Mazano ekutanga eCrispreservation yetsika tsika

Mazano ekutanga eCrispreservation yetsika tsika

Masero anoenderera mberi metabolism yavo panguva yakajairwa mabasa ekuita, ayo anoda kutora chikamu kwemari dzakasiyana siyana. Kana tembiricha inodonha pazasi -70 ° C, dziripo dzinomira kushanda. Naizvozvo, munharaunda ine zvakanyanyisa kudziya kwakadzika, masero anogona kumisa zviitiko zvavo zvevanyoreri uye isa durmant mamiriro anobvumira kuchengetedza kwenguva refu.

1. Kuedza

Base zvinyorwa:

Chinyorwa cheChikamu Chete

Serum (kujairika FBS / NBS)

Ultrapure Workbench

Stile Dimethyl Sulfoxide (DMSO)

Sterile PBS Phosphathe Buffer Munyiti Solution

Pipetting pfuti

Magetsi pipetting pfuti

Kugadzirira kwekuedzwa

A) Kugadzirira kwemhinduro yeKusika:

General masero: 55% sosi yemasikati + 40% Bovine Serum (FBS / NBS) + 5% DMSO

Masero akakosha: 90% Bovine Serum (FBS / NBS) + 10% DMSO

Aliquot iyo yakagadzirirwa cryopreserresvation mhinduro mune 15 ml centrifige tube [dendere] uye chengetedza pa 4 ° C kuti ushandiswe gare gare.

(b) Kugadzirira kwemasero kuti kuve nechando:

Usati wasimudza masero, sarudza masero ane chimiro chakanaka chekukura uye mune iyo logarithmic yekurayira chikamu uye kutsiva neyemagetsi nyowani gumi nemaviri isati yarira kuchengetedza cell mamiriro.

2. Kuedza maitiro

1. Cherekedza dentity yemasero kuti afukidzwe, idzo inenge 80% ~ 90%. Shandisa pipette pfuti kune espating the yekare svikiro, wedzera zvisina kufanira pbs kuti ugeze masero 1-2 nguva, uye bvisa svikiro rakasara munzvimbo yemhando yetsika.

2. Wedzera huwandu hwakakodzera hwe trypsin kana yekugaya muto kubvumira iyo trypsin kuti ipinde masero, uye vaise mucubator yekugaya. Cherekedza mamiriro emasero ari pasi peMicroscope: Iyo cytoplasm iri kudzoreredza uye masero haasisiri kubatana mumashiti. Panguva ino, wedzera mhinduro yekumisa kumisa maitiro ekugaya.

3. Nyoro bubble masero ane tip kuti aumbe cell kumiswa uye centrifige the cell kumiswa pa1000 rpm kwemaminitsi 3-5.

Rasa iyo Supernatant, wedzera huwandu hwakakodzera hwemhinduro dzinotonhora, uye zvinyoronyoro pipette kuita kuti masero aiverenge zvakaenzana. Gadzirisa iyo cell density neCryopreserrdved Svikiro kuti iite yekupedzisira dentity 5 × 106 / ml ~ 1 × 107 / ml.

4. Shandisa pipette pfuti kupatsanura iyo cryogenic machubhu zvinoenderana neyekutarisirwa kugona, uye shandisa otomatiki capping muchina kuti ubate uye chisimbiso.

5. Iyo yakajairika Cryopreservation chirongwa chinotonhorera che -1 ° C to -2 ° C / Min, uye masero ekucheka anogona kuve akaomeswa zvishoma maererano nematanho ekutevera: 4 ° C (20min) → - 20 ° C (30min) → -80 ° C ° C (Overnight)

Inogona zvakare kuchengetwa zvakananga mufreezer pa -80 ° C usiku uye wozoendeswa kune mvura nitrogen yekuchengetedza.

Yongyue Medical ndeye yakakwira-tech bhizinesi inokosha muR & D, kugadzirwa uye kutengesa kwe biomeidical zvinoparadza. Pipette matipi, pombi, pclt plate, zvigadzirwa zvetsika zvigadzirwa uye zvimwe zvigadzirwa zvekushandisa zvinoparadza. Yongyue Medical Yakakodzera Kuvimba Kwako uye Chisarudzo! Tikugashirei kubvunza!