Q: Kuv yuav tsum xaiv microsomes thiab Hepatocytes rau metabolic Stability Choj? Kuv puas tuaj yeem xaiv ib tus ntawm lawv kom ua tiav qhov yuav tsum tau ua?
A: Hauv metabolic kev kawm ruaj khov, kev xaiv ntawm daim tawv nqaij microsomes lossis hepatocytes feem ntau yog qhov sib xyaw, nyob rau ib qho nrog tus ntsuas metabolic siab yog xaiv. Feem ntau, daim siab micreem tau, tshwj xeeb tshaj yog thaum lub compound molecules yog ntau cov roj av ntxiv - Hepatocytes tuaj yeem siv rau kev sim thaum muaj pov thawj ntawm ob qho kev sib tw - Feem ntau, ib lub kaw lus metabolic tuaj yeem xaiv tau kom tau raws li qhov kev xav tau; Yog tias cov xwm txheej hauv txhua qhov kev tso cai, nws yog qhov zoo tshaj plaws los xaiv ob qho tib si hauv tib lub sijhawm.
Q: Vim li cas nws thiaj tsim nyog siv thawj tus hepatocytes rau enzyme inductionay?
A: CYP Ezyme - Txhaum kev ua yeeb yam yuav tsum tau mus ntawm "kev sau ntawv" rau "post - Kev txhais cov protein" kom tau txais cov hnoos qeev nquag "kom tau txais cov protein acein. Yog li, qhov kev xeem ntawv yuav tsum yog lub siab mob hlwb, tsis yog lier microsomes lossis mob siab s9.
Q: Vim li cas kuv thiaj yuav tsum xaiv peb tus neeg pub nyiaj tib neeg thawj tus Hepatocytes rau Enzyme Induction Induction?
A: Raws li kev qhia rau cov lus qhia rau cov kev sib cuam tshuam tshuaj yeeb, tsawg kawg peb tus neeg pub nyiaj yuav tsum tau ntsuas cais. Ntxiv rau cov tsim cov ntsiab lus tseem ceeb, kev xaiv ntawm peb tus neeg pub nyiaj rau kev sim kuj tseem ceeb dua rau kev ntsuas tus inter - cov kev sib txawv. Yog tias qhov tshwm sim los ntawm tsawg kawg ib tus neeg pub nyiaj ntau tshaj ib tus neeg ua haujlwm tau ua tiav, cov neeg sib tw txiav yeeb tuaj yeem cuam tshuam thiab ua raws li kev ntsuam xyuas - yuav tsum muaj.
Q: Vim li cas thiaj tsom tau ntawm CYP enzyme induction thiab tsis on ugt enzyme induction?
A: Qhov laj thawj rau kev tsom mus rau CYIE Induction Induction yog tias kev siv tsheb ntawm CYPEST induction tau kawm kom meej meej. Hauv lwm lo lus, vim li cas rau tsis xav tau txoj kev kawm ntawm Ugtase Induction yog tias kev siv tshuab tseem tsis meej; Txawm li cas los xij, qhov no tsis txhais tau tias auvtase yuav tsis yog ntxias. Cov lus qhia tseem muaj lub xeev uas tsis muaj cov qauv qhia qauv rau cov cim lossis cov neeg thauj neeg thauj khoom thiab theem II metabolism enzymes.
Q: Ntev npaum li cas tuaj yeem ua tiav thawj tus hepatocytes muaj sia nyob tom qab kev rov ua xyem xyav, thiab ntev npaum li cas tuaj yeem tswj hwm yam tsis muaj kev rov ua haujlwm ntxiv tom qab resuscitation?
A: Pasteurized thawj Hepatocytes feem ntau siv rau enzyme induction ua. Tom qab kev rov ua haujlwm ntawm tes, cov hlwb raug tshem tawm hauv phaj kis nruab nrab, haum kom tsim nyog, thiab kev coj noj coj ua hauv collagen - coated daim hlau; Tom qab ntawd cov cell tuaj yeem muab tshuaj txhuam hauv 4 ~ 6 teev. Tom qab lub hlwb tau txuas nrog phab ntsa, nruab nrab ntawm nruab nrab yog hloov thiab tswj hwm rau 18 h kom ua kom muaj kev kho dua tshiab ntawm lub xeev. Tom ntej no, enzyme induction inductionay tuaj yeem ua los xyuas cov metabolic kev ua ub no thiab mrna induction qib. Los ntawm cov kev pom ntawm lub voj voog tag nrho, Hepatocytes tuaj yeem tswj hwm rau 6 ~ 7 hnub tom qab phab ntsa ua vaj tawv. Raws li lub sijhawm mus los ntawm, lub phab ntsa phab ntsa uas muaj kev cuam tshuam ntawm phab ntsa Cell
Yog tias cov lus tsis zoo yuav tsis muaj kev coj noj coj ua, peb tau txheeb xyuas tias lawv tuaj yeem tswj hwm hauv lub xeev zoo tsis pub dhau 4 ~ 6 teev. Peb tseem tsis tau ua tiav cov ntaub ntawv pov thawj rau lub sijhawm ntev dua.
Q: Hepatocytes tuaj yeem siv tau ob qho kev ncua thiab cov hlwb uas raug ncua nyob ntawm cov kev sib tham sib txawv media siv?
A: Feem ntau cov hlwb los ntawm cov ntaub so ntswg thiab cov plab hnyuv siab raum yog walled, tab sis tsis yog txhua tus hlwb muaj ntsa hauv - vitro. Seb cov hlwb yog phab ntsa - ua rau muaj nyob ntawm lub xeev ntawm cov hlwb lawv tus kheej; Nyob rau tib lub sijhawm, phab ntsa - Cov qog hlwb yuav tsum muaj kev coj noj coj ua me me (E.hm.hm.heej, fibronectin, fibronection qhov tseeb) uas tuaj yeem koom nrog hauv cov txheej txheem ntawm tes txuas. Hauv kev xaus, cov txuj ci cell tuaj yeem siv ua cov hlwb ncua, tab sis cov hlwb tshem tawm tsis tas muaj rau kev siv tus kheej.
Q: Dab tsi yog qhov zoo / tsis zoo ntawm kev siv daim ntawv qhia phab ntsa Phab Ntsa raug tshem tawm kev ncua kev ncua hepatocytes? Dab tsi yog qhov kev txiav txim siab?
A: Tom qab Hepatocytes yog cais, lawv tuaj yeem yog kev coj noj coj ua lossis hauv phab ntsa nplaum. Qhov kev ua si ntawm Cytochrome P450 enzyme los ntawm kev ncua kev kawm hauv tuam txhab Hepatocytes muaj feem ntau nrog rau thawj 4 ~ 6H, txo qis nrawm nrog lub sijhawm ntev. Yog li, ncua kev kawm hepatocytes feem ntau siv rau kev kawm metabolic kev kawm lossis metabolite profiling kev kawm. Lub Hepatocytes hauv phab ntsa nplaum muaj sijhawm txaus kom muaj kev puas tsuaj los ntawm kev puas tsuaj los tswj cov yam ntxwv ntawm kev puas tsuaj thiab kev ua haujlwm metabolic ntawm ib txwm hepatocytes. Li no, lawv feem ntau siv rau cov kev tshawb fawb enzyme induction induction, metabotoxicity, metabolic ruaj khov ntawm cov tshuaj kawm qeeb metabolizing.
Tam sim no, peb tsis muab lub vev xaib online yuav khoom. Cov khoom ntxiv rau lub tawb nqa khoom tom qab sau npe thiab nkag mus yog rau kev siv xwb. Yog tias koj xav yuav peb cov khoom, thov cia koj cov ntaub ntawv tiv toj los ntawm kev sib cuag peb channel, thiab peb yuav tiv tauj koj nyob rau lub sijhawm los ua kom tiav peb cov kev pabcuam.
Q: Yuav ua li cas tsis yog - cov protein sib xyaw cuam tshuam rau kev sim ua?
A: Yog tias cov tshuaj sib tw khi cov ntaubns tsis - tshwj xeeb rau microsomal protein, qhov kev tshwm sim hauv kev hloov kho cov kab ke. Raws li cov protein ntau nce, lub txiaj ntsig km tus nqi nce ntxiv, ua rau muaj kev txiav tawm qis qis. Tsis tas li, cov tshuaj khi ntawm cov tshuaj uas muaj protein nyob rau hauv microsomes tuaj yeem ua rau muaj kev txawv txav loj hauv cov chaw ua hauj lwm txawv thiab sib txawv.
Q: Qhov pom zoo ntawm lub siab tawv ntawm cov khoom siv theem kuv lossis theem II met IIt II hauv kev siv zog rau lawv tau zoo ua ntej ntxiv rau lub system?
A: Tsis tas yuav dilute the hepatic micromomes ua ntej; Cov concentration ntawm microsomatic nyob rau hauv cov khoom siv yog 20 mg / ml thiab qhov kawg concitor hauv kev sim ntsuas yog 0.1 mg / ml, uas tuaj yeem muab ntxiv rau kev sib npaug.
Q: Dab tsi yog cov pom zoo ntawm cell ntom rau metabolic stability testing ntawm thawj hepatocytes? Puas yog qhov kev tshem tawm cov kev txiav txim siab tib yam li kev ntsuas microsomes?
A: Cov kev pom zoo ntawm cell density rau thawj Hepatocyte Metabolic Stability Assay yog 0.5 txog 2 × 106 Cov Hlwb / ml, thiab cov kev hloov pauv thiab kev ua cov txiaj ntsig ntawm cov hepatic microsomal metabolic stability assentay yeej xwm yeem.
Q: Cov khoom xyaw tseem ceeb hauv koj lub PBS tsis yog dab tsi? Puas yog nws muaj kcl thiab Nacl?
A: Lub ntsiab Cheebtsam ntawm peb PBS tsis yog k2Hpo4 thiab kh2PO4, thiab yog KCl thiab Nacl.
Q: Lub hom phiaj ntawm phosphate buffers yog dab tsi? Vim li cas koj thiaj li xav tau phosphate?
A: Phosphate Buffers yog xaiv rau lawv lub hom phiaj ntawm kev ua pa ntawm lub cev ib puag ncig, thiab phosphate yog ib qho tseem ceeb tshaj plaws tsis muaj ib puag ncig lub cev.
Q: Qhov niaj zaus teeb tsa rau enzyme - Indusen Receptor siab? Puas muaj kev daws teeb meem yog tias cov solubility ntawm lub kaw lus yuav tsum tau kuaj yog neeg pluag?
A: Kev xav tau enzyme tsis xav tau teeb tsa nrog qib kev xav tau zoo dua li ntawm kev txiav txim siab ntawm tib neeg ntau dua li kev txiav txim siab ntshav siab ntau dua li cov tshuaj tiv thaiv ntshav ntau zaus hauv tib neeg. Yog hais tias solubility nyob rau theem aqueous yog neeg pluag, ib lub cev muaj zog muaj peev xwm xaiv tau raws li nws cov kuab tshuaj, piv txwv li. DMOs, tab sis tus nqi ntawm cov organic kuab tshuaj ntxiv rau qhov system yuav tsum tau tswj hwm.
Q: Kev tshawb xyuas hauv metabolic stability kev kawm, yog qhov yuav tsum siv ob lub tshuab kuaj, lub siab micratocytes thiab Hepatocytes, rau kev sim?
A: Hepatic microsomes yog ze li spherical memickas - Thawj Hepatocytes (Phcs) yog Hepatocytes Ceg tsis yog tom qab kev sib cais ntawm cov tsiaj hauv lub siab, tshwj xeeb tshaj yog cov khoom lag luam metapolic, tshwj xeeb tshaj yog cov khoom lag luam metapolic, tshwj xeeb tshaj yog cov kev khaws cia ntawm cov khoom siv hauv cov tsiaj txhu. Hauv qhov chaw kawm ruaj khov, tsis tas yuav xav txog tus nqi, thiab ob lub tshuab kuaj khoom tuaj yeem xaiv tau rau qhov kev xeem tib lub sijhawm; Lossis cov kev ntsuas tsim nyog tuaj yeem raug xaiv raws li cov khoom lag luam metabolic ntawm cov compound, thiab lub hauv paus ntsiab lus yog tias cov kab ke twg muaj qhov siab metabolic tus nqi yog xaiv. Nyob rau hauv dav dav, daim siab microsomes yog qhov kev xaiv zoo tshaj plaws thaum lub compound molecules yog ntau cov metabolism yog qhov tseem ceeb metabolism yog qhov tseem ceeb metabolism (tshwj xeeb ntawm CYP); Thaum ob - theem metabolism yog qhov tseem ceeb ntawm kev siv hluav taws xob tseem ceeb, tsis pom zoo nyob hauv daim ntawv ntsuam xyuas microscomes, thiab cov tshuaj ntsuam tuaj yeem nqa tawm siv cov hepatocytes.
Q: Yog lub concentration ntawm microsomes tshuaj ntsuam hauv Metabolic Strability Test? Dab tsi yog cov txiaj ntsig ntawm kev siab dhau lossis qis dhau?
A: Hauv kev ntsuas kev ruaj khov, cov protein ntau yam tseem ceeb ntawm 0.1 mg Siab dhau kev xav ntawm microsis cov protein yuav tsis - tsis yog los ntawm cov tshuaj ntawm cov tshuaj mus rau microsinomal protein; Thaum lub sijhawm qis dua ntawm microsomein cov protein yuav ua rau tsis muaj cov tshuaj metabolism tsis txaus ntseeg ntawm cov tshuaj.