WPI PicoPump PV830, Инструкция по эксплуатации

Страница: 15 / 28

PV830 Pneumatic PicoPump
World Precision Instruments 11
Techniques In Microinjection
One of the more demanding tasks for which the PicoPump has been designed is the
microinjection of fluid into cells. Difficulties encountered will not usually involve the
mechanics of the PicoPump, but rather the manufacture of suitable micropipettes.
Both care and steady hands are needed in making pipettes to take full advantage of
the features of the PicoPump. Here we list several important things to keep in mind
when working with the PicoPump.
Setting the Hold Pressure
The Hold pressure is used to counterbalance the capillary action of the fluid
backfilling into the pipettes. When an empty pipette is inserted into the fluid, a
meniscus can be seen to rise up from the capillary tip.
Hold pressure may be set
by adjusting the regulator setting (with the Hold/vent switch in the hold position)
until the meniscus stops at the desired position. In many applications, colored dye
or fluorescent dye is dissolved in the injection fluid. The capillary effect can be seen
with the color change at the tip of the pipette. When the fluid flows into the pipette,
the color of the tip becomes lighter. If the hold pressure is higher than the capillary
pressure, the fluid oozes out of the pipette. The solution around the pipette will be
colored. Adjusting the
Hold pressure prevents this from happening.
Understanding how capillary action causes the backfilling of the pipette helps you
to correctly use the
Hold pressure. The backfilling flow rate is determined by the
pressure of capillary action and the tip size. Since the tip size is often determined
by the requirement of the application, controlling the pressure of capillary action
becomes the main option to eliminate backfilling.
The pressure of capillary action
is determined by the inner diameter of the glass capillary where the meniscus
of air/liquid interface is located. It has nothing to do with the pipette tip size, a
common misunderstanding.
If we assume the pipette tip is near a cylindrical shape, the pressure of capillary
action can be described by the LaPlace equation:
P=4
γ
cos
θ
/d
where
γ
is the surface tension,
θ
is the contact angle between the water and glass,
and d is the inner diameter of the capillary where the meniscus is located. In most
cases, we can assume the contact angle for glass and water is zero (unless the
glass surface is treated). From this equation we can see that the capillary pressure
can vary a thousand times when the meniscus is moved from a 0.5µm ID tip to
the 0.5mm shank. The pressure at 0.5µm tip is about 80 PSI (in aqueous solution)
while at the shank will be only 0.08 PSI. Using one regulator to counterbalance the
pressure in such a large dynamic range is not practical. The hold regulator in this
instrument is optimized to work in the 0.2–10 PSI range. 10 PSI can counterbalance
a meniscus at the section of tip where the inner diameter is 4µm. In practice, this is
the highest pressure ever needed. On the lower pressure end, it becomes difficult to
exactly counterbalance the capillary pressure when the meniscus is at the shank of