WPI MICRO-ePUMP, Instruction Manual

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18 World Precision Instruments
Manufacturing Micropipettes
Pulling suitable micropipettes is one of the biggest obstacles to taking full advantage
of the MICRO-ePUMP . Both care and steady hands are required. The volume of fluid
ejected is markedly dependent on the micropipette tip size.
When using micron-sized tips a reduction in tip-size of a few percent may give an
order of magnitude difference in the flow rate. With tip sizes less than 1 µm, pressure
ejection becomes increasingly difficult and special steps must be taken.
The most important of these steps is cleaning the glass. Small amounts of dust or
grease can easily clog micron-sized tips. Cleaning with chromic acid solutions before
pulling the electrode is commonly performed, but care must be taken to thoroughly
rinse the pipettes to remove all traces of the chromic acid, which has some affinity for
glass. Some researchers prefer hydrochloric or nitric acid.
Silanization of the glass is also recommended for small tips. With 1 µm and smaller
tips, capillary action becomes prohibitively large, and the hydrophilic surface of the
glass greatly limits the flow of fluid through the tip. Silanization decreases the surface
tension and allows the fluid to flow smoothly through the tip. For similar reasons,
we don’t recommend use of a capillary with an internal filament. Some of the many
papers on the art of silanization are listed in the bibliography.
Calibrating Volume by Measuring Droplet
For ejected volumes greater than 1 nL, visual inspection using a microscope can be
an accurate gauge of volume. A single pulse deposits a drop of fluid on the tip of the
micropipette. The volume of this drop may be calculated by measuring the radius of
the drop and assuming the drop to be spherical. Fig. 33 may be helpful in determining
the volume for a given radius. See “Appendix A: Droplet Volume” on page 24 for a
comparison of spherical and cubical volumes.
1 pl
10 pl
100 pl
1 nl
10 nl
100 nl
1 µl
10 µm 100 µm 1 mm
Volume of a Spherical Droplet
V =
π
d
3
6
V
olume
Diameter
Fig. 33—(Left) The volume of a droplet increases as the diameter increases. See “Appendix A:
Droplet Volume” on page 24.
Fig. 34—(Right) WPI’s MicroFil is helpful in backfilling glass pipette tips. The MicroFil’s tip is
thin and very flexible.