Equipment for Culture Chambers

2.1.1. Cover Slips and Cleaning Materials

1. 24 x 24 mm2 cover slips (we use Marienfeld, product no. 5370).

2. Porcelain staining racks for cover slips (product no. 8542-E42, Thomas Scientific, www.thomassci.com).

4. Long anatomical forceps, 200 mm (Bochem 18/8).

5. Pyrex beaker, 1-L capacity.

6. Cleaning solution 7X® PF 1% in distilled water (ICN Biomedicals, Inc., cat. no. 76-671-210).

7. Standard distilled water.

8. Ultrapure water (double distilled, or Milli-Q).

9. Staining glass box with cup, 90 x 60 x 50 mm3.

10. Ethanol absolute grade (GR) for analysis (Merck, cat. no. K30598083).

2.1.2. Culture Chamber Mounting

1. 24 x 24-mm2 Cover slips, treated as discussed in Subheading 3.1.1.

2. Aluminum slides with a 20-mm hole bored through its center, treated as discussed in Subheading 3.1.3. (we make them ourselves from aluminum sheets).

3. Acetone GR for analysis (Merck).

4. Tissue papers. (We use professional wipes; Kimberly-Clark, code 7102.)

6. Bunsen burner.

7. Nail polish.

8. Slides support. We make it ourselves by attaching two foam stripes to the bottom of a box that can be closed afterward. The slides must touch the support only at the two extremes.

2.1.3. Microscope Slide Cleaning and Recycling

1. Eight staining glass boxes with cup, 90 x 60 x 50 mm3.

2. Removable staining tray for slides, 76 x 26 mm2.

3. Wire clip for staining tray.

4. Aceton GR for analysis (Merck cat. no. K30688514).

5. Xylene 99+%, high-performance liquid chromatography (HPLC) grade (Sigma-Aldrich, cat. no. 31,719-5).

6. Ethanol, absolute GR for analysis (Merck, cat. no. K30598083).

2.2. Equipment for Preparing Living Spermatocytes

1. Newly eclosed Drosophila males, expressing the GFP-fusion protein of interest in testis.

2. Standard CO2 source.

3. Standard dissecting microscope with a white-light source.

4. Heavy white mineral oil (Trinity Biotech GmbH, cat. no. 400-5) specific gravity (77°F): 0.875-0.885.

5. Two straight dissection forceps (A. Dumont inox No. 5).

6. Filter paper, extremely thin, cut into triangles of approx 3 cm/side (Schleicher & Schuell 595, cat. no. 10311614).

7. Sterile needles (BD Microlance™ 3, 27-gage 3/4, 0.4 x 19 mm, No. 20. (Becton Dickinson, cat. no. 302200).

2.3. Image Recording and Processing Equipment for Phase-Contrast Microscopy

1. Inverted microscope. We use a Leica DM IRB/E.

2. Charge coupled device (CCD) camera. We use a CCD Cohu High performance Model 4912-5100.

3. Temperature controller. We use either a temperature-controlled chamber specially built for the microscope, or a Bioptechs objective controller.

4. Objectives. We use either an oil 63 x /1.32 PLAN APO Ph3 or an oil 100 x /1.25 C PLAN Ph3.

5. Shutter. We use a Vincent V Uniblitz electronic.

6. Shutter driver. Uniblitz Model D122.

7. Computer for recording. We use a Power Macintosh 9600/233, with a video card compatible with the camera and the shutter driver.

8. Acquisition software: NIH-Scion Image, version 1.62. (www.rsb.info.nih.gov/ nih-image).

9. Computer for image processing. Any computer having enough RAM memory and processing speed. We use a single processor 500-MHz power Macintosh G4 with 384MB RAM memory.

10. Processing software: NIH-Scion Image, version 1.62. (www.rsb.info.nih.gov/nih-image).

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