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Materiali in metode
3.1. Kultivacije v stresalni bučki
Pre-kulture suspenzijskih celic MDCK.SUS2 so bile gojene v shaker flasks (125 mL polikarbonatne Erlenmeyerjeve bučke, #431143, Corning®, New York City, NY, ZDA) s 50 mL delovne prostornine (wv) , v inkubatorju Multitron Pro (Infors HT, Bottmingen, Švica) pri 37 °C in atmosferi 5 % CO2 s frekvenco stresanja 180 vrt/min. Celice so bile pasirane vsake 3–4 dni z gostoto sejanja 0,5–0,8 × 106 celic/mL.
IAV A/Puerto Rico/8/34 (H1N1) semenski virus je bil uporabljen za okužbo, ustvarjen v adherentnih celicah MDCK (ECACC # 84121903). Semenski virus je imel titer 1,1 × 109 TCID50/mL.
Celice so bile gojene v kemično definiranem mediju brez beljakovin in živalskih komponent, Smif8 (Smif8 PGD 2×, dopolnjen s 5 mM glutamina in 8 mM piruvata), posebej razvit za gojenje suspenzijskih celic MDCK [95].
Poskusi rasti in okužbe za validacijo modela so bili izvedeni z uporabo 500 mL stresalnih bučk (#4113-0500, Nalgene™, Thermo Scientific, Waltham, MA, ZDA) z začetnim volumnom gojenja 200 ml pri 150 vrt./min. Izvedeni sta bili dve kultivaciji, ena lažna okužba (Kultivacija 1), kjer so celice rasle približno 200 ur, in ena okužba (Kultivacija 2), kjer so bile celice okužene z IAV približno 49 ur po inokulaciji. Za dosego sinhrone okužbe celične populacije je bil uporabljen moi = 10. Zaradi razmeroma nizke koncentracije celic v času okužbe (2,1 × 106 celic/mL) in uporabljenega visokega moi ni bila potrebna niti zamenjava medija niti dodajanje tripsina (za aktivacijo virusa). V obeh gojenjih je bila pred vzorčenjem dodana sterilna Milli-Q voda, da se kompenzira izhlapevanje vode (1–2 mL/dan), saj je bil poskus izveden v nehidriranem inkubatorju.
Vir iz: https:/ /www.mdpi.com/2218-1989/12/3/239/htm
The FAI climbed 5.9 percent year-on-year in the first 11 months of 2018, quickening from the 5.7-percent growth in Jan-Oct, the National Bureau of Statistics (NBS) said Friday in an online statement.
The key indicator of investment, dubbed a major growth driver, hit the bottom in August and has since started to rebound steadily.
In the face of emerging economic challenges home and abroad, China has stepped up efforts to stabilize investment, in particular rolling out measures to motivate private investors and channel funds into infrastructure.
Friday's data showed private investment, accounting for more than 60 percent of the total FAI, expanded by a brisk 8.7 percent.
NBS spokesperson Mao Shengyong said funds into weak economic links registered rapid increases as investment in environmental protection and agriculture jumped 42 percent and 12.5 percent respectively, much faster than the average.
In breakdown, investment in high-tech and equipment manufacturing remained vigorous with 16.1-percent and 11.6-percent increases respectively in the first 11 months. Infrastructure investment gained 3.7 percent, staying flat. Investment in property development rose 9.7 percent, also unchanged.