Cryovialsziqhele ukusetyenziselwa ugcino cryogenic yemigca yeseli kunye nezinye izinto ezibalulekileyo zebhayoloji, kwi dewars ezaliswe nitrogen engamanzi.

Kukho amanqanaba amaninzi abandakanyekayo ekugcinweni ngempumelelo kweeseli kwinitrogen engamanzi. Ngelixa umgaqo osisiseko kukukhenkce okucothayo, olona buchule busetyenziswayo buxhomekeke kuhlobo lweseli kunye ne-cryoprotectant esetyenziswayo. Kukho iingqwalasela ezininzi zokhuseleko kunye neendlela ezifanelekileyo zokuthathela ingqalelo xa ugcina iiseli kumaqondo obushushu aphantsi ngolo hlobo.

Esi sithuba sijolise ekunikeni isishwankathelo sendlela i-cryovials egcinwa ngayo kwi-nitrogen engamanzi.

Yintoni iCryovials

I-Cryovials zincinci, iibhotile ezivaliweyo ezilungiselelwe ukugcina iisampulu zolwelo kumaqondo obushushu aphantsi kakhulu. Baqinisekisa ukuba iiseli ezigcinwe kwi-cryoprotectant azidibani ngokuthe ngqo ne-nitrogen engamanzi, ukunciphisa umngcipheko wokuqhekeka kweeseli ngelixa zixhamla kwisiphumo sokupholisa ngokugqithisileyo se-nitrogen engamanzi.

Iingqayi zidla ngokufumaneka kwiindidi zemiqulu kunye noyilo - zinokuthi zifakwe ngaphakathi okanye ngaphandle ngeentambo eziphantsi okanye ezinqamlekileyo. Iifomathi eziNdlongo nezingazali nazo ziyafumaneka.

Ngubani OsebenzisaIiCyrovialsUkugcina iiSeli kwiNitrogen engamanzi

Uluhlu lwe-NHS kunye neebhubhoratri zabucala, kunye namaziko ophando akhethekileyo kwibhanki yegazi, i-epithelial cell biology, i-immunology kunye ne-stem cell biology isebenzisa i-cryovials ukuze i-cryopreserve cells.

Iiseli ezigcinwe ngale ndlela ziquka iiseli ze-B kunye ne-T, iiseli ze-CHO, i-Hematopoietic Stem kunye ne-Progenitor Cells, i-Hybridomas, i-Intestinal Cells, i-Macrophages, i-Mesenchymal Stem kunye ne-Progenitor Cells, i-Monocytes, i-Myeloma, i-NK Cells kunye ne-Pluripotent Stem Cells.

Isishwankathelo sendlela yokugcina iiCryovials kwiNitrogen yoLiquid

I-Cryopreservation yinkqubo egcina iiseli kunye nolunye ulwakhiwo lwebhayoloji ngokuzipholisa kumaqondo obushushu aphantsi kakhulu. Iiseli zinokugcinwa kwinitrogen engamanzi iminyaka ngaphandle kokulahleka kokusebenza kweeseli. Olu ludweliso lweenkqubo ezisetyenzisiweyo.

Ukulungiselela iSeli

Indlela echanekileyo yokulungiselela iisampulu iya kwahluka ngokuxhomekeka kuhlobo lweeseli, kodwa ngokubanzi, iiseli ziqokelelwa kwaye zifakwe kwi-centrifuged ukuphuhlisa i-pellet ene-cell-rich. Le pellet iphinda imiswe kwi-supernatant exutywe ne-cryoprotectant okanye i-cryopreservation medium.

Cryopreservation Medium

Esi sixhobo sisetyenziselwa ukugcina iiseli kwiindawo ezinobushushu obuphantsi eziza kulawulwa ngokuthintela ukwakheka kweekristale zangaphakathi kunye nezangaphandle zeseli kwaye ngenxa yoko ukufa kweeseli. Indima yabo kukubonelela ngendawo ekhuselekileyo, ekhuselayo yeeseli kunye nezicubu ngexesha lokukhenkceza, ukugcinwa, kunye neenkqubo zokunyibilika.

I-medium efana ne-frozen plasma (FFP), isisombululo se-plasmalyte esine-heparinised okanye i-serum-free, izisombululo zezilwanyana ezingenazo zixutywe kunye ne-cryoprotectants ezifana ne-dimethyl sulphoxide (DMSO) okanye i-glycerol.

I-pellet yesampulu ehlaziyiweyo kwakhona ifakwe kwi-polypropylene cryovials njengeInkampani yaseSuzhou Ace Biomedical Cryogenic Storage Vials.

Kubalulekile ukuba ungagcwalisi i-cryovials njengoko oku kuya kwandisa umngcipheko wokuqhekeka kunye nokukhutshwa kwezinto ezinokuthi zikhutshwe (1).

Ireyithi yokuNqanda eLawulwayo

Ngokubanzi, isantya somkhenkce esilawulwa kancinci sisetyenziselwe ukugcinwa ngempumelelo kweeseli.

Emva kokuba iisampulu zifakwe kwi-vials cryogenic, zibekwe kumkhenkce omanzi okanye kwi-4 ℃ efrijini kwaye inkqubo yokukhenkceza iqaliswe kwimizuzu emi-5. Njengesikhokelo esiqhelekileyo, iiseli zipholiswa ngomlinganiselo we -1 ukuya -3 ngomzuzu (2). Oku kuphunyezwa kusetyenziswa isixhobo esipholileyo esicwangcisiweyo okanye ngokubeka iingqayi kwibhokisi egqunyiweyo ebekwe kwiqondo lokukhenkceza elilawulwa ngu--70°C ukuya ku-90°C.

Ukutshintshela kwi-Nitrogen elulwelo

Iingqayi ze-cryogenic ezikhenkcezisiweyo zithunyelwa kwitanki yenitrogen engamanzi kangangexesha elingenammiselo ukuba ubushushu obungaphantsi kwe -135℃ bugciniwe.

La maqondo obushushu asezantsi kakhulu angafunyanwa ngokuntywiliselwa kulwelo okanye kwisigaba somphunga wenitrogen.

Ulwelo okanye iSigaba soMphunga?

Ugcino kwisigaba solwelo lwenitrogen luyaziwa ngokugcina ubushushu obubandayo obungaguquguqukiyo ngokupheleleyo, kodwa kaninzi alucetyiswa ngezi zizathu zilandelayo:

• Imfuneko yemithamo emikhulu (ubunzulu) yenitrogen engamanzi nto leyo enokuba yingozi. Ukutshisa okanye ukuphefumla ngenxa yoku kungumngcipheko wokwenene.
• Iimeko ezibhaliweyo zokungcoliseka okunqamlezayo ziiarhente ezosulelayo ezifana ne-aspergillus, i-hep B kunye nokusasazeka kwentsholongwane nge-liquid nitrogen medium (2,3)
• Ukubanakho kwenitrogen engamanzi ukuvuza kwiibhotile ngexesha lokuntywiliselwa. Xa isusiwe kwindawo yokugcina kwaye ifudunyezwe kubushushu begumbi, i-nitrogen ikhula ngokukhawuleza. Ngenxa yoko, i-vial inokuqhekeka xa isusiwe kwindawo yokugcina i-nitrogen engamanzi, idala ingozi kuzo zombini inkunkuma ephaphazelayo kunye nokuvezwa kweziqulatho (1, 4).

Ngezi zizathu, ugcino lobushushu obuphantsi kakhulu buxhaphake kakhulu kwisigaba somphunga senitrogen. Xa iisampuli kufuneka zigcinwe kwisigaba solwelo, i-cryoflex tubing ekhethekileyo kufuneka isetyenziswe.

Icala elisezantsi kwisigaba somphunga kukuba iqondo lobushushu elithe nkqo linokwenzeka elikhokelela kuguquko lobushushu phakathi kwe -135℃ kunye ne -190℃. Oku kufuna ukuba kujongwe ngononophelo nangenkuthalo amanqanaba enitrogen yolwelo kunye nokwahluka kobushushu (5).

Abavelisi abaninzi bacebisa ukuba iicryovials zikulungele ukugcinwa ukuya kuthi ga kwi -135℃ okanye zisetyenziswe kwisigaba somphunga kuphela.

Ukunyibilikisa iiseli zakho zeCryopreserved

Inkqubo yokunyibilika icinezela inkcubeko engumkhenkce, kwaye ukuphathwa ngendlela eyiyo kunye nobuchule kuyafuneka ukuqinisekisa ukusebenza kakuhle, ukuchacha, kunye nokusebenza kweeseli. Iiprothokholi ezichanekileyo zokunyibilika ziya kuxhomekeka kwiintlobo zeeseli ezithile. Nangona kunjalo, ukunyibilika ngokukhawuleza kuthathwa njengomgangatho woku:

• Ukunciphisa nayiphi na impembelelo ekubuyiselweni kweselula
• Nceda ukunciphisa ixesha lokuvezwa kwi-solutes ekhoyo kwimidiya yokukhenkcela
• Nciphisa nawuphi na umonakalo ngokwenziwa ngokutsha komkhenkce

Iibhafu zamanzi, iibhafu ezinamaso, okanye izixhobo ezizisebenzelayo ezikhethekileyo ziqhele ukusetyenziselwa ukunyibilikisa iisampulu.

Amaxesha amaninzi umgca weseli om-1 uyanyibilika ngexesha lemizuzu emi-1 ukuya kwemi-2, ngokujija ngobunono kwibhafu yamanzi eyi-37℃ kude kubekho nje intwana yomkhenkce eshiyekileyo kwivial phambi kokuba ihlanjwe kwindawo yokukhula efudunyeziweyo.

Kwezinye iiseli ezifana neembumba ezanyisayo, ukufudumala kancinci kubalulekile ukuze ziphile.

Iiseli ngoku zilungele ukukhuliswa kweeseli, ukuhlukaniswa kweeseli, okanye kwimeko yeeseli ze-haematopoietic stem-uphononongo olusebenzayo ukuqinisekisa ukuthembeka kweeseli ze-donor stem phambi konyango lwe-myeloablative.

Yinto eqhelekileyo ukuthatha ii-aliquots ezincinci zesampulu ehlanjwe kwangaphambili esetyenziselwa ukwenza ubalo lweeseli ukumisela ugxininiso lweeseli zokubekwa kwinkcubeko. Unokuthi emva koko uvavanye iziphumo zeenkqubo zokuhlukaniswa kweeseli kunye nokumisela ukusebenza kweeseli.

IiNdlela eziLungileyo zokuGcina iiCryovials

I-cryopreservation eyimpumelelo yeesampulu ezigcinwe kwi-cryovials ixhomekeke kwizinto ezininzi kwiprotocol kuquka ukugcinwa okufanelekileyo kunye nokugcinwa kwerekhodi.

• Ukwahlula iiseli phakathi kweendawo zokugcina– Ukuba imiqulu ivumela, yahlula iiseli phakathi kweengqayi kwaye uzigcine kwiindawo ezahlukeneyo ukunciphisa umngcipheko welahleko yesampulu ngenxa yokusilela kwezixhobo.
• Thintela ungcoliseko olunqamlezayo-Khetha ukusebenzisa enye ibhotile ye-cryogenic okanye i-autoclave ngaphambi kokusetyenziswa okulandelayo
• Sebenzisa iibhotile ezinobungakanani obufanelekileyo kwiiseli zakho– iingqayi ziza kuluhlu lwemiqulu phakathi kwe-1 kunye ne-5mls. Kuphephe ukuzalisa ii-vials ukunciphisa umngcipheko wokuqhekeka.
• Khetha iibhotile ze-cryogenic zangaphakathi okanye zangaphandle– Iingqayi ezifakwe ngaphakathi zicetyiswa ziiyunivesithi ezithile kumanyathelo okhuseleko – zinokuthintela ungcoliseko ngexesha lokuzaliswa okanye xa zigcinwe kwinitrogen engamanzi.
• Thintela ukuvuza- Sebenzisa izitywina ezifakwe kabini ezibunjwe kwi-screw-cap okanye i-O-rings ukuthintela ukuvuza kunye nongcoliseko.
• Sebenzisa iibhakhowudi ze-2D kunye nelebhile yeebhotile– ukuqinisekisa ukulandeleka, iingqayi ezineendawo zokubhala ezinkulu zenza ukuba ibhotile nganye ibhalwe ngokwaneleyo. Iibhakhowudi ze-2D zinokunceda ngolawulo lokugcina kunye nokugcinwa kwerekhodi. Iikhowudi ezinombala ezinemibala ziluncedo ekuchongeni lula.
• Ukugcinwa okwaneleyo kokugcinwa- Ukuqinisekisa ukuba iiseli azilahlekanga, iinqanawa zokugcina kufuneka zihlale zibeke iliso kubushushu kunye namanqanaba e-nitrogen engamanzi. Iialam kufuneka zifakelwe ukwazisa abasebenzisi ngeempazamo.

ImiGaqo yoKhuseleko

I-nitrogen yolwelo iye yaba yinto eqhelekileyo kuphando lwangoku kodwa inomngcipheko wokwenzakala okumandundu ukuba isetyenziswe ngokungalunganga.

Isixhobo esifanelekileyo sokuzikhusela (PPE) kufuneka sinxitywe ukunciphisa umngcipheko weqabaka, ukutsha kunye nezinye izehlo ezimbi xa uphethe initrogen engamanzi. Nxiba

• Iiglavu zeCryogenic
• Idyasi yeLabhoratri
• Isikhuselo sobuso esinempembelelo epheleleyo esigqume intamo
• Izihlangu ezivaliweyo
• Ifaskoti yeplastiki ye-splashproof

Izikhenkcezisi zenitrogen ezimanzi kufuneka zibekwe kwiindawo ezinomoya owaneleyo ukunciphisa umngcipheko wokuphefumla – initrogen ephuncukileyo iyaba ngumphunga kwaye isuse ioksijini esemoyeni. Iivenkile ezinkulu zevolumu kufuneka zibe neenkqubo ze-alamu ephantsi ye-oxygen.

Ukusebenza ngababini xa uphethe initrogen engamanzi ilungile kwaye ukusetyenziswa kwayo ngaphandle kweeyure eziqhelekileyo zokusebenza kufuneka kungavunyelwa.

IiCryovials zokuxhasa ukuhamba kwakho komsebenzi

Inkampani yaseSuzhou Ace Biomedical inikeza ukhetho olubanzi lweemveliso ezihlangabezana neemfuno zakho zokugcinwa kwe-cryopreservation kwiintlobo ezahlukeneyo zeeseli. Ipotfoliyo ibandakanya uluhlu lwee-tubesas kunye noluhlu lwee-cryovials eziyinyumba.

Iikhriyovi zethu zezi:

• Lab Screw Cap 0.5mL 1.5mL 2.0mL Cryovial Cryogenic Vials Conical Bottom Cryotube nge Gasket

  ● 0.5ml, 1.5ml, 2.0ml iinkcukacha, kunye nesiketi okanye ngaphandle kwesiketi
  ● Uyilo oluyiconical okanye oluzimeyo, olunyumba okanye olungazaliyo zombini ziyafumaneka
  ● Iityhubhu ze-Screw cap zenziwe nge-polypropylene yodidi lwezonyango
  ● I-PP Cryotube Vials inokukhenkcezwa ngokuphindaphindiweyo kwaye inyibilikiswe
  ● Uyilo lwe-cap yangaphandle lunokunciphisa amathuba osulelo ngexesha lonyango lwesampulu.
  ● Screw cap cryogenic iityhubhu Imisonto ye-Screw jikelele ukuze isetyenziswe
  ● Iityhubhu zilungele iirotor eziqhelekileyo
  ● Iityhubhu zeCryogenic tube o-ring zilingana ne-intshi e-1 kunye ne-intshi ezi-2, i-48well, i-81well, i-96well kunye ne-100well iibhokisi zesikhenkcisi.
  ● Ikwazi ukuchaneka ukuya kwi-121°C kwaye ikhenkceza ukuya ku -86°C

  ISIQENDU INOMBOLO	IZINTO	UMQULU	ICAPUMBALA	PCS/IBHAGA	IBHAKA / ICALA
  UMTHETHO05-BL-N	PP	0.5ML	Mnyama, Mthubi, Bhulowu, Obomvu, uMsobo, Mhlophe	500	10
  UMTHETHO15-BL-N	PP	1.5ML	Mnyama, Mthubi, Bhulowu, Obomvu, uMsobo, Mhlophe	500	10
  UMTHETHO15-BL-NW	PP	1.5ML	Mnyama, Mthubi, Bhulowu, Obomvu, uMsobo, Mhlophe	500	10
  UMTHETHO20-BL-N	PP	2.0ML	Mnyama, Mthubi, Bhulowu, Obomvu, uMsobo, Mhlophe	500	10