Ukuhlalutya i-leukocytes kwigazi elipheleleyo luvavanyo lwesiqhelo kwilebhu yeklinikhi okanye kwibhanki yegazi.Ukugxininiswa kunye nokusebenza kwe-leukocytes yisalathisi esibalulekileyo njengokulawula umgangatho wokugcinwa kwegazi.Ngaphandle kwe-leukocyte, igazi elipheleleyo liqulethe inani elikhulu leeplatelet, iiseli ezibomvu zegazi, okanye i-debris yeselula, ezenza kube nzima ukuhlalutya igazi elipheleleyo ngokuthe ngqo phantsi kwe-microscope okanye i-counter cell cell counter.Iindlela eziqhelekileyo zokubala iiseli ezimhlophe zegazi zibandakanya inkqubo ye-RBC lysis, edla ixesha.
I-AOPI Ubalo lwe-Dual-fluoresces luhlobo lovavanyo olusetyenziselwa ukufumanisa ukugxila kweeseli kunye nokusebenza.Isisombululo siyindibaniselwano ye-acridine orange (ibala eliluhlaza-fluorescent nucleic acid) kunye nepropidium iodide (ibala elibomvu-elikhanyayo le-nucleic acid).I-Propidium iodide (PI) yidayi yokukhutshwa kwe-membrane engena kuphela kwiiseli ezineembrane eziphazamisekileyo, ngelixa i-acridine orange iyakwazi ukungena kuzo zonke iiseli kubemi.Xa zombini iidayi zikhona kwi-nucleus, i-propidium iodide ibangela ukunciphisa i-acridine orange fluorescence nge-fluorescence resonance energy transfer (FRET).Ngenxa yoko, iiseli ze-nucleated ezine-membrane ezingaguqukiyo zingcolisa i-fluorescent eluhlaza kwaye zibalwa njengeziphilayo, kanti iiseli ze-nucleated ezinemibrane edityanisiweyo kuphela zibala umbala obomvu we-fluorescent kwaye zibalwa njengezifileyo xa usebenzisa inkqubo ye-Countstar® Rigel.
I-Countstar Rigel sisisombululo esifanelekileyo kwiimvavanyo ezininzi ezintsonkothileyo zamanani abantu beeseli, ezivumela ukuhlalutya ngokukhawuleza iiseli zegazi ezimhlophe kwigazi elipheleleyo.
Inkqubo yoMfuniselo:
1.Thatha i-20 µl yesampuli yegazi kwaye uhlambulule isampuli kwi-180 µl ye-PBS.
2.Yongeza isisombululo se-12µl AO/PI kwisampulu ye-12µl, exutywe ngobunono kunye nepipette;
3.Zoba 20µl umxube kwisilayidi segumbi;
4.Vumela iiseli ukuba zihlale kwigumbi malunga nomzuzu omnye;
5.Tyala isilayidi kwisixhobo se-Countstar FL;
6.Khetha i-assay "AO / PI Viability", emva koko Faka iSampuli ye-ID kule sampuli.
7.Khetha Dilution ratio, Cell Type, cofa 'Run' ukuqalisa uvavanyo.
Isilumkiso: I-AO kunye ne-PI inokuba yi-carcinogen.Kuyacetyiswa ukuba umqhubi anxibe izixhobo zokuzikhusela (PPE) ukuthintela ukunxibelelana ngokuthe ngqo nolusu kunye namehlo.
Isiphumo:
1. Umfanekiso weNdawo eqaqambileyo wegazi elipheleleyo
Kumfanekiso oqaqambileyo wegazi lonke, ii-WBC's azibonakali phakathi kweeseli ezibomvu zegazi.(Umfanekiso 1)
Umzobo 1 Umfanekiso wentsimi oqaqambileyo wegazi elipheleleyo.
2. Umfanekiso weFluorescence wegazi elipheleleyo
Idayi ye-AO kunye ne-PI zombini zibala i-DNA kwinucleus yeeseli.Ngoko ke, iiPlatelets, iiseli ezibomvu zegazi, okanye i-debris yeselula ayikwazi ukuchaphazela ukugxininiswa kwe-leukocytes kunye nesiphumo sokuphila.I-leukocyte ephilayo (eluhlaza) kunye ne-leukocyte efile (eBomvu) ibonakala ngokulula kwimifanekiso ye-fluorescence.(Umfanekiso 2)
Umfanekiso we-2 Fluorescence Imifanekiso yegazi elipheleleyo.(A).Umfanekiso weAO Channel;(B) Umfanekiso wePI Channel;(C) Hlanganisa imifanekiso ye-AO kunye ne-PI Channel.
3. Ukugxila kunye nokusebenza kwe-leukocytes
I-software ye-Countstar FL ibala ngokuzenzekelayo iiseli zamacandelo amathathu egumbi kwaye ibale ixabiso eliphakathi kwenani elipheleleyo le-WBC cell count (1202), i-concentration (1.83 x 106 cells / ml), kunye ne-% yokuphila (82.04%).Imifanekiso yonke yegazi kunye nedatha ingathunyelwa ngaphandle ngokulula njengePDF, uMfanekiso okanye i-Excel yohlalutyo olongezelelweyo okanye ukugcinwa kwedatha.
Umzobo we-3 umfanekiso weskrini we-Countstar Rigel Software
