PHEM Buffer recipe
PHEM buffer has been used primarily for tissues and cell cultures being processed for immunocytochemical studies. Most antigens, especially intra cellular ones, stain better using PHEM than those processed using PBS, it also has a more limited effect on biochemical reactions and enzymes. Very good preservation of cellular ultrastructure, notably microtubules, is also an advantage of PHEM use.
References:
- Schliwa, Manfred, and Jonathan Van Blerkom. “Structural interaction of cytoskeletal components.” The Journal of Cell Biology 90.1 (1981): 222-235.
- Jacqueline Montanaro: Improved ultrastructure of marine invertebrates using non-toxic buffers
Preparation recipe for 500ml of Buffer PHEM at 0.2M
Dissolve in 400 ml distilled water this reagents powder in following order:
| Reageant powder | Number C.A.S | weight gram/ Molarity | Ref. em-grade |
| HEPES | 7365-45-9 | 0.598 gr. / 5mM. | GP19 |
| EGTA | 67-42-5 | 0.380 gr. / 2mM | GP27 |
| MgSO4 , 7H2O | 10034-99-8 | 0.098 gr / 0,8mM | GP28 |
| P.I.P.E.S. | 5625-37-6 | 1.814 gr / 12mM | GP21 |
| WATER D. | Make up to 500ml final volume and adjust to the desired pH with NaOH | ||
Filter the solution to 0.22µ.
