PIPES Buffer

1,4 Piperazine bis (2‐ethanosulfonic acid) Useful pH range: 6.4 ‐ 7.2

Reference:

Good, N.E., et al., J. Biochem. 5, 467 ‐ 1966
Baur, P.S., et al., J. of Microscopy, 109, 315 ‐ 1977

Introduction

PIPES is a hydrogen‐ion organic‐based buffer, used in the preparation of a variety of mammalian tissues for EM. It produces excellent results in the preservation of ultrastructural details, especially when long fixation is required. This buffer does not appear to contribute extraneous anions or cations to the tissue, thus permitting elemental determinations by means of energy dispersive X‐Ray analysis.

Baur, et al., have demonstrated that the ultrastructural aspects of various tissues (human skin, human hypertrophic scars, mollusc neuronal tissue, oyster gill, hamster ovary, cell in culture, rat skin, lung and skeletal muscle, and canine cardiac muscle) fixed with NaOH‐PIPES‐bufferedglutaraldehyde solution were of superior quality with more detail between organelles. The cytoplasm is more homogeneous in consistency, mitochondria is very electron‐dense, microfilaments and microtubules are more commonly observed in PIPES tissue and to a lesser degree in phosphate‐treated cells.

The following procedure is recommended

Fixation for 24 to 48 hours in a 3% glutaraldehyde, 0.1M PIPES solution buffered with NaOH to pH 7.6.

Rinse the PIPES buffer for 20 minutes, using three changes.

Post fixation is with 1% 0s04 with 0.1M PIPES buffer for 1 ‐ 2 hours.

Other Useful Chemicals for Electron Microscopy Sciences

PIPES ‐ C₈H₁₈N₂0₆S₂ M.W. 353.3 ‐ C022
8% solution ‐ Glutaraldehyde EM grade ‐ 10mL ampoules
10% solution ‐ Glutaraldehyde EM grade ‐ 10mL ampoules
25% solution ‐ Glutaraldehyde EM grade ‐ 10mL ampoules ‐ C001
50% solution ‐ Glutaraldehyde EM grade ‐ 10mL ampoules ‐ C003
Osmium tetroxide, 0s0₄ ‐ one gram ampoule ‐? C010
Osmium tetroxide ‐ 4% aqueous solution ‐ 5mL ampoules
Osmium tetroxide ‐ 4% aqueous solution ‐ 10mL ampoules ‐? C011
Sodium hydroxide pellets ‐ 100 grams ‐ C201