Equipment

1. Falcon organ culture dish, 1 mL; Falcon tissue culture supplies. This dish has an inner well holding 1 mL of solution and an outer well that can be filled with media to facilitate maintenance of adequate humidity.

2. 30-gauge Becton Dickinson needles attached to 1-mL syringes. This size is optimal for flushing embryos from the uterine horns.

3. Micropipet glass capillary, 100-^L volume pulled by heat to a fine point. The tip is broken off using a diamond-tipped pencil.

4. Braking pipet and loading tools, made by hand as described in ref. 6.

5. Vacuum drying tube (cat no. 6699-10; Ace Glass Inc., Vineland, NJ). The individual slides are stored back to back in a wooden slide holder containing a total of six slides.

6. Dissecting microscope with magnification of 0.5X or 1X objective with 10X oculars.

7. Teflon or polyethylene block; 25 X 120 X 5 mm drilled with 60 holes of 3-mm diameter used as an oil well.

8. Oil well oil: 30% Hexadecane and 70% USP light mineral oil. This mixture gives the correct viscosity to protect the added fluid droplet from access to CO2 from air and still allow the droplet to fall rapidly to the bottom of the well when delivered from the constriction pipet. To ensure that the oil is free of disturbing impurities, the oil mixture is washed 3 times by shaking vigorously in a separatory funnel with 1 volume of 0.5 M NaOH. The oil phase is removed and washed, and 0.01 M HCl and six changes of water until the washings are neutral. The oil is finally cen-trifuged at 10,000 rpm and the oil phase removed from the water phase. The last traces of water are removed by drying the oil in a vacuum dessicator.

9. Freeze-dryer attached to —35°C freezer with connectors for vacuum tubes.

10. Borosilicate glass fluorometer tubes, 10 X 75 mm.

11. Fluorimeter; a filter-type fluorimeter with a photomultiplier tube and continuously variable sensitivity from Optical Technology Devices (Elmsford, NY).

12. Constriction pipet (Bie and Berntsen Glass, Copenhagen, Denmark). These fine-tipped pipets should be made of quartz and bent at an angle of 45° so that it is possible to pipet into the oil with the hand steadied by resting on the working surface.

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