Note: Requires a pressure bomb attached to a tank of compressed helium

1. Analytical columns with integrated emitter tips are manufactured using 360-µm o.d. × 75 µm i.d. fused silica capillary tubing.
2. To form the emitter tip, use a butane lighter to remove ~3 cm of polyimide coating 5 cm from one end of the fused silica.
3. Clean this area with methanol to remove any remaining charred polyimide coating before inserting the silica into the puller.
4. Using the manufacturer’s recommended settings for the laser puller, pull a 15-µm tip, by avoiding the polyimide coating.
5. Inspect the tip under a microscope; if the tip is closed, etch with HF to create an opening. In the fume hood, transfer 50 µl of HF to a microcentrifuge tube and fill another tube with 0.5 M ammonium formate. Dip the electrospray tip in HF for 1–2 min. Then thoroughly rinse the tip with water.

! CAUTION Take appropriate safety precautions when using HF. Wear proper protective equipment, and perform the work in a properly vented fume hood. Wear safety goggles when you are working with the pressure bombs.

6. Prepare a slurry of 5 µm BEH packing material by transferring the packing material to a glass vial containing a mini magnetic stir bar. Add ACN to the vial. As the 3.5-µm BEH packing material is added only to keep the smaller 1.7-µm material in the column, a very dilute slurry is recommended (0.1 mg, 3.5-µm packing material, 1 ml ACN). Place this into the pressure bomb.
7. Insert the analytical column, with pulled tip pointing upward, through the high-pressure bomb. Be careful when you are handling the column, as the tip is fragile and easily damaged. Slowly adjust the position of the column until is 1–2 mm above the bottom of the tube.
8. Set the pressure regulator on the helium tank to 1,000 psi.
9. Increase the pressure in the pressure bomb by slowly turning the valve to the open position.
10. Increase the pressure in the pressure bomb by slowly turning the valve to the open position.
11. Fill ~5 mm of the tip with 5-µm BEH slurry. If desired, a lower pressure of helium (200–300 psi) can be used for this step.
12. Prepare a slurry of 1.7-µm BEH packing material by transferring the packing material (~0.5 mg) to a glass vial containing a mini magnetic stir bar. Add 1 ml of chloroform to the vial.
13. Fill the analytical column with 1.7-µm BEH packing material to a length of ~30 cm.
14. Release the pressure by turning the valve to the closed position slowly to avoid column unpacking.