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Kungani i-cell culture i-lyse amangqamuzana abomvu egazi kuqala?

2022-12-23

Isingeniso esiyisisekelo
I-Erythrocyte lysate ingenye yezindlela ezilula futhi ezilula zokususa amangqamuzana egazi abomvu, okungukuthi, ukuhlukanisa amangqamuzana abomvu egazi nge-lysate, engalimazi amangqamuzana e-nucleated futhi angasusa ngokuphelele amangqamuzana abomvu egazi. I-Lysate cleavage iyindlela ethambile yokukhipha amangqamuzana egazi abomvu, esetshenziselwa ikakhulukazi ukuhlukaniswa nokuhlanza amangqamuzana ezicubu ahlakazwa ukugaya kwe-enzyme, ukuhlukaniswa nokuhlanzwa kwama-lymphocyte, nokususwa kwamangqamuzana abomvu egazi ekuhlolweni kweprotheyini yezicubu kanye ne-nucleic. ukukhishwa kwe-asidi. Amangqamuzana ezicubu atholwe yi-lysate yamangqamuzana abomvu egazi awanawo amaseli abomvu egazi, futhi angaphinde asetshenziselwe isiko eliyinhloko, ukuhlanganiswa kwamaseli, ukugeleza kwe-cytometry, ukuhlukaniswa nokukhishwa kwe-nucleic acid namaprotheni, njll.

Imiyalo yokusetshenziswa
Isampula yeseli yezicubu
1. Izicubu ezintsha zagaywa amanyikwe/i-enzyme noma i-collagenase futhi zahlakazwa ekumisweni kweseli elilodwa, futhi i-supernatant yalahlwa yi-centrifugation.

2. Thatha i-ELS lysate kusuka esiqandisini ngo-4â, engeza i-ELS lysate kumvula yeseli ngesilinganiso esingu-1:3-5 (engeza u-3-5ml we-lysate ku-1ml weseli elihlanganisiwe), vuthela ngobumnene futhi uhlanganise.

3. I-Centrifuge ku-800-1000rpm imizuzu engu-5-8 futhi ulahle uketshezi oluphezulu olubomvu olucacile.

4. Ingxenye enemvula yaqoqwa futhi yafakwa phakathi nendawo ngesixazululo sika-Hank noma isisombululo se-serum-free culture izikhathi ezingu-2-3.

5, uma ukuqhekeka kungaqediwe/kuphelele kungaphindaphindwa izinyathelo 2 no-3.

6. Amaseli okumiswa kabusha okuhlolwa okulandelayo; Uma i-RNA ikhishiwe, kungcono ukwenza kanjalo esixazululweni esilungiselelwe kusukela ku-Isinyathelo sesi-4 usebenzisa amanzi e-DEPC

Amangqamuzana egazi abomvu anomjikelezo wokuphila omfushane kakhulu, izinsuku eziyi-120 kuphela, kodwa akhiqiza igazi ngokushesha okukhulu, futhi kulokhu akwazi ngokukhethekile ukuhlukana kwamangqamuzana, futhi angamangqamuzana ahlukanisayo ashesha kakhulu kunawo wonke, ngakho leli ngqamuzana libaluleke kakhulu, ngakho-ke iwusizo kakhulu kusiko lwamaseli. Kulula kakhulu, ayinawo ama-organelles kuwo, ama-membranes weseli kanye namaprotheni.

Ngaphambilini:Iyini imisebenzi yekhithi ye-ELISA?

Olandelayo:Izidingo ze-reagent kanye nokumelwa okuyisisekelo kokugxila kwesixazululo.