Troubleshooting; Controlling Pipette Tip Shapes; Problem: What Glass Should I Use: Thick Or Thin; Problem: The Resistance Of My Pipettes Is To Low. How Do I Pull A - Sutter Instrument P-2000 Operation Manual

6. TROUBLESHOOTING

6.1 Controlling Pipette Tip Shapes

6.1.1 Problem: WHAT GLASS SHOULD I USE: THICK OR THIN?

In general, the thicker the wall in relation to the outside diameter of the glass the finer the
tip will be, and the thinner the wall the larger the tip will be. Thin wall glass will give the
best results in most experiments as it will have the largest pore for a given tip size. This
means it will have a lower resistance and will allow for easier injection of solutions.
However, in many cases with small cells the thin wall glass will not form tips fine enough to
obtain good penetrations. In this case, heavier wall glass must be used.

6.1.2 Problem: THE RESISTANCE OF MY PIPETTES IS TO LOW. HOW DO I PULL A

HIGHER RESISTANCE PIPETTE?

The first point to note is that there is very little correlation between tip size and electrode
resistance. Most of the resistance of a microelectrode is in the shank of the electrode behind
the tip. Electrode tips that are .1 micron in diameter can vary in resistance from 20 Mega-
ohms to 1000 Mega-ohms depending on the length of the electrode and what is used for the
filling solution. If the same solution is used then resistance may give an indication of how
well the electrode will penetrate a cell as the electrode with the higher resistance will
probably have a longer shank and a smaller cone angle at the tip. This combination will aid
in the penetration of cells where the cell is not a surface cell.

6.1.3 Problem: OK, BUT I STILL WANT A SMALLER TIP THAN I AM GETTING.

The first thing to try in most cases is to increase the HEAT value. This will generally
decrease the tip size but it will also give a longer shank. If the higher resistance is not a
problem, this is generally the best solution. Continuing to increase the HEAT is not the final
answer as too high a HEAT can lead to very high resistances. It is usually a good idea to
incrementally (and alternately) increase the HEAT and the PULL settings until the desired
pipette geometry has been achieved.

6.1.4 Problem: HOW DO I INCREASE THE SIZE OF MY PATCH-PIPETTE?

Reduce the HEAT. Try dropping the HEAT 10 units at a time to see if this will increase the
size of the tips. You may also need to try reducing the PULL and VELOCITY setting to
achieve the desired tip shape and size
6.1.5 Problem: THE TIPS OF MY PATCH-PIPETTES VARY IN SIZE FROM PULL TO
PULL.
This can happen when a pipette is formed in two or more LOOPS. If the pipette is formed in
three LOOPS in one case and then on the next pull it forms in four LOOPS the tips will not
be the same. Adding one unit in the VELOCITY value will in most cases cause the pipette to
be formed in three LOOPS or subtracting 1 unit should cause the pipette to form in 4
LOOPS. It is always good technique when a program is developed that produces a desired
pipette, to try increasing and decreasing the VELOCITY value to be sure that you are in a
stable region. The best procedure in developing a very reliable pipette program is to change
P-2000 MICROPIPETTE PULLER OPERATION MANUAL -– REV. 2.4C (20160412)
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