Making Patch-pipettes and Sharp Electrodes with a Programmable Puller

Summary

This video shows how to use a programmable puller to make patch pipettes and sharp electrodes for electrophysiology. The same procedure can be used to make a variety of glass tools, including injection needles.

Abstract

Glass microelectrodes (also called pipettes) have been a workhorse of electrophysiology for decades. Today, such pipettes are made from glass capillaries using a programmable puller. Such instruments heat the capillary using either a metal filament or a laser and draw out the glass using gravity, a motor or both. Pipettes for patch-clamp recording are formed using only heat and gravity, while sharp electrodes for intracellular recording use a combination of heat, gravity, and a motor. The procedure used to make intracellular recording pipettes is similar to that used to make injection needles for a variety of applications, including cRNA injection into Xenopus oocytes. In general, capillary glass <1.2 mm in diameter is used to make pipettes for patch clamp recording, while narrower glass is used for intracellular recording (outer diameter = 1.0 mm). For each tool, the puller is programmed slightly differently. This video shows how to make both kinds of recording pipettes using pre-established puller programs.

Protocol

Pulling pipettes

Using a microelectrode puller such as the Sutter P-97 Flaming/Brown, pull a set of approximately 10-20 pipettes.

1. Select your capillary glass. We use borosilicate capillaries (Sutter BF150-110-10, 1.5mm outer diameter, 1.1mm inner diameter, 10cm long). Store the glass carefully so it remains clean and dust free.
2. Design a pulling program. We use a 5 step program, with descending heat and velocity at each step, and a small pull on the final step. Sutter's pipette cookbook is an excellent reference for developing suitable programs.
3. Examine the pipette tips under a microscope to determine opening di....

Discussion

The protocol illustrated here is in daily use in electrophysiology laboratories and is also used to make injection needles for cells and animals. With a programmable puller, it is easy to make pipettes for a variety of uses.  With attention and care, the filament on your puller will last for one year or more.  Good luck with your experiments.

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Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Micropipette Puller	Instrument	Sutter Instrument Co.	P-97	Or similar instrument (e.g. Sutter P-87 or P-2000)
Glass Capillaries	Reagent	Sutter Instrument Co.	BF150-86-10	Or, similar capillary glass. To make filling the pipette easier, use a capillary with a glass filament.