Multichannel Systems HiClamp Manual - Manual (Page 77)

Appendix
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11.3 About Preparation of Xenopus Oocytes
The materials and methods described in the following are kindly provided by the Bayer AG,
Leverkusen. The described procedures have been optimized over several years for best
performance of the oocytes and highest throughput. We recommend that you follow the
instructions for obtaining best results. Refer to standard protocols if you need more detailed
information on the subject.
Warning: Only qualified personnel should be allowed to perform laboratory work. Always
make sure you fulfill the requirements of local regulations and laws. Work according to good
laboratory practice to obtain best results and to minimize risks.
11.3.1 Materials
Recommended products are listed under “Sources of Supply” in chapter “Ordering
Information”.
Biological materials
Female frogs of Xenopus laevis.
Technical Equipment
 Shaker for the tubes (during defolliculation)
 Stereo microscope for quality control of oocytes
 8- / 12-channel pipette or Tecan Columbus Microplate Washer for filling the well plates
and washing the oocytes
 96 well plates with conical bottom
It is very important that the well plates are produced carefully and have minimum variations.
Do not use coated plates, because oocytes will not adhere to the well bottom of coated plates!
Check each single plate before use. The plate should be even and it should not be distorted
in any way.
Note: If you use warped plates, you will encounter problems during injection. Check each plate
carefully before use.
Oocyte sieve for grading oocytes by size: Remove the bottom of a 50 ml Falcon tube.
Place a polyimide mesh with an 800 μm grid over the cut end and fix it with glue.
Oocyte transfer pipette: Cut and fire polished glass Pasteur pipette with an opening of ca.1.5 mm.
The cut end should be straight, that is, with a fixed diameter of about 1.5 mm, for at least 10 mm
so that a number of oocytes can line up inside the pipette as pearls on a string. See also chapter
“Plating Oocytes”.
 Rubber ball for oocyte transfer pipette
 Glass tool for moving oocytes: Glass Pasteur pipette, sealed over a Bunsen burner, and melted
into a golf-club like shape.
 Petri or cell culture dishes, 100 mm
 Petri dishes, 60 mm
 Beaker, 100 ml
 Razor blade
 Forceps
 Parafilm and standard laboratory equipment