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Ukuqonda i-Paraformaldehyde: Izicelo & Izingozi
Kwadalwa ngo 12.04
Ukuqonda i-Paraformaldehyde: Izicelo & Izingozi
Isingeniso: Indima ye-Paraformaldehyde ekuhloleni kweBiological Fixation
I-Paraformaldehyde iyisithako esibalulekile esisetshenziswa kakhulu ocwaningweni lwezobuchwepheshe bezempilo, ikakhulukazi ekugcineni izicubu nezicubu. Njengendlela ye-polymeric ye-formaldehyde, i-paraformaldehyde isebenza njengomthombo wegesi ye-formaldehyde uma ixutshwe namanzi, ethinta ama-proteins futhi igcine ukwakheka kwezicubu. Le nqubo yokugcinwa ibalulekile ukuze kugcinwe amasampula ezempilo esimweni esifana nempilo ukuze kuhlolwe ngemicroscope kanye nokuhlaziywa kwemolekyuli. Izici zokugcinwa kwe-paraformaldehyde ziyasiza ukuvimbela i-autolysis nokubola, kuqinisekisa ukuzinza kwe-antigens nezakhiwo zezicubu ukuze kuhlolwe ngemva kokufaka imibala nokuthwebula.
E-histology nase-cytology, i-paraformaldehyde ibhekwa kakhulu njengezinga eliphezulu lokugcina ngenxa yokukwazi kwayo ukugcina ubunjalo bezicubu ngaphandle kokuphazamiseka okukhulu. Ukusetshenziswa kwayo kabanzi emalabhorethri emhlabeni wonke kugcizelela ukubaluleka kwayo ocwaningweni nasekuxilongeni. Ngaphezu kwalokho, izici ze-chemical ze-paraformaldehyde zenza ukuthi ikwazi ukungena ezicubwini kahle futhi iphinde iphendule namaqembu e-amino kumaprotheni, yakha amabhuloho e-methylene aqinisa isampula. Ngokubheka lezi zinzuzo, ukuqonda izakhiwo, izicelo, kanye nezingozi ezingaba khona ze-paraformaldehyde kubalulekile kubacwaningi abasebenza nezampula ze-biological.
Izici ze-Paraformaldehyde: Iziphumo kanye Nezindlela Zokubopha
I-Paraformaldehyde iyisidakamizwa esimhlophe, esiqinile esakhiwe ngama-polymerized formaldehyde units. Lapho kushiswa noma kuhlanganyelwa ezinhlelweni zamanzi, iyadedelela ukukhulula i-formaldehyde, i-aldehyde esebenzayo eyakha ama-crosslinks e-covalent phakathi kweqembu le-amino kuma-proteins. Le ndlela yokuxhumanisa ibhekelele umphumela wokubopha, "ivala" kahle izingxenye ze-cell. Izinga lokubopha elitholakalayo lincike ezimeni ezifana nokugxila kwe-paraformaldehyde, i-pH, izinga lokushisa, kanye nesikhathi sokub exposure.
Uma kuqhathaniswa nezinye izixazululo, i-paraformaldehyde inikeza ibhalansi phakathi kokugcina izicubu nokugcina ama-antigen, okwenza kube ukukhetha okufanele kwe-immunohistochemistry kanye ne-immunofluorescence protocols. Ivame ukulungiswa entsha ngokuxuba uthuli lwe-paraformaldehyde emanzini anokuphathwa kwe-phosphate (PBS) nokulungisa i-pH ibe ne-neutrali. Ukuqina nezimo zokugcina kubalulekile, njengoba ukuphathwa okungafanele kungaholela ekwehliseni nasekunciphiseni ukusebenza kokufaka. Lezi zici ziqhakambisa isidingo seziqondiso ezijwayelekile ukuze kuqinisekiswe ukuphindaphindwa nokugcina okuhle.
Izicelo KwiSifundo: Ukusetshenziswa kuHistology, Cytology, kanye Nokugcina I-Protein
I-Paraformaldehyde isetshenziswa kakhulu emikhakheni ehlukene yocwaningo, kuhlanganise ne-histology, cytology, molecular biology, kanye ne-neuroscience. Ku-histology, isetshenziswa ukuvimbela izingxenye zezicubu, igcina imininingwane ye-cellular futhi ivumela izifundo ezichanile ze-morphological. Abacwaningi be-cytology basebenzisa ukuvimbela kwe-paraformaldehyde ukuze bagcine ama-smears we-cell kanye nezixazululo ukuze zihlolwe nge-microscope. Ngaphezu kwalokho, ukuvimbela kwe-paraformaldehyde kubalulekile ekuvikelweni kwe-protein epitopes ngesikhathi sokuhlolwa kwe-antibody, okuyisisekelo ku-immunohistochemistry, western blotting, kanye ne-flow cytometry.
Ekuhloleni kwezimiso zengqondo, ukujolisa kwe-paraformaldehyde kugcina izicubu zengqondo nezakhiwo ze-neuronal, kuthuthukisa ukufundwa kwemijikelezo ye-neural kanye nokuhlonza kweprotein. Lokhu ku-fixative kusiza futhi ekugcineni ama-nucleic acids kanye nama-lipids ngezinga elithile, kusekela izindlela ezihlukahlukene zokuhlaziya. Ngaphezu kwalokho, amasampula agcinwe nge-paraformaldehyde ahambisana nezindlela zokubona ezithuthukile ezifana ne-confocal microscopy kanye ne-electron microscopy ngemuva kokucubungula okwengeziwe. Lezi zicelo ezibanzi zikhombisa ubuchwepheshe be-paraformaldehyde kanye nokubaluleka kwayo ezinsizakalweni ze-biological.
Izinto okufanele zicatshangelwe ngaphambi kokufaka: Izizathu nezinkinga
Ukufaka ngaphambi kokufaka kubhekisela ekwelashweni kokuqala kwezibonelo ze-biological ngaphambi kokufaka okukhulu okwenziwa nge-paraformaldehyde. Le nqubo kwesinye isikhathi isetshenziswa ukuze iqinise izibonelo noma ikhiphe izingxenye ezingaphandle kodwa ingase ilethe izinkinga. Enye yezizathu zokufaka ngaphambi kokufaka ukuwunciphisa umphumela wokungcoliswa kwangemuva noma ukuthuthukisa ukungena kokufaka. Nokho, ukufaka ngaphambi kokufaka kungase kuphinde kushintshe izimo ze-protein, kwehlise ama-antigens abucayi, noma kuholele ekufakeni okungalingani, kuthinta ikhwalithi yokuhlaziywa okuphuma.
Abacwaningi kufanele bahlolisise ngokucophelela isidingo seziqondiso zokufaka ngaphambi kokufaka ngokuya ngohlobo lwesampula kanye nokuhlaziywa okuhlosiwe. Kwezinye izimo, ukweqa ukufaka ngaphambi kokufaka nokusebenzisa i-paraformaldehyde ngqo kuholela ekugcinweni okungcono nasekuphindaphindeni. Ukuqonda lezi zinkinga kubalulekile ukuze kuthuthukiswe iziqondiso zokufaka, kuncishiswe ama-artifact, futhi kugcinwe i-antigenicity ukuze kutholakale imiphumela ethembekile yokuhlola i-immunostaining.
Imihlahlandlela Yokuhlola Amantshontsho: Izindlela Ezinhle Ngemva Kokulungiswa
Ngemuva kokufakwa kwe-paraformaldehyde, izinqubo zokuhlola ama-antibody zidinga ukuhlela ukuze zifeze izimpawu ezithile nezinamandla. Inqubo yokufaka ingase ibhidlize ama-epitope, okwenza kudingeke izindlela zokubuyisela i-antigen ezifana nokubuyiselwa kwe-epitope okubangelwa ukushisa noma ukugaywa kwe-enzyme. Ukukhetha ama-buffer wokuvimba afanele kanye nezimo zokuhlala kusiza ukunciphisa ukuhlanganiswa okungafanele kanye noise yangemuva. Kubalulekile futhi ukusebenzisa ama-antibody aphakanyisiwe aphumelele ahambisana nezibonelo ezifakiwe.
Abacwaningi kumele balandele izinqubo ezijwayelekile, kuhlanganise nezinyathelo zokuhlanza ezinembile kanye nokusebenzisa izilawuli ukuze kuqinisekiswe ukujoliswa kokufaka umbala. Ukuhlonishwa okukhethekile kufanele kunikezwe isikhathi sokufaka nokugxila kwe-parafomaldehyde, njengoba ukufaka kakhulu kunganciphisa ukufinyelela kwe-antibody. Ngokulandela lezi zindlela ezinhle, ososayensi bangakwazi ukwandisa ubuhlakani nokunembile kokuhlaziywa kwe-immunohistochemical kanye ne-immunocytochemical besebenzisa amasampula afakwe ku-parafomaldehyde.
Ukuhlola Nokutholakele: Isifinyezo Sokuhlolwa Kwamagciwane Nezimpumelelo
Ukuhlolwa okuningi kwezicubu ze-antibody ezisuswe ku-paraformaldehyde kukhombisa ubukhulu obuhlukahlukene bokuthinteka kuye ngento ethile kanye nezimo zokufaka. Izifundo eziningi zikhuluma ukuthi iningi le-antibodies ligcina amandla awo okuxhumana ngemuva kokufakwa uma izinyathelo ezifanele zokubuyisela izinto zisetshenziswa. Nokho, ezinye izindawo ezithile zihluleka ekuxhumaneni futhi zingase zidinge izindlela zokufaka ezihlukile noma izixazululo ezithambile.
Ukwahlulelwa kwamuva kubonise ukuthi ukuvikelwa kwe-paraformaldehyde kugcina ubunjalo bezicubu ngenkathi kugcinwa ukuqashelwa kwe-antibody kwezinhlobonhlobo zamaprotheni, kuhlanganise nezithako ze-membrane receptors, izakhi ze-cytoskeleton, kanye nezinto eziphilayo ze-nuclear. Lezi zifundo ziqhakambisa ukwethembeka kwe-paraformaldehyde njengokwakhiwa kokuvikela emisebenzini ye-immunostaining, kodwa futhi ziqhakambisa ukubaluleka kokwenza kahle ngokwezifiso kwantoni ngayinye ye-antibody kanye nohlobo lwezicubu.
Ukubopha Ngemuva Kokufaka Umbala: Izindlela Nezinto Eziphathelene Nokuzinza KweDye
Emaphutheni athile, ukulungiswa kwenziwa ngemva kokufaka umbala ukuze kuqinisekiswe ukuthi umbala uhlala unqamuki futhi kugwenywe ukulahleka kwesignali ngesikhathi sokuthwebula. Ukulungiswa ngemva kokufaka umbala nge-parafomaldehyde kungathuthukisa ukuqina kwesignali ye-fluorescence, ikakhulukazi kumabala e-fluorescent abuthakathaka ekukhanyeni noma ekonakaleni kwemichilo. Nokho, le ndlela idinga ukuphathwa ngokucophelela ukuze kugwenywe ukuguqulwa kokubekwa noma amandla kombala.
Abacwaningi kumele balinganisela phakathi kokubambelela okwanele ukuze bagcine isignali nokugwema ukuxhumana okweqile okungase kuphumelele ukukhanya. Ngaphezu kwalokho, ukukhetha imidiya yokufaka kanye nezithako zokulwa nokuphuma kwefaded kuhlangene nokubambelela ukuze kuqinisekiswe ukuzinza kwemibala. Lezi zinqubo zibalulekile ikakhulukazi ekubukeni okuphakeme kwe-microscopy nasekugcineni amasampula isikhathi eside.
Izinto Ezikhethekile: GFP Fluorescence Nezindlela Ezingezona Ezokukhanya
Iphrotheni eluhlaza elikhanyayo (GFP) kanye nezinsalela zayo zisetshenziswa kabanzi njengemakha ye-in vivo ocwaningweni lwezobuchwepheshe. Nokho, ukuvimbela kwe-parafomaldehyde kunganciphisa ukukhanya kwe-GFP ngenxa yokuxhumana kwemichiza nokushisa. Abacwaningi bavame ukusebenzisa izikhathi zokuvimbela ezimfishane, ukunciphisa izinga le-parafomaldehyde, noma izixazululo ezihlukile ezifana ne-methanol ukuze bagcine izimpawu ze-GFP.
Ngaphezu kwalokho, izindlela ezintsha zokubopha kanye nezinhlelo ezithuthukisiwe ziye zathuthukiswa ukuze zilinganiswe ukugcina izicubu nokugcina ukukhanya kwe-GFP. Ukuqonda lezi zici kubalulekile ezexperimentini ezihilela amaprotein akhanyayo. Izinketho ezifana nokubopha nge-cryo noma ukuboniswa kwezicubu eziphilayo zingase zengezelele ukubopha kwe-paraformaldehyde uma ukugcina ukukhanya kubalulekile.
Isiphetho: Ukubaluleka Kwezindlela Zokubopha Ocwaningweni Lwezobunjiniyela
I-Paraformaldehyde iseyindida engabalulekile ekugcineni emiphumeleni ye-biological kanye ne-biomedical research, inikeza ukugcinwa okuphumelelayo kwemorphology ye-tissue kanye ne-protein antigenicity. Ukusetshenziswa kwayo kabanzi ku-histology, immunohistochemistry, kanye ne-molecular biology kugcizelela ukuhamba kwayo nokwethembeka. Nokho, izinqubo zokugcinwa kumele ziqondiswe ngokucophelela kumkhiqizo, imigomo yocwaningo, kanye nezicelo ezilandelayo ukuze kugwenywe ama-artifacts futhi kuthuthukiswe ikhwalithi yedatha.
Ulwazi ngezimpawu ze-paraformaldehyde, izindlela zokufaka, kanye nezindlela ezinhle kakhulu kubalulekile kubacwaningi ukuze bakwazi ukusebenzisa amandla ayo aphelele. Ngaphezu kwalokho, izinkampani ezifana neHebei Jintaida Chemical Co., Ltd.uhola imboni ngokuhlinzeka ngemikhiqizo ye-paraformaldehyde yekhwalithi ephezulu ehambisana nezindinganiso eziqinile, kusekela intuthuko yezesayensi nezimboni emhlabeni jikelele.
Izinkomba
Izifundo ezikhethiwe kanye nezibuyekezo mayelana nokufakwa kwe-paraformaldehyde, izici zayo zamakhemikhali, kanye nezicelo zayo ezinqubweni zokuhlola imiphumela yokuvikela kanye ne-microscopy zinikeza ukuqonda okubalulekile kubacwaningi. Lokhu kufaka phakathi imibhalo eyisisekelo ngezindlela ze-histological, izindatshana ezibuyekeziwe ngabanye mayelana nokuthuthukiswa kokufakwa kwe-antibody, kanye nezincwadi zobuchwepheshe ezivela kubakhiqizi bamakhompiyutha.
Izinsiza Ezengeziwe
Ukuze baqhubeke nokufunda, abafundi bangahlola izihloko ezihlobene ezifana neukuhlola i-flow cytometryamathiphu kanyeI-ELISA izindlela, evamise ukusebenzisa amasampula aqinisiwe nge-paraformaldehyde ukuze kuhlaziywe amaseli nokulinganiswa kweprotein. Lezi zinsiza zinikeza ukuqonda okuphelele kanye nezikhombisi-ndlela ezisebenzayo zokusebenza kwezocwaningo lwezinto eziphilayo.
Xhumana
Shiya ulwazi lwakho futhi sizokuthinta.
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