PC-505B Manual, Rev. 201013
Warner
Instruments
A Harvard Apparatus Company
27
Preliminary
Mount the electrode
Position the Ag-AgCl reference electrode or KCl-agar salt bridge into the recording
chamber and connect to the ground pin on the headstage case.
NOTE
: If using a Ag/AgCl wire or pellet reference electrode, make sure that your Ag
wire/pellet is fully chlorided. Any bare Ag
2+
that contacts your solutions will introduce an
undesired junction potential. This is also true for the Ag/AgCl wire in the recording electrode
in your pipette holder.
Insert a properly pulled and filled micropipette into the electrode holder and gently but
firmly insert the prepared electrode holder into the headstage.
NOTE
: Stray capacitance can be reduced by Sylgard coating your pulled pipettes. (But not
the tip!) For a detailed discussion on this and other noise sources, see Chapter 1 in Patch-
Clamp Applications and Protocols, by Levis and Rae (1995) Humana Press.
Submerge the electrode tip
The following operations need to be performed quickly since any delay will foul the tip
and inhibit seal formation. Also, it is advisable to locate the cell you want to patch before
lowering the tip into solution.
Just before lowering the electrode pipette tip into the bath, carefully aspirate the surface
of the bath with a small tube connected to a vacuum line, or quickly wipe the surface with
a small clean piece of lens tissue. This will remove any debris on the surface.
Next, apply continuous, gentle, positive air pressure (about 1-2 kPa or 10-20 cm of
water) to the side port of the electrode holder to prevent any remaining debris from fouling
the pipette tip during insertion and lower the tip into the bath.
Keep the pipette tip in the bath and maintain the air pressure in the electrode holder as
you proceed with the steps below.
Zero the pipette offset
On entering the bath, the junction potential between the pipette and reference
electrodes will induce an appreciable
current through the pipette. This is
indicated by the panel meter I
m
reading in
pA, and on your data acquisition system
or oscilloscope from the I
m
output
terminal.
Note
: If the data trace is not a straight line
but instead looks like the one to the right,
then push (once) the
SPEED TEST
button on
the rear of the instrument.
