Ii-antibodies, ezikwabizwa ngokuba yi-immunoglobulins (Ig), ziyi-glycoprotein ezibophelela ngokukodwa kwii-antigens.
 
Ukulungiswa kwe-antibody eqhelekileyo kuveliswa ngokugonywa kwezilwanyana kunye nokuqokelela i-antiserum.Ngoko ke, i-antiserum ihlala iqulethe izilwa-buhlungu ezichasene nezinye ii-antigens ezingahambelaniyo kunye namanye amacandelo eprotheni kwi-serum.Iimolekyuli ze-antigen ngokubanzi ziqulathe ii-epitopes ezininzi ezahlukeneyo, ngoko ke izilwa-buhlungu eziqhelekileyo zikwangumxube wezilwa-buhlungu ezichasene nee-epitopes ezininzi ezahlukeneyo.Nkqu nezilwa-buhlungu zeserum eziqhelekileyo ezijoliswe kwi-epitope enye zisaqulunqwa zizilwa-buhlungu ezingafaniyo eziveliswa zii-clones zeeseli ze-B ezahlukeneyo.Ke ngoko, ii-antibodies ze-serum eziqhelekileyo zikwabizwa ngokuba zii-polyclonal antibodies, okanye ii-polyclonal antibodies ngokufutshane.
 
I-antibody ye-Monoclonal (i-antibody eyi-monoclonal) li-antibody efanayo kakhulu eveliswe yi-cell clone ye-B enye kwaye ijoliswe kuphela kwi-epitope ethile.Ngokuqhelekileyo ilungiselelwe iteknoloji ye-hybridoma-iteknoloji ye-antibody ye-hybridoma isekelwe kwi-teknoloji ye-cell fusion, ukudibanisa iiseli ze-B kunye nokukwazi ukukhupha ama-antibodies athile kunye neeseli ze-myeloma ezinamandla okukhula okungapheliyo kwi-B-cell hybridomas.Le seli ye-hybridoma ineempawu zeseli yomzali.Inokukhula ngokungenasiphelo nangokungafiyo kwi-vitro njengeeseli ze-myeloma, kwaye inokudibanisa kwaye ikhuphe izilwa-buhlungu ezithile ezifana ne-splenic lymphocytes.Ngokusebenzisa i-cloning, umgca we-monoclonal ophuma kwiseli enye ye-hybridoma, oko kukuthi, i-hybridoma cell line, inokufumaneka.Izilwa-buhlungu ezizivelisayo zizilwa-buhlungu ezilingana ne-homogenous ngokuchasene ne-antigenic determinant, oko kukuthi, izilwa-buhlungu ze-monoclonal.
 
Ii-antibodies zikhona njenge-monomer enye okanye ngaphezulu ezimile njengo-Y (oko kukuthi, izilwa-buhlungu ze-monoclonal okanye izilwa-buhlungu zepolyclonal).Imonoma nganye emile okwe-Y yenziwe ngamatyathanga e-polypeptide e-4, kubandakanywa amatyathanga amabini afanayo anzima kunye namatyathanga amabini afanayo okukhanya.Itsheyini elikhanyayo kunye nekhonkco elinzima libizwa ngokobunzima bemolekyuli.Umphezulu wesakhiwo se-Y ngummandla oguquguqukayo, oyindawo yokubopha i-antigen.(Icatshulwe kwi-Detai Bio-Monoclonal Antibody Concept)
 
Ulwakhiwo lwe-antibody
Itsheyini elinzima
Zintlanu iindidi zemixokelelwane enzima ye-mammalian Ig, ebizwa ngoonobumba besiGrike u-α, δ, ε, γ, kunye no-μ.Izilwa-buhlungu ezihambelanayo zibizwa ngokuba yi-IgA, IgD, IgE, IgG, kunye ne-IgM.Amakhonkco anzima ahlukeneyo ahluke ngobukhulu kunye nokubunjwa.α kunye ne-γ ziqulethe malunga ne-450 amino acids, ngelixa i-μ kunye ne-ε iqulethe malunga ne-550 amino acids.
Ikhonkco ngalinye elinzima linemimandla emibini: ummandla osisigxina kunye nommandla oguquguqukayo.Zonke izilwa-buhlungu zohlobo olufanayo zinengingqi efanayo engaguqukiyo, kodwa kukho umahluko phakathi kwezilwa-buhlungu zeentlobo ezahlukeneyo.Imimandla engaguqukiyo yamatyathanga anzima γ, α, kunye ne-δ yenziwe ngamacandelo amathathu e-Ig kwi-tandem, kunye nommandla we-hinge ukwandisa ukuguquguquka kwayo;imimandla engaguqukiyo yamatyathanga anzima μ kunye no-ε yenziwe ngamathambeka angama-4 e-Ig.Ummandla oguquguqukayo wetsheyini elinzima le-antibody eliveliswa ngamaseli e-B ahlukeneyo ahlukile, kodwa ummandla oguquguqukayo we-antibody oveliswa yi-cell B enye okanye i-cell clone yeseli iyafana, kwaye ummandla oguquguqukayo wekhonkco ngalinye elinzima malunga ne-110 amino acids ubude., Kwaye wenze i-domain ye-Ig enye.
 
Ikhonkco elikhanyayo
Zimbini kuphela iindidi zamatyathanga okukhanya kwizilwanyana ezanyisayo: uhlobo lwe-lambda kunye nodidi lwe-kappa.Ikhonkco ngalinye lokukhanya linemimandla emibini edibeneyo: ummandla osisigxina kunye nommandla oguquguqukayo.Ubude bekhonkco lokukhanya malunga ne-211 ~ 217 amino acids.Amatyathanga okukhanya amabini akwi-antibody nganye ahlala efana.Kwizilwanyana ezanyisayo, ikhonkco lokukhanya kwi-antibody nganye linodidi olunye kuphela: i-kappa okanye i-lambda.Kwezinye ii-vertebrates ezisezantsi, ezifana neentlanzi ze-cartilage (i-cartilage fishes) kunye ne-bony fish, ezinye iintlobo zamatyathanga okukhanya afana ne-iota (iota) uhlobo nazo zifunyenwe.
 
Amacandelo eFab kunye neFc
Icandelo le-Fc linokudityaniswa ngokuthe ngqo kunye ne-enzymes okanye idayi ye-fluorescent ukulebula amajoni omzimba.Yinxalenye apho i-antibody rivets kwi-plate ngexesha lenkqubo ye-ELISA, kwaye ikwayinxalenye apho i-antibody yesibini ibonwa kwaye iboshwe kwi-immunoprecipitation, i-immunoblotting kunye ne-immunohistochemistry.Ii-antibodies zinokuthi zenziwe nge-hydrolyzed zibe ngamacandelo amabini e-F (ab) kunye necandelo elinye le-Fc nge-proteolytic enzymes ezifana ne-papain, okanye zinokwaphulwa ukusuka kummandla we-hinge nge-pepsin kunye ne-hydrolyzed kwi-F (ab) 2 yecandelo kunye ne-Fc enye.Amaqhekeza e-antibody e-IgG ngamanye amaxesha aluncedo kakhulu.Ngenxa yokungabikho kwecandelo le-Fc, icandelo le-F (ab) aliyi kudlula kunye ne-antigen, kwaye ayiyi kubanjwa ngamaseli omzimba kwizifundo ze-vivo.Ngenxa yamaqhekeza amancinci e-molecular kunye nokungabikho komsebenzi wokudibanisa (ngenxa yokungabikho kwecandelo le-Fc), icandelo le-Fab lihlala lisetyenziselwa i-radiolabeling kwizifundo zokusebenza, kwaye icandelo le-Fc lisetyenziswa kakhulu njenge-agent yokuthintela kwi-histochemical staining.
 
Iingingqi eziguquguqukayo neziqhubekayo
Ummandla oguquguqukayo (ummandla we-V) ufumaneka kwi-1/5 okanye i-1/4 (equlethe malunga ne-118 amino acid iintsalela) ye-chain ye-H kufuphi ne-N-terminus kunye ne-1/2 (equlethe malunga ne-108-111 amino acid iintsalela) kufuphi ne-N-terminus ye-chain L.Ummandla ngamnye we-V uneringi ye-peptide eyenziwe ngamabhondi e-intra-chain disulfide, kwaye iringi nganye yepeptide iqulethe malunga nama-67 ukuya kuma-75 entsalela ye-amino acid.Ukubunjwa kunye nokucwangciswa kwee-amino acids kummandla we-V kumisela i-antigen yokubopha ngokukodwa kwe-antibody.Ngenxa yeentlobo ezisoloko zitshintsha kunye nokulandelelana kwee-amino acids kummandla we-V, iintlobo ezininzi ze-antibodies ezineempawu ze-antigen ezibophelelayo ezahlukeneyo zinokuqulunqwa.Imimandla ye-V ye-L chain kunye ne-H chain ibizwa ngokuba yi-VL kunye ne-VH, ngokulandelanayo.Kwi-VL kunye ne-VH, ukubunjwa kwe-amino acid kunye nokulandelelana kweminye imimandla yendawo ineqondo eliphezulu lokuhluka.Le mimandla ibizwa ngokuba yi-hypervariable regions (HVR).Ukuqulunqwa kwe-amino acid kunye nokulungiswa kwamalungu angeyiyo i-HVR kwingingqi ye-V ngokufanelekileyo, ebizwa ngokuba ngummandla wesikhokelo.Kukho imimandla emithathu ye-hypervariable kwi-VL, edla ngokufumaneka kwiintsalela ze-amino acid 24 ukuya kuma-34 kunye nama-89 ukuya kuma-97 ngokulandelelanayo.Ii-HVR ezintathu ze-VL kunye ne-VH zibizwa ngokuba yi-HVR1, i-HVR2 kunye ne-HVR3, ngokulandelelanayo.Uphando kunye nohlalutyo lwe-X-ray crystal diffraction lubonise ukuba ummandla we-hypervariable ngokwenene yindawo apho i-antibody antigen ibophelela khona, ngoko ibizwa ngokuba ngummandla wokugqiba ukuhambelana (CDR).I-HVR1, i-HVR2 kunye ne-HVR3 ye-VL kunye ne-VH inokubizwa ngokuba yi-CDR1, i-CDR2 kunye ne-CDR3 ngokulandelelanayo.Ngokubanzi, i-CDR3 ineqondo eliphezulu lokungaguquguquki.Ummandla we-hypervariable ikwayindawo ephambili apho i-idiotypic determinants of Ig molecules zikhona.Kwiimeko ezininzi, i-chain ye-H idlala indima ebaluleke kakhulu ekubopheni i-antigen.
Ummandla ongaguqukiyo (ummandla C)ifumaneka kwi-3/4 okanye i-4/5 (malunga ne-amino acid 119 ukuya kwi-terminal C) ye-chain ye-H kufuphi ne-C terminus kunye ne-1/2 (iqulethe malunga ne-105 amino acid iintsalela) kufuphi neC terminus ye-L chain.Ummandla ngamnye osebenzayo we-H chain uqulethe malunga ne-110 i-amino acid iintsalela, kwaye iqulethe i-peptide ring eyenziwe ngama-amino acid angama-50-60 adityaniswe ngamabhondi e-disulfide.Ukwakhiwa kwe-amino acid kunye nolungiselelo lwalo mmandla ngokuthe rhoqo kwisilwanyana esifanayo Ig isotype L chain kunye nohlobo olufanayo H chain.Ngokufanayo, inokubophelela ngokukodwa kwi-antigen ehambelanayo, kodwa isakhiwo sommandla wayo we-C siyafana, oko kukuthi, sine-antigenicity efanayo.I-antibody ye-IgG yesibini yehashe (okanye i-anti-antibody) inokudityaniswa nezi zimbini A indibaniselwano yezilwa-buhlungu (IgG) ngokuchasene neexotoxins ezahlukeneyo zenzeka.Esi sisiseko esibalulekileyo sokulungiselela izilwa-buhlungu zesibini kunye nokusebenzisa i-fluorescein, isotopes, i-enzymes kunye nezinye izilwa-buhlungu ezibhalwe phantsi.
 
 
Iimveliso ezinxulumeneyo:
Isethi yeSeli eNgqo RT-qPCR