雨生红球藻的冷冻扫描电镜制样条件初探

Preliminary study on the sample preparation conditions of Cryo-scanning electron microscopy for Haematococcus pluvialis

为了探索针对雨生红球藻的冷冻扫描电镜制样条件,以固体琼脂和液体培养的藻细胞为材料,比较了琼脂培养基直接粘台和藻液滴于预先粘台的滤纸两种粘台方式和-90℃下分别升华2 min、10 min、20 min和30min四种升华时间对成像效果的影响。结果表明:上述两种粘台方式均可获得满意的观察效果,比较而言滤纸法效果更好;-90℃升华2 min即可将藻细胞表面完全暴露,获得满意的观察效果,而长达30 min的升华时间也不会对细胞表面结构造成损伤。

To investigate the sample preparation conditions of Cryo-scanning electron microscopy （ Cryo-SEM ） for Haematococcus pluvialis, we compared two sample fixing methods and four sublimation times. The agar cultures were directly pasted on the sample stub by conductive glue, while the liquid cultures were dropped on the filter papers which had been pasted on the stub in advance. Then the samples were sublimated at -90℃ for 2 min, 10 min, 20 min and 30 min respectively. It was found that both of the two sample fixing methods could receive good Cryo-SEM images. The method using filter paper was better. For Haematococcus pluvialis cells, being sublimated at -90 ℃ for 2 min was enough. However, the prolonged sublimation time （ up to 30 min） would not cause damages to the cell surface structures.