离心法分离毕赤酵母活细胞和死细胞

本文以毕赤酵母为研究对象，探索出一种分离活酵母细胞的新方法。研究发现，通过改变淋巴细胞分离液和50%聚蔗糖溶液的比例，获得不同密度的酵母细胞分离液，进而通过离心分层的方法可使毕赤酵母活细胞主要存留于离心液的上层。当酵母细胞分离液的密度为1.1467 g/mL (27.5%淋巴细胞分离液+72.5%聚蔗糖溶液)，分离液上下层中酵母活细胞的分配比例差别达到最大，分别为94.67%(分离液上层)和5.33%(分离液下层)。毕赤酵母细胞浓度为4.35×108~1.13×109/mL时，活细胞在分离液上下两层的分配比例约为95%和5%。低毕赤酵母细胞存活率有利于离心分离。本方法可用于有效分离培养液中的毕赤酵母活细胞。

Separation of alive and dead Pichia pastoris cells by centrifugation

A new method of separating alive Pichia pastoris cells from dead Pichia pastoris cells using centrifugation approach was presented, in which alive Pichia pastoris stayed in the up layer of centrifugation solution by changing the density of centrifugation solution through adjusting the ratio of Ficoll-Hypaque solution and 50% polysucrose solution. When the centrifugation method was carried out with the Pichia pastoris cell density at 4. 35×108/mL ~ 1. 13×109/mL (27. 5% of Ficoll-Hypaque solution and 72. 5% of 50% polysucrose solution), it led to 95% and 5% alive stayed in up and down layer of centrifugation solution respectively, which was the highest proportion. And, low ratio of alive Pichia pastoris cells provided higher distribution ratio of up layer centrifugation solution. In summary, this method effectively separate alive Pichia pastoris from dead Pichia pastoris cells.