A practical guide to the use of monolayer purification and affinity grids

Methods Enzymol. 2010;481:83-107. doi: 10.1016/S0076-6879(10)81004-3.

Abstract

Lipid monolayers have traditionally been used in electron microscopy (EM) to form two-dimensional (2D) protein arrays for structural studies by electron crystallography. More recently, monolayers containing Nickel-nitrilotriacetic acid (Ni-NTA) lipids have been used to combine the purification and preparation of single-particle EM specimens of His-tagged proteins into a single, convenient step. This monolayer purification technique was further simplified by introducing the Affinity Grid, an EM grid that features a predeposited Ni-NTA lipid-containing monolayer. In this contribution, we provide a detailed description for the use of monolayer purification and Affinity Grids, discuss their advantages and limitations, and present examples to illustrate specific applications of the methods.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Cryoelectron Microscopy / methods*
  • Crystallography / methods*
  • Lipids / chemistry*
  • Nickel / chemistry
  • Nitrilotriacetic Acid / chemistry
  • Proteins / chemistry
  • Proteins / ultrastructure

Substances

  • Lipids
  • Proteins
  • Nickel
  • Nitrilotriacetic Acid