From lipids to membranes

Published in Protocols & Methods

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Membranes are amazing things: they surround the cell, the nucleus, and many other intracellular organelles. They keep stuff inside, they keep other stuff out. There are protocols for extracting them from cells, and there are protocols for making them from scratch.

Preparing them “artificially” has the obvious advantage that you can exquisitely control their composition and properties, and a number of assays looking at membrane function and behaviour start with this step.

Short aside

For some reason these protocols always remind me of a certain German physicist who made some scientific observations that started at the kitchen sink. I can never remember her name or any of the details, but the miracle of the internet allowed me pin down her name: ‘Agnes Pockels’, and that the experiments related to detergents and surface tension. If you have access to Nature, then you can read some of the letters that she published in the 1890s:  “Surface Tension” (The text of this letter can be accessed here as well. Lord Rayleigh asked Nature to publish a letter that she had written to him and for some reason this makes me feel quite emotional. ), “Relations between the surface-tension and relative contamination of water surfaces” and “On the spreading of oil upon water“.

Back to the present, and back to the topic!

Here is a taste of some of the experimental possibilities covered in Nature Protocols so far.

Fusion of single proteoliposomes with planar, cushioned bilayers in microfluidic flow cells
Erdem Karatekin & James E Rothman

In this protocol, you make both a planar supported bilayer (SBL) and some small unilamellar vesicles (SUV).

The SBL is put together in a microfluidic flow channel…

…and docking and fusion events can be detected using conventional far-field epifluorescence or total internal reflection fluorescence microscopy.

A single vesicle-vesicle fusion assay for in vitro studies of SNAREs and accessory proteins
Jiajie Diao, Yuji Ishitsuka, Hanki Lee, Chirlmin Joo, Zengliu Su, Salman Syed, Yeon-Kyun Shin, Tae-Young Yoon & Taekjip Ha

Here, the assays are looking at the fusion of vesicles in solution with vesicles velcro’ed (biotin-neutravidin) to the bottom of a flow chamber. Both assays involve FRET pairs: either paired-dyes in reacting membranes or on the two ends of a DNA hairpin.

Generation of phospholipid vesicle-nanotube networks and transport of molecules therein
Aldo Jesorka, Natalia Stepanyants, Haijiang Zhang, Bahanur Ortmen, Bodil Hakonen & Owe Orwar

This protocol starts by taking a soybean polar extract and using it to prepare giant unilamellar vesicles (GUV) attached to multilamellar membrane reservoirs. By some very fancy manipulations, it is possible to make networks of vesicles connected by nanotubes that can be used for experiments relating to transport of molecules.

An earlier post on Membrane Proteins is related summary; and other protocols can be found by browsing our content.

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