Amfani da taka tsantsan na ultrafiltration centrifuge tube
1) Zaɓi bututun ultrafiltration mai dacewa.Lura cewa UF membranes sun bambanta a matakin jurewarsu ga sunadarai daban-daban.Yawanci, za a iya zaɓin bututun ultrafiltration tare da yanke-yanke nauyin kwayoyin halitta na 10 kDa tare da yanke nauyin kwayoyin halitta wanda bai kamata ya fi 1/3 na nauyin kwayoyin halitta na furotin mai sha'awa ba, kamar 35 kDa.Idan nauyin kwayoyin furotin na sha'awa yana kusa da 10 kd, ana iya amfani da bututun ultrafiltration tare da yanke nauyin kwayoyin halitta na 3 KD.
(2) ultrafiltration, sabo da siya, ya bushe, tare da ultrapure ruwa ƙara kafin amfani da ruwa gaba daya wuce ta cikin membrane, kankara wanka ko firiji pre sanyaya na ƴan mintuna.Sai a zubar da ruwan, watau sinadarin protein, da nawa aka kara, duk wanda bai wuce farar layin da ke saman bututun ba.Aikin yana da haske, kuma bututun ultrafiltration yana buƙatar a sanyaya shi a kan kankara kafin ƙara maganin furotin.
3) Duka taro da tsakiyar nauyi sun kasance don isa ga ma'auni.Lura cewa saurin juyawa da haɓakawa ba su da sauri sosai, in ba haka ba kai tsaye suna lalata membrane ultrafiltration.An fara aikin ultrafiltration na centrifugal (centrifuge precooled zuwa digiri 4).Bayan RPM na centrifuges daban-daban sun canza zuwa g, ya bambanta.An daidaita haɓakar centrifuge zuwa mafi ƙanƙanta, rage matsa lamba akan membrane.Lura, tabbatar da barin centrifuge bayan centrifuge ya kai saurin inda za a nufa, ko kuma idan kuna da matsala, ba za ku iya rike shi a karon farko ba.An daidaita madaidaicin membrane zuwa sandar sandar bisa ga umarnin (al'amarin centrifuge na kusurwa shine membrane zuwa axis perpendicular).A cikin amfani mai amfani, saurin jujjuyawar gabaɗaya yana ƙasa da wancan a cikin umarnin, ta yadda za'a iya tsawaita rayuwar bututun centrifuge.
3) Duka taro da tsakiyar nauyi sun kasance don isa ga ma'auni.Lura cewa saurin juyawa da haɓakawa ba su da sauri sosai, in ba haka ba kai tsaye suna lalata membrane ultrafiltration.An fara aikin ultrafiltration na centrifugal (centrifuge precooled zuwa digiri 4).Bayan an canza RPM na centrifuges daban-daban zuwa g, ya bambanta.An daidaita haɓakar centrifuge zuwa mafi ƙanƙanta, rage matsa lamba akan membrane.Lura, tabbatar da barin centrifuge bayan centrifuge ya kai saurin inda za a nufa, ko kuma idan kuna da matsala, ba za ku iya rike shi a karon farko ba.An daidaita madaidaicin membrane zuwa sandar sandar bisa ga umarnin (al'amarin centrifuge na kusurwa shine membrane zuwa axis perpendicular).A cikin amfani mai amfani, saurin jujjuyawar gabaɗaya yana ƙasa da wancan a cikin umarnin, don a iya tsawaita rayuwar bututun centrifuge.
(4) Lokacin da aka mayar da hankali ga sauran 1ml, ɗauki 50ul na buffer bayani, ƙara 10ul na gudana ta hanyar kuma duba idan akwai launin shudi, a matsayin hukunci ko bututun ultrafiltration ya rasa furotin.Idan bututun ya yoyo, sake zuba saman saman sannan a zuga cikin sabon bututu don fara ultrafiltration.Don tantance daidai idan ba a rasa bututun ba, centrifuge na 10min tare da 5mgml na BSA kafin tafiya, yana gudana akan manne furotin, ko ɗanyen ɗanyen Bradford, kuma ci gaba da ƙara sauran bayanan furotin mai ƙarfi (wanda ke aiki akan kankara kuma yana hana sunadarai daga dumama) har sai An ƙara duk abin da aka tattara.Kula da lokacin centrifugation idan hazo na sunadaran suna faruwa, yana haifar da rufewar bututu.Idan hazo ya faru, ƙayyade ko takamaiman dalilin hazo shine ƙwayar furotin da ya wuce kima ko buffer da bai dace ba;Za a iya magance na farko ta hanyar ultrafiltration na tubes ultrafiltration da yawa a lokaci guda, rage yawan taro, da kuma na karshen ta hanyar musayar hanyoyin buffer daban-daban har sai babu hazo na furotin.
(4) Lokacin da aka mayar da hankali ga sauran 1ml, ɗauki 50ul na buffer bayani, ƙara 10ul na gudana ta hanyar kuma duba idan akwai launin shudi, a matsayin hukunci ko bututun ultrafiltration ya rasa furotin.Idan bututun ya yoyo, sake zuba saman saman sannan a zuga cikin sabon bututu don fara ultrafiltration.Don tantance daidai idan ba a rasa bututun ba, centrifuge na 10min tare da 5mgml na BSA kafin tafiya, yana gudana akan manne furotin, ko ɗanyen ɗanyen Bradford, kuma ci gaba da ƙara sauran bayanan furotin mai ƙarfi (wanda ke aiki akan kankara kuma yana hana sunadarai daga dumama) har sai An ƙara duk abin da aka tattara.Kula da lokacin centrifugation idan hazo na sunadaran suna faruwa, yana haifar da rufewar bututu.Idan hazo ya faru, ƙayyade ko takamaiman dalilin hazo shine ƙwayar furotin da ya wuce kima ko buffer mara dacewa;Za a iya magance na farko ta hanyar ultrafiltration na tubes ultrafiltration da yawa a lokaci guda, rage yawan taro, da kuma na karshen ta hanyar musayar hanyoyin buffer daban-daban har sai babu hazo na furotin.
(5) Ana amfani da ƴan matakai na farko don tattara furotin, kuma idan za'a canza buffer, a hankali ƙara sabon buffer (ultrafiltration ta hanyar 0.22um ultrafiltration membrane) zuwa kusan 1ml na jimlar furotin, kuma sake mayar da hankali zuwa kusan 1ml na uku. lokuta a jere, tare da ƙarar ƙarshe ta ƙarshe dangane da adadin furotin da ake so, gabaɗaya bai wuce 500ul ba, amma kuma zuwa cikin 200ul.Cimma 1000 × ko fiye a lokuta uku, da gaske gwargwadon canjin buffer, kamar yadda aka ƙididdige ta aƙalla 10 × ko makamancin haɓaka girma kowane lokaci.
(5) Ana amfani da ƴan matakai na farko don tattara furotin, kuma idan za'a canza buffer, a hankali ƙara sabon buffer (ultrafiltration ta hanyar 0.22um ultrafiltration membrane) zuwa kusan 1ml na jimlar furotin, kuma sake mayar da hankali zuwa kusan 1ml na uku. lokuta a jere, tare da ƙarar ƙarar ƙarshe ta ƙarshe dangane da adadin furotin da ake so, gabaɗaya bai wuce 500ul ba, amma kuma zuwa cikin 200ul.Cimma 1000 × ko fiye a lokuta uku, da gaske gwargwadon canjin buffer, kamar yadda aka ƙididdige ta aƙalla 10 × ko makamancin haɓaka girma kowane lokaci.
Lokacin aikawa: Nuwamba-09-2022