64
•
Guide to Electrophysiological Recording
The suction port on the HL-U series holders provided with the
MultiClamp 700B should therefore be left open.
•
Using filling solutions with low pH, or adding small concentrations of
polyvalent cations like Th
4+
, may reduce the size of the tip potential and
therefore the magnitude of any changes (Purves, 1981).
Filling Solutions
The best filling solution to use depends on the preparation under investigation and
the experience of the investigator. Although KCl gives one of the lowest tip
resistances for a given tip diameter, a KCl-filled electrode is not necessarily the
fastest to settle after a current pulse. K-citrate is sometimes faster.
It is important to be aware that during current-passing, large amounts of ions from
inside the microelectrode can be ionophoresed into the cell. For example, if current
is passed by the flow of ion species A from the microelectrode into the cell, then
after 50 seconds of current at 1 nA (or 1 s of current at 50 nA) the change in
concentration of A inside a cell 100
µ
m in diameter is 1 mM. If A is an
impermeant ion, the cell may swell due to the inflow of water to balance the
osmotic pressure. The injection of a permeant ion, such as chloride, can
significantly alter the equilibrium potential for that ion.
Impaling Cells
Start with the MultiClamp 700B in IC mode (I-Clamp). Fill a microelectrode with
internal solution and secure it firmly in the pipette holder. Be sure to support the
headstage with your other hand so that the micromanipulator will not have to
absorb your force. Advance the electrode until its tip enters the bath. Press the
Pipette Offset button to null the offset.
Note:
Check the stability of the bath electrode and microelectrode. Drifts in
Primary output: Membrane Potential indicates that the electrode wires
probably need to be rechlorided. Also check for a changing tip potential by
passing a steady current, as described above.
MultiClamp 700B Theory and Operation, Copyright 2005 Axon Instruments / Molecular Devices
