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the pull takes place in less than eight seconds. If the pull occurs in less than three seconds,
decrease the HEAT until the pull takes place in 4 to 8 seconds. For the best micropipette
reproducibility with the finest tips, you should select a HEAT value that melts the glass in 5
to 6 seconds. For microinjection pipettes, select a HEAT value that melts the glass in about 7
seconds or longer.
4.8.2.2 PULL Strength
Low values of PULL strength in the range of 50 to 75 will give larger tips appropriate for
injection needles, while 80 to 150 give smaller tips appropriate for sharp microelectrodes. The
PULL strength can be set to any value desired with no danger of damaging the instrument.
4.8.2.3 VELOCITY (Trip Point)
The VELOCITY value determines the point at which the heat is turned off. VELOCITY
reflects the speed at which the two puller bars are moving during the weak pull. The lower
the VELOCITY value, the slower the speed of the bars when the trip point occurs. Although
VELOCITY can safely be set to any value from 1-255, all values over a maximal trip point
(usually about 150) will produce equivalent results. As the pull progresses, the speed of the
puller bars, as measured by the velocity transducer, reaches a point where further increases
in the VELOCITY trip point will not change the time point at which the heat is turned off.
VELOCITY is typically set between 80 to 90 for microelectrodes or 50 to 80 for microinjection
pipettes.
In a multiple cycle program, it is possible for the glass to separate before the trip velocity is
attained. Thus, the glass is subjected to heating as it separates. Such an occurrence can lead
to difficulties in forming tips as well as lack of reproducibility. If you are using a one-line,
looping program, try decreasing the VELOCITY a few units at a time. If your program is a
multi-line program, decrease the VELOCITY in the next to last line of the program.
Decreasing the VELOCITY will increase the amount of glass left in the last cycle of the
program, thus allowing the glass to attain the trip velocity before separating.
4.8.2.4 TIME Mode (Cooling)
The TIME parameter controls the length of time the cooling air is active (one unit of TIME is
equivalent to 0.5 ms). In order to produce effective cooling, the air must be supplied to the
filament and glass during the time the tip is being formed. When pulling sharp electrodes,
the hard pull lasts several tens of milliseconds. Because of this fact, increasing cooling TIME
values beyond a certain range (typically 200 to 250) will have no effect. Values of TIME
under 200 will cool the glass less as the tip is being formed and lead to a longer taper.
However, once TIME values become too short (values in the range 110 to 130) cooling
becomes ineffective. The glass will not form a tip and instead forms a wispy fiber. The very
finest tips for a given PULL and HEAT will be formed at an air setting of about 5 units
higher than the lowest TIME value that forms a tip. Because of this quite narrow working
range of usable TIME values for making micropipettes, it is not recommended to vary
cooling, and therefore electrode tip length, by adjusting TIME. Adjusting the cooling air
pressure and/or switching to the Delay mode of active cooling are both more effective means
of controlling tip length (see below).
P-1000 FLAMING/BROWN MICROPIPETTE PULLER SYSTEM OPERATION MANUAL – REV. 3.02 (20161118)
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