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Kutheni le nto inkcubeko yeeseli idibanisa iiseli ezibomvu zegazi kuqala?

2022-12-23

Intshayelelo esisiseko
I-Erythrocyte lysate yenye yezona ndlela zilula kwaye zilula zokususa iiseli ezibomvu zegazi, oko kukuthi, ukwahlula iiseli ezibomvu zegazi nge-lysate, engonakalisi iiseli ze-nucleated kwaye inokususa ngokupheleleyo iiseli ezibomvu zegazi. I-Lysate cleavage yindlela yokususa iiseli ezibomvu zegazi, esetyenziswa ikakhulu ukwahlula kunye nokucocwa kweeseli zezicubu ezisasazwe yi-enzyme digestion, ukwahlukana kunye nokucocwa kwe-lymphocytes, kunye nokususwa kweeseli ezibomvu zegazi kuvavanyo lweprotheni yezicubu kunye ne-nucleic. ukutsalwa kweasidi. Iiseli zezicubu ezifunyenwe yi-lysate yeeseli ezibomvu zegazi aziqukethe iiseli ezibomvu zegazi, kwaye zingasetyenziselwa ngakumbi inkcubeko yokuqala, ukudibanisa kweeseli, ukuhamba kwe-cytometry, ukuhlukana kunye nokutsalwa kwe-nucleic acid kunye neprotheni, njl.

Imiyalelo yokusetyenziswa
Isampulu yeseli yezicubu
1. Izicubu ezitsha zagaywa yi-pancreas / enzyme okanye i-collagenase kwaye zahlakazwa kwi-single cell suspension, kwaye i-supernatant yalahlwa yi-centrifugation.

2. Thatha i-ELS lysate kwifriji ngo-4â, yongeza i-ELS lysate kwi-cell precipitate kwi-recipitate ye-1: 3-5 (yongeza i-3-5ml ye-lysate kwi-1ml yeseli edibeneyo), vuthela ngobumnene kwaye udibanise.

3. I-Centrifuge kwi-800-1000rpm imizuzu emi-5-8 kwaye ulahle ulwelo oluphezulu olubomvu olucacileyo.

4. Inxalenye ye-precipitated yaqokelelwa kwaye i-centrifuged kunye nesisombululo se-Hank okanye isisombululo senkcubeko ye-serum-free ngamaxesha angama-2-3.

I-5, ukuba ukuqhekeka akuphelelanga / kugqityiwe kunokuphinda kuphindwe amanyathelo 2 kunye ne-3.

6. Iiseli zokurhoxiswa kwiimvavanyo ezilandelayo; Ukuba i-RNA ikhutshwe, kungcono ukwenza njalo kwisisombululo esilungiselelwe ukusuka kwiNyathelo 4 usebenzisa amanzi e-DEPC

Iiseli ezibomvu zegazi zinomjikelezo wobomi obufutshane kakhulu, iintsuku ezingama-120 kuphela, kodwa zivelisa igazi ngokukhawuleza, kwaye kulo mzekelo ziyakwazi ukuhlukana kweeseli, kwaye zezona iiseli ezahlula ngokukhawuleza kuzo zonke, ngoko le seli ibaluleke kakhulu. ngoko ke iluncedo kakhulu kwinkcubeko cell. Ilula kakhulu, ayinayo nayiphi na i-organelles kuyo, iimbumba zeeseli kunye neeprotheni kuphela.

Ngaphambili:Yeyiphi imisebenzi yekhithi ye-ELISA?

Okulandelayo:Iimfuno ze-reagent kunye nokumelwa okusisiseko koxinzelelo lwesisombululo.