov kawg ntawm cov txheej txheem. Cov txheej txheem yog ua nyob rau hauv ib tug tangential flow ultrafiltration (TFF) system, qhov twg tshiab, huv si formulation tsis tu ncua ntxiv mus rau lub circulating concentrated viral tov, thaum lub qub hnyav thiab me me - molecule impurities raug tshem tawm. Qhov kev ua haujlwm no tau txais txiaj ntsig zoo thiab maj mam tshem tawm cov ntsev seem, cov kuab tshuaj organic, cov khoom lag luam nuclease degradation, thiab cov kua dej tsis huv uas tseem tshuav los ntawm cov txheej txheem purification.
Qhov tseem ceeb yog kom muaj qhov ntim tsis tu ncua lossis siv cov kev hloov kho me me kom ntseeg tau tias tus kab mob concentration ua tau raws li cov qauv kev qhia tshwj xeeb. Rau cov kab mob tsis yooj yim, cov kab mob hauv cov kab mob xws li kab mob rhinotracheitis bovine infectious rhinotracheitis virus (IBRV), ib puag ncig zoo ntawm hydrodynamic ib puag ncig ntawm cov diafiltration theem nrab yog qhov tseem ceeb rau kev khaws cov khoom zoo thiab kev tiv thaiv kab mob. Thaum kawg, cov kauj ruam no muab lub hauv paus ruaj khov rau kev tsis ua haujlwm tom ntej (yog tias xav tau), kev sib txuas ntxiv lossis cov khoom ruaj khov ntxiv, thiab kev ua tiav zaum kawg, kom ntseeg tau tias cov khoom kawg nkag mus rau hauv kev tsim nrog cov ntsiab lus, cov xwm txheej zoo sib xws, thiab kev sib raug zoo. Yog li ntawd nws yog ib qho ntawm cov kauj ruam tseem ceeb rau kev ua kom cov tshuaj tiv thaiv kev nyab xeeb, ruaj khov, thiab batch-rau-batch sib xws.
IBRV yog ib lub hnab ntawv, ob npaug -kab tawm kab DNA kab mob nrog ib lub hnab ntawv kwv yees li spherical. Mature IBRV hais muaj ib txoj kab uas hla ntawm 160-230 nm. Raws li, siv 100, 300, lossis 500 kDa ultrafiltration membranes tuaj yeem khaws IBRV thaum tshem tawm qee cov kab mob sib kis. Lub ultrafiltration rov qab tus nqi ntawm Jiuling Technology membrane cassettes sib txawv nrog cov khoom noj, tab sis feem ntau mus txog 90-95%.