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Arichard:Protocols/Glass micropipette fabrication
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==Glass micropipette fabrication== * Sutter Model P-90 ** Heat #1 = 700 ** Pull = 600 * Sutter borosilicate glass with filament ** O.D. 1 mm, I.D. 0.5 mm, length 10 cm ** Item #: BF100-50-10 ===Prep=== * Aliquot ~150 ul nuclease-free H2O into 0.2 ul tube. * Aliquot ~150 ul Sigmacote into 0.2 ul tube. * UV treat, lids open. ===Protocol=== * Each glass tube makes 2 pipettes ====Pulling==== # Turn the machine on # Position the glass tube in the groove on the upper clamp of the puller. Ensure that the metal resistive heating element is located at the center of the tube. Tighten the upper clamp. # Slide the lower clamp upwards until it stops. Tighten the lower clamp. # Press pull. The heating element will glow and the bottom clamp will drop in 5-10 seconds. # Allow 5-10 seconds for the glass to cool before removing the pulled pipettes. ====Bending==== # Light bunsen burner. Fully open the carb at bottom. Smallest flame is ideal. # Hold pipette by blunt end. # Carefully position the pipette such that the flame heats ~3/4 towards the tip. Use tweezers to guide the bend to ~15 degrees. # Allow 5-10 seconds to cool. # Break the tip against a smooth clean surface. (Flame often seals the tip closed) ====Coating==== * Do not work with Sigmacote outside the fume hood. # Dip each tip into Sigmacote aliquot. Ensure that liquid is drawn into the tip. Tip diameters will vary so it may take a few seconds to draw up the Sigmacote. # Mount pipette onto the microscope manipulator. Press clear after attaching pipette to flush. # Hold water tube under pipette and rinse 3x up and down. Press clear again. # UV treat.
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