Environmental Engineering Laboratory

By: Ngoc Thuan Le

Preparation for SEM

Apparatus

The microbial structure of granules was examined with a scanning electron microscope
(SEM; Jeol JSM-6380LV(Japan)).

Procedure

1. The granule samples were gently washed with phosphate buffer solution and allowed
to settle naturally.
2. Granules then were fixed with 4% paraformaldehyde and left for 4 h.
3. The fixed granules were dehydrated by successive passages through 40, 60, 80 and
100% ethanol.
4. Then they were dried either by Freeze Dryer or Critical Point Dryer (model E3000).
5. And finally observed by SEM.

Reagents
- Phosphate buffer solution
o 0.2 M sodium phosphate buffer (1000 ㎖)
o NaH2PO4.2H2O (MW = 156.01) 31.202 g
o DW 800 ㎖
o Adjust pH 7.3 with NaOH pellet.
o Add DW to 1000 ㎖.
- 4% paraformaldehyde: 10.8ml paraformaldehyde (37%), bring to 100ml with
DW.
- Ethanol

Reference:
“The effects of shear force on the formation, structure and metabolism of aerobic granules”
J.-H. Tay · Q.-S. Liu · Y. Liu Appl Microbiol Biotechnol (2001) 57:227–233 DOI
10.1007/s002530100766