Hi, I am Jan Dolzer,
Product Manager for the Sutter Instrument P-2000 Laser-based Micropipette Puller.
What I am going to show you today is something you can think of as the "Birth" of a glass pipette.
We have taken a very close look at the events that take place as a quartz glass capillary
is heated and then pulled into a patch pipette. And then we have captured this exciting process on film.
By looking at these events and understanding them in detail
we will go a long way towards becoming proficient pipette cooks, which enables us
to create our own pipettes and to shape them the way we actually need them.
Going forward, we will produce a series of tutorial videos that will show and deal with
what happens if you change individual parameters of the pull program.
The title of the series is "Program 71", simply because that is the program number
that we used during the production of the video footage.
Today, we will set the baseline: That is, we will show what happens
if you pull glass of a particular specification with a sample program, that is Program #1.
What you will get is a patch pipette. The tutorial videos will then use that as a
starting point and show what happens if you change the parameters one at a time.
We have also produced an introductory video that explains the point of view of the camera
in those close-up shots of the pipettes in the process of being pulled.
So if you find ourself saying: "I have no clue what I am looking at here",
Go to our website
or take a look at our YouTube channel and find the video called: "Tutorial Intro".
That will make things much clearer.
Your puller comes pre-programmed with a two-line sample program in position 1, which can be
used as a good starting point for pulling patch pipettes from 1-mm quartz glass.
Should the sample program have been overwritten, you can copy the parameters
from the subtitle here.
We are viewing the pipette from the back of the puller, above the axis of the laser beam.
As the quartz capillary is heated, it starts glowing white, and only then it becomes pliable.
The first pull draws the capillary to an hourglass. The second pull separates the glass after
a very short heating time.
All this happened too fast for most of us to follow. So let's take a closer look at
a few details of the pull. During the initial heating phase, dust particles light up first.
Even though the capillary in this video was cleaned, small dust particles cannot be completely
avoided with reasonable effort. In the video, they help us judge movement of the glass.
In this slow motion clip individual frames are 16.7 ms apart. When the glass is glowing white,
the weak pull starts drawing the halves of the capillary apart, and the hourglass begins to form.
Note how the narrow section of the hourglass has already darkened and hardened,
while material is being pulled from the shoulders, which are still brightly lit.
The final hourglass of the first pull measures 6.5 mm shoulder to shoulder.
In the second pull, the pipettes are separated after only a short,
flashing glow in the center of the hourglass.
Here is a shot of another pull with the same parameters in brighter illumination.
The frame rate is 100 Hz this time. Individual frames can be visually separated now.
While the glowing of the glass cannot be seen as clearly, the glass itself is better to see under the
brighter illumination. But even with the higher frame rate, the second pull only covers 3 - 4 frames.
Let's focus on the formation of the hourglass during the first pull, and on how the hourglass
relates to the taper of the final pipettes. Here is the original video again.
After the hourglass has formed, an image of the final pipette is superimposed and aligned at the shoulders.
Note how the pipette tip extends more than 1 mm into the other side.
A close-up shows a smooth taper with an angle of about 1.5 degrees all the way to the tip.
There is only a hint of a shoulder from the second pull, just above the marker in the center of the frame.
At higher magnification under the microscope, the very tip of the pipette becomes visible.
The diameter of the pipette opening is of the order of one micron - more easily resolved
looking down the actual microscope than captured on camera.
Now let's take a look at the right side and see whether the pull has produced symmetrical pipettes.
Again, the image of the final pipette is superimposed and aligned on the video.
The tip protrudes about 1 mm to the left of the center, just as the other side did.
And the pipette is virtually a mirror image of the left half.
Now that we have learned how to pull a patch pipette with a sample program, go out and do it!
Keep in mind that Program #1 only works with the glass that we have used today.
If you use a different type of glass, your results may and will vary.
And you may need to do some fine-tuning, even if you use this same type of glass.
Enjoy. Have fun!
Stay tuned for a series of tutorial videos that show the effect of changes in individual parameters.
Check out our website or our YouTube channel for a growing collection of tutorial videos.
If you need any tech support go to www.sutter.com
or give us a call at +1 415 883 0128.