Qhov sib txawv ntawm monoclonal thiab polyclonal antibodies yog dab tsi?

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• Monoclonal antibodies yog dab tsi?
  • 1. Kev Txhais Tau
  • 2. Keeb Kwm
  • 3. Monoclonal Antibody Ntau Lawm
  • 4. Txoj kev Cloning
  • 5. Daim ntawv thov
• Dab tsi yog polyclonal antibody?
  • 1. Polyclonal antibody txhais li cas?
  • 2. Qauv
  • 3. Kev cais tawm
  • 4. Daim ntawv thov
• Monoclonal vs Polyclonal antibodies
  • 1. Lub Tswv Yim
  • 2. Ntau lawm
  • 3. Cons thiab Pros
  • 4. Lwm qhov sib txawv
• xaus

Monoclonal antibodies yog dab tsi?

1. Lus txhais

Muaj qhov sib txawv loj ntawm monoclonal thiab polyclonal antibodies. Cov kab mob monoclonal yog cov khaub ncaws ntau tshuaj tiv thaiv uas tau tsim los ntawm ib leeg B cell clone thiab lub hom phiaj tsuas yog qhov tshwj xeeb epitope. Hybridoma antibody thev naus laus zis yog tsim los ntawm kev siv thev naus laus zis ntawm tes, uas suav nrog kev nkag siab zoo B cells thiab myeloma cells nrog lub peev xwm ua kom tsis txawj tuag mus rau hauv B cell hybrids qog.

Ib leeg hybridoma ntawm tes nrog cov yam ntxwv no tau siv los cog cov pej xeem ntawm tes los npaj cov tshuaj tiv thaiv tshwj xeeb tiv thaiv epitope, uas yog, a monoclonal tshuaj tiv thaiv.

2. Keeb kwm

Kev tsim kho ntawm cov kab mob monoclonal tau hla plaub theem, uas yog:
Murine monoclonal tshuaj tiv thaiv, chimeric antibody, humanized monoclonal antibody thiab tag nrho tib neeg cov tshuaj tiv thaiv kab mob monoclonal.

Siab tib neeg cov kab mob monoclonal: Cheeb tsam sib txawv thiab thaj tsam tas li ntawm cov tshuaj tiv thaiv kab mob yog tib neeg keeb kwm, tshem tawm cov tshuaj tiv thaiv kab mob thiab cov tshuaj lom.

Cov thev naus laus zis cuam tshuam rau kev npaj ua tiav tib neeg cov tshuaj tiv thaiv feem ntau suav nrog:
Tib neeg siv tshuab thev naus laus zis, EBV hloov pauv B lymphocyte thev naus laus zis, phage zaub thev naus laus zis, transgenic nas tshuaj tiv thaiv kab mob npaj thev naus laus zis thiab tib leeg B cell tshuaj tiv thaiv kab mob.

Tib neeg thiab tag nrho humanized tshuaj tiv thaiv tshuaj npaj los ntawm tib neeg thiab muaj cov tshuaj tiv thaiv tib neeg muaj peev xwm kov yeej ntau yam tsis txaus ntawm cov tshuaj tiv thaiv tsiaj thiab cov tshuaj tiv thaiv chimeric vim tias lawv muaj kev sib raug zoo, tshwj xeeb, thiab muaj tshuaj lom tsawg thiab muaj kev phiv. Nws tau dhau los ua qhov tsis sib xws hauv kev txhim kho tshuaj kho tshuaj tiv thaiv kab mob.

3. Monoclonal Antibody Ntau Lawm

1. Txhaj tshuaj rau tsiaj
   Kev txhaj tshuaj tiv thaiv tsiaj yog txheej txheem tiv thaiv cov nas nrog lub hom phiaj antigen kom cov nas tsim cov tshuaj tiv thaiv kab mob B lymphocytes. Feem ntau, poj niam BALB/c nas hnub nyoog 6-8 lub lis piam raug xaiv thiab txhaj tshuaj raws li txoj cai tau txhaj tshuaj tiv thaiv kab mob ua ntej. Antigen nkag mus rau cov kab mob hauv lub cev tiv thaiv kab mob los ntawm cov ntshav ncig lossis cov qog ntshav ncig, txhawb qhov sib thooj B lymphocyte clones, ua kom muaj zog, nthuav tawm, thiab sib txawv rau hauv kev nkag siab B lymphocytes.
   
2. Cell Fusion
   Cov nas raug tua nrog cov pa roj carbon dioxide, thiab cov hnoos qeev raug tshem tawm aseptically. Tom qab ntawd nyem thiab tsoo nws hauv cov tais zaub mov kom ua rau lub cev raug tshem tawm. Cov npaj syngeneic myeloma cells thiab nas spleen cells tau sib xyaw hauv qee qhov kev faib ua feem, thiab ntxiv cov fusion txhawb nqa polyethylene glycol. Raws li lub luag haujlwm ntawm polyethylene glycol, ntau yam lymphocytes tuaj yeem sib xyaw nrog cov cell myeloma los tsim cov cell hybridoma.
   
3. Xaiv cog qoob loo
   Lub hom phiaj ntawm kev xaiv kab lis kev cai yog tshuaj xyuas fused hybridoma cells, thiab HAT xaiv nruab nrab feem ntau siv. Hauv HAT nruab nrab, cov tsis siv myeloma hlwb tuag vim tsis muaj hypoxanthine-guanine-phosphoribosyltransferase thiab tsis muaj txoj hauv kev khaws cia los ua ke DNA.
   
   Txawm hais tias tsis siv cov lymphocytes muaj hypoxanthine-guanine-phosphoribosyltransferase, lawv tsis tuaj yeem muaj sia nyob ntev hauv vitro.
   
   Tsuas yog cov fused hybridoma cells tuaj yeem muaj sia nyob thiab nthuav dav hauv HAT nruab nrab vim tias lawv tau txais hypoxanthine-guanine-phosphoribosyltransferase los ntawm cov qog hlwb thiab lub peev xwm ntawm kev loj hlob tsis tu ncua los ntawm myeloma cells.
   
4. Kuaj thiab cloning ntawm hybridoma zoo clones
   Tsuas yog qee qhov ntawm cov menyuam sib xyaw hybridoma loj hlob hauv HAT nruab nrab yog cov cell uas tso tawm qhov kev txiav txim siab tshwj xeeb monoclonal antibodies. Yog li, kev tshuaj xyuas thiab cloning yog qhov tsim nyog. Txoj kev lim dej limiting feem ntau yog siv rau kev coj noj coj ua ntawm cov cell hybridoma. Siv cov txheej txheem tiv thaiv kab mob rhiab, nrawm, thiab tshwj xeeb, cov kab mob hybridoma zoo uas tuaj yeem tsim cov tshuaj tiv thaiv kab mob monoclonal uas xav tau raug tshuaj xyuas thiab cloned thiab nthuav dav. Tom qab kev txheeb xyuas qhov tseeb ntawm hom immunoglobulin, subclass, qhov tshwj xeeb, kev nyiam, epitope ntawm kev lees paub antigen thiab nws qhov hnyav ntawm cov tshuaj tiv thaiv monoclonal zais cia los ntawm nws, nws yuav tsum tau khov hauv lub sijhawm.
   
5. Kev tsim ntau ntawm cov tshuaj tiv thaiv monoclonal
   Kev npaj loj ntawm cov kab mob monoclonal feem ntau lees txais tsiaj hauv vivo induction thiab hauv vitro kev coj noj coj ua.

4. Txoj kev Cloning

Cov qhov dej zoo uas tau sim rau tshuaj tiv thaiv kab mob tuaj yeem tsim thiab cloned kom tau txais cov tshuaj tiv thaiv kab mob monoclonal zais los ntawm cov xeeb ntxwv ntawm ib lub cell. Feem ntau hais lus, ua ntej lub sijhawm cloning, zoo dua. Vim tias nyob rau lub sijhawm no, ntau yam hybridoma cells loj hlob zoo nyob rau tib lub sijhawm, sib tw rau kev noj zaub mov thiab chaw, thiab cov cell tsim cov tshuaj tiv thaiv tshwj xeeb tuaj yeem ua rau poob qis thiab tshem tawm.

Txawm li cas los xij, lub sijhawm cloning yuav tsum tsis txhob ntxov dhau, txwv tsis pub cov yam ntxwv ntawm tes tsis ruaj khov, thiab tus lej ntawm cov cell me me thiab poob yooj yim. Lub cloned positive hybridoma cells, tom qab lub sijhawm kev coj noj coj ua, tseem yuav plam lub peev xwm los tsim cov tshuaj tiv thaiv vim yog kev hloov pauv ntawm tes lossis poob ntawm cov chromosomes tshwj xeeb, yog li lawv yuav tsum tau cloned ntau thiab ntau dua. Tus naj npawb ntawm cloning yog txiav txim siab los ntawm lub peev xwm tso pa tawm thiab tiv thaiv kab mob ntawm antigens. Feem ntau hais lus, tus naj npawb ntawm clones ntawm lub zog tiv thaiv kab mob antigen tuaj yeem tsawg dua, tab sis tsawg kawg 3 txog 5 clones tuaj yeem ua rau nws ruaj khov. Muaj ntau txoj hauv kev ntawm cloning, suav nrog txwv txoj kev dilution, micromanipulation txoj kev, cov txheej txheem mos agar phaj thiab txoj kev ua kom sib cais ua kom pom kev zoo.

5. Daim ntawv thov

Txij li thaum lub advent ntawm cov kab mob monoclonal, lawv tau siv rau hauv ntau thaj chaw ntawm cov tshuaj vim lawv cov yam ntxwv tshwj xeeb.

1. Kuaj tshuaj kuaj kab mob reagents
   Raws li kev kuaj mob reagent hauv cov chaw kuaj tshuaj kho mob, cov kab mob monoclonal tau siv dav hauv thev naus laus zis xws li kev sib txuas enzyme-txuas immunosorbent assay, radioimmunoassay, txhaj tshuaj tiv thaiv kab mob thiab ntws cytometry vim lawv qhov tshwj xeeb muaj zog, siab dawb huv, thiab zoo ib yam. Thiab daim ntawv thov ntawm cov kab mob monoclonal tau txhawb nqa txoj kev loj hlob ntawm cov khoom siv coj mus muag.
   
   Cov khoom siv ua lag luam ua los ntawm cov tshuaj tiv thaiv kab mob monoclonal tau siv dav hauv:
   1. Tshawb nrhiav cov kab mob pathogenic antigens thiab antibodies;
   2. Tshawb nrhiav cov qog nqaij hlav antigens;
   3. Tshawb nrhiav cov cell tiv thaiv kab mob thiab lawv cov pab pawg tsawg;
   4. Kev txiav txim siab cov tshuaj hormones;
   5. Kev txiav txim siab ntawm cytokines.
      
      Kev lees paub ntawm antigens los ntawm cov tshuaj tiv thaiv monoclonal yog qhov sib txawv heev ntawm cov tshuaj tiv thaiv polyclonal. Cov khoom siv sib txawv siv cov tshuaj tiv thaiv monoclonal sib txawv thiab muaj qhov sib txawv antigen paub qhov chaw, ua rau qee qhov sib txawv hauv cov txiaj ntsig ntsuas. Yog li, qhov teeb meem kev tsim qauv xav tau kev kawm ntxiv.
      
2. Purification ntawm cov protein
   Cov kab mob monoclonal yog ligands tseem ceeb hauv kev sib raug zoo chromatography. Cov tshuaj tiv thaiv kab mob monoclonal tau nthuav tawm ntawm cov txheej txheem tsis sib xws (xws li Spehrose 2B, 4B, 6B, thiab lwm yam) thiab npaj rau hauv kem chromatography. Thaum cov qauv ntws los ntawm kab ntawv chromatographic, cov antigen yuav raug cais tau tshwj xeeb tuaj yeem khi rau theem ruaj khov monoclonal antibody, thiab cov khoom seem tsis tuaj yeem khi rau nws. Tom qab cov kab lus chromatographic tau ua tiav tag nrho, lub zog ionic lossis pH ntawm eluate tau hloov pauv, cov antigen kom raug cais tawm yog cais tawm ntawm cov tshuaj tiv thaiv, thiab cov antigen kom huv tuaj yeem tau txais los ntawm kev sau cov eluate.
   
3. Kev kho mob qog nqaij hlav thiab siv tshuab thev naus laus zis
   Cov tshuaj tiv thaiv kab mob monoclonal tau hais tawm tsam qee qhov qog nqaij hlav antigen txuas nrog tshuaj siv tshuaj kho mob lossis tshuaj kho kab mob, thiab lub hom phiaj cuam tshuam ntawm cov tshuaj tiv thaiv kab mob monoclonal tau siv los nqa cov tshuaj lossis cov tshuaj kho hluav taws xob mus rau lub cev lub hom phiaj thiab ncaj qha tua lub hom phiaj ntawm tes, uas yog hu ua kev kho mob qog noj ntshav.
   
   Ib qho ntxiv, xov tooj cua tuaj yeem ua tau los ntawm kev txuas cov cim hluav taws xob nrog cov tshuaj tiv thaiv kab mob monoclonal thiab txhaj tshuaj rau hauv cov neeg mob txhawm rau pab kho mob qog noj ntshav. Monoclonal antibodies feem ntau yog tua cov tshuaj tiv thaiv kab mob, thiab cov ntshav ntawm cov tsiaj sib txawv tuaj yeem ua rau tsis haum rau tib neeg. Yog li ntawd, kev npaj ntawm tib neeg-tib neeg monoclonal antibodies lossis humanized tshuaj tiv thaiv yog qhov tseem ceeb dua, tab sis tsis muaj kev nce qib tseem ceeb hauv qhov no.

Dab tsi yog polyclonal antibody? 

1. Polyclonal antibody txhais li cas?

Antigens feem ntau yog tsim los ntawm ntau qhov kev txiav txim siab antigenic. Ib hom kev txiav txim siab antigenic txhawb lub cev, thiab tshuaj tiv thaiv tsim los ntawm B lymphocyte tau txais cov tshuaj tiv thaiv kab mob hu ua Monoclonal Antibody. Lub cev tau txhawb nqa los ntawm ntau qhov kev txiav txim siab antigenic, thiab ntau yam tshuaj tiv thaiv kab mob monoclonal tau tsim raws li. Cov tshuaj tiv thaiv kab mob monoclonal no tau sib xyaw ua ke los tsim polyclonal tshuaj tiv thaiv.

2. qauv

Cov qauv txheej txheem ntawm cov tshuaj antigen uas tuaj yeem ua rau lub cev tsim cov tshuaj tiv thaiv hu ua epitope. Muaj peev xwm muaj ntau qhov sib txawv antigenic txiav txim siab ntawm antigen, yog li lub cev tsim ntau yam tshuaj tiv thaiv sib txawv, thiab cov tshuaj tiv thaiv zaum kawg yog cov ntshav ntshav. Cov pej xeem ntawm cov ntshav ntshav uas tsuas yog ua rau ntawm ib qho tshuaj tiv thaiv kab mob antigenic yog clone. Cov tshuaj tiv thaiv tshwj xeeb uas tsim los ntawm clone hu ua monoclonal antibody. Cov kab mob monoclonal tuaj yeem tshwj xeeb khi rau ib qho tshwj xeeb antigenic tus txiav txim siab, ib yam li lub foob pob hluav taws raug tsoo lub hom phiaj. Ntawm qhov tod tes, txawm hais tias nws yog tib qho kev txiav txim siab antigenic, ntau qhov clones tuaj yeem tsim cov tshuaj tiv thaiv kab mob hauv lub cev, tsim kev sib xyaw ntawm ntau cov tshuaj tiv thaiv monoclonal, hu ua polyclonal tshuaj tiv thaiv.

3. Classification

Antigens feem ntau yog tsim los ntawm ntau yam epitopes. Ib hom epitope txhawb lub cev. Cov tshuaj tiv thaiv kab mob tsim los ntawm B lymphocyte tau txais lub antigen hu ua monoclonal antibody. Lub cev tau txhawb los ntawm ntau yam kev txiav txim siab antigenic, thiab ntau yam tshuaj tiv thaiv kab mob monoclonal tau tsim raws li. Cov tshuaj tiv thaiv kab mob monoclonal no tau sib xyaw ua ke los tsim cov tshuaj tiv thaiv kab mob polyclonal. Tsuas yog ntau qhov kev txiav txim siab antigenic, tib hom epitope tseem tuaj yeem txhawb lub cev los tsim tsib hom tshuaj tiv thaiv kab mob: IgG, IgM, IgA, IgE thiab IgD.

Polyclonal antibodies yog ib pawg immunoglobulins zais los ntawm lub cev cov ntshav ntshav uas txhawb los ntawm heterologous antigens (macromolecular antigens, hapten conjugates). Cov tshuaj tiv thaiv kab mob Polyclonal tau siv dav hauv kev tshawb fawb thiab kuaj mob vim tias lawv muaj peev xwm paub ntau yam epitopes thiab ua rau muaj dej nag tshwm sim nrog lub sijhawm npaj luv thiab tus nqi qis.

4. Daim ntawv thov

Ib lub polyclonal antiserum zoo muaj ntau yam tshuaj tiv thaiv tiv thaiv epitopes sib txawv ntawm qee yam antigen. Txij li polyclonal antiserum feem ntau muaj cov tshuaj tiv thaiv tiv thaiv epitopes sib txawv ntawm qee yam antigen, suav nrog cov tsis muaj peev xwm tiv taus epitopes, nws kuj tseem yuav muaj qhov cuam tshuam hauv qhov piv txwv tob. Hauv kev ua kom tawv nqaij ntawm cov ntaub so ntswg ua ntu ntu, Polyclonal cov tshuaj tiv thaiv kab mob feem ntau siv. Raws li qhov xav tau sib txawv ntawm qhov kev sim, polyclonal antibodies tau siv los sau cov kab mob sib xws.

Ib qho ntxiv, hauv kev ua liaj ua teb, polyclonal tshuaj tiv thaiv yog siv rau kev saib xyuas ntawm qhov chaw ntawm cov tshuaj tua kab; hauv kev siv tshuaj kho mob, tshuaj tiv thaiv kab mob polyclonal feem ntau yog siv rau kev kuaj kab mob, kuaj mob thiab kho, xws li cov tshuaj tiv thaiv kab mob tiv thaiv kab mob rau kev hloov pauv thiab kho cov kab mob autoimmune.

Monoclonal vs Polyclonal antibodies

1. tswvyim

Cloning: hais txog asexually reproduced cell line, uas yog pawg ntawm cov kab xov tooj ntawm tes tsim los ntawm kev faib thiab rov ua dua tshiab ntawm ib tus zuj zus ntawm tes. Hauv txhua tus tswv cuab ntawm tsev neeg no, yog tias tsis muaj kev hloov pauv, cov noob yog tib yam nkaus.

Polyclonal tshuaj tiv thaiv (pAb): Txhaj tshuaj tiv thaiv tsiaj nrog antigen uas muaj ntau yam epitopes, uas tuaj yeem txhawb ntau yam B cell clones los tsim cov tshuaj tiv thaiv sib txawv rau ntau yam epitopes. Cov tshuaj tiv thaiv kab mob uas tau txais yog qhov sib xyaw ntawm ntau yam tshuaj tiv thaiv.

Cov kab mob monoclonal (mAb): Cov tshuaj tiv thaiv kab mob homologous tsim los ntawm clone ntawm B cells uas paub txog epitope. Siab zoo ib yam, muaj zog tshwj xeeb, muaj lub npe siab, me me lossis tsis muaj kev cuam tshuam hla.

2. Qhuav

Qhov sib txawv ntawm kev npaj tshuaj tiv thaiv kab mob monoclonal:
B lymphocytes kho nrog cov antigens tshwj xeeb thiab myeloma cells tau fused kom tau txais cov cell hybridoma. Tom qab tshuaj xyuas los ntawm HAT nruab nrab thiab kuaj pom lub ntsiab lus los ntawm ELISA, tau txais cov clone zoo, thiab tom qab ntawd ua cov kab lis kev cai ntawm tes lossis txhaj cov cell mus rau hauv lub plab kab noj hniav ntawm cov tsiaj (feem ntau yog kab laug sab/c nas) thiab coj lawv nrog ascites, sau cov supernatant/ascites thiab ntxuav lawv kom tau txais cov kab mob monoclonal. .

Kev npaj ntawm polyclonal tshuaj tiv thaiv tsis yog qhov nyuaj li cov tshuaj tiv thaiv monoclonal. Koj tsuas yog yuav tsum tau txhaj tshuaj antigen (siab dua qhov ua kom huv, zoo dua) ncaj qha rau hauv lub cev tsiaj txhawm rau txhaj tshuaj tiv thaiv kab mob. Tom qab 3 txog 4 qhov kev txhaj tshuaj tiv thaiv kab mob, tus titer tau txais kev tsim nyog los ntawm ELISA, thiab cov ntshav tau muab tso rau hauv ib nrab. Tus supernatant tuaj yeem ua kom huv kom tau txais cov tshuaj tiv thaiv polyclonal. Yog li ntawd, kev npaj ua ntu zus ntawm cov tshuaj tiv thaiv kab mob polyclonal yog luv dua li cov tshuaj tiv thaiv kab mob monoclonal, thiab tus nqi pib npaj qis dua li ntawm cov tshuaj tiv thaiv monoclonal.

3. Cons thiab Pros

A. Monoclonal antibody

Zoo:

• Kev lees paub tshwj xeeb ntawm epitope ntawm antigen
• Immortal hybridoma cell kab muaj peev xwm los tsim cov tshuaj tiv thaiv kab mob tsis txwv
• Immortal hybridoma cell kab muaj peev xwm los tsim cov tshuaj tiv thaiv kab mob tsis txwv
• Tsawg suab nrov tom qab thiab hla kev tawm tsam
• Zoo heev affinity thiab purification

Zoo heev affinity thiab purification:

• Kev tsim cov tshuaj tiv thaiv kab mob monoclonal siv sijhawm thiab xav tau txuj ci txuj ci siab.
• Lawv tuaj yeem tsim ntau yam tshuaj tiv thaiv tshwj xeeb, tab sis yuav tsis pom nyob hauv ntau hom.
• Raug rau kev hloov pauv hauv epitopes. Txawm tias kev hloov pauv me me hauv kev hloov pauv tuaj yeem ua rau muaj peev xwm txo qis kev sib khi.

B. Polyclonal antibody

Zoo:

• Tus nqi yog pheej yig thiab kev tsim khoom yog nrawm dua.
• Vim yog kev lees paub ntawm ntau yam epitopes, tag nrho cov tshuaj tiv thaiv kev sib raug zoo rau antigen yog siab dua.
• Kev nkag siab siab rau tshuaj xyuas cov protein tsawg.
• Muaj peev xwm muaj zog txhawm rau ntes lub hom phiaj protein (pom zoo raws li ntes cov tshuaj tiv thaiv kab mob hauv cov qhaub cij ELISA).
• Cov tshuaj tiv thaiv kab mob sib kis tuaj yeem ua rau nrawm nrawm rau lub hom phiaj antigen (pom zoo rau kev tshuaj xyuas uas xav tau ceev cov protein; xws li IP lossis ChIP).
• Cov tshuaj reagents siab dua txhawm rau txheeb xyuas cov protein ntuj.
• Yooj yim rau ob niam txiv nrog cov cim tshuaj tiv thaiv kab mob, nws tsis zoo li yuav cuam tshuam rau kev muaj peev xwm khi.

tsis zoo:

• Qhov sib txawv ntawm cov tsiaj sib txawv ntawm lub sijhawm sib txawv.
• Txij li nws tau lees paub ntau yam epitopes (koom nrog cov tshuaj tiv thaiv kab mob ua kom pom qhov tsawg kawg nkaus ntawm kev ua haujlwm sib cuam tshuam), qhov muaj peev xwm hla kev ua kom rov zoo siab.

4. Lwm qhov sib txawv

xaus

Qhov sib txawv nruab nrab ntawm monoclonal thiab polyclonal antibodies yog qhov loj heev, thiab cov kev sib txawv no ua rau lawv muaj txiaj ntsig hauv cov haujlwm sib txawv. Cov kab mob monoclonal nyob hauv qhov chaw tseem ceeb hauv tshuaj, thaum polyclonal tshuaj tiv thaiv ci ntsa iab hauv thaj tsam ntawm kev ua liaj ua teb thiab nrhiav kab mob thiab kho mob.

References

• ProteoGenix: POLYCLONAL VS. MONOCLONAL: YUAV UA LI CAS YUAV TSUM xaiv qhov zoo tshaj plaws tiv thaiv kab mob rau koj txoj haujlwm?
• LibreTexts: Kev Siv Tau Zoo ntawm Monoclonal thiab Polyclonal Antibodies