Yeast Cell Micromanipulation
Advantages:
- Perfectly flat and light-transmitting
- Minimal time and effort to construct
- Non-breakable support stem
- Minimal shadow in field of view
Materials Needed:
- 3/4 inch fiber-optic strand with polished ends (for source, see below)
- closed end capillary 1 mm diam.
- aluminum rod 2mm x 5 inches
- superglue
- tape
- 2 microscope slides 1 inch x 3 inches
- small, steady flame (for example, a well-shielded pilot from Touch-o-matic burner)
- small, non-serrated forceps (Millipore is preferable)
- toothpick with Parafilm tip
- Might need hacksaw or bolt cutter
Protocol:
- Make a right angle bend 3/8 inch from closed end of capillary tube. With the capillary tube secured to a horizontal support, heat the bend point with the tip of the flame. As the glass softens, the tip will fall by gravity straight down into the flame. Remove the flame.
- This step is optional, but will reduce shadow. Stretch out the short closed-end segment to make it thinner. Direct the flame to the middle (not at the bend!) of the short segment. Grasp the closed-end tip with forceps and pull it gently and deliberately. Keep your line straight to preserve the 90 degree angle. Cut off excess glass to leave a suitable length of 5/16 to 1/2 inch.
- Place the capillary tube parallel to the aluminum rod so that the bent end extends 1 inch beyond the rod. Tape them together. One short piece of tape is enough.
- Check the length of your two-part needle support. Measure the distance between the viewing site of your microscope and the micromanipulator holder. Shorten your rod if necessary using a hacksaw or bolt cutter.
- Attach a fiber optic strand to the short bent segment of the capillary tube using superglue.
- Working under good light, place a fiber optic strand on a glass slide. Strands are handled conveniently with forceps or a Parafilm-tipped toothpick. If convenient, check under low power (40-50X) magnification to be sure the ends are squared off (not broken).
- Move the fiber so that one end extends into open air beyond the edge of the slide, or even slants down off the slide. The opposite end will be the working tip.
- Apply a drop of super-glue to the second slide, and moisten the capillary bent segment in it. Bring the glue-moistened segment parallel to the optic fiber so that the bend meets it about 1/8 inch from the non-working end (the end off the slide). On contact, the fiber will attach by surface tension. Caution: Avoid touching the slide with glue - the fiber may stick to the slide! And avoid getting glue on the bottom end of the fiber - light transmission will be impaired (but the needle will still function).
- Adjust the angle and height of the fiber needle. While the glue is still moist, you can make small adjustments with a toothpick or your fingertip. Try to perfect the right angle. The distance from the bend to working tip should be a little over 1/2 inch.
Sources:
- Fiber optic needle source:
Cora Styles Lab Supplies, LLC
PO Box 431
Talent, OR 97540
Phone: 541-535-6775
Fax: 678-693-5704
E-mail: products@CoraStyles.com
