Preparation of Slides
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Preparation of Slides
Last updated: October 6, 1999
Materials | Qty | Order info | |
---|---|---|---|
Glass microscope slides | 60 | Gold Seal #3010 | |
Slide rack | 2 | Shandon Lipshaw #121 (800-245-6212) | <= Each rack holds 30 slides |
Slide chamber | 6 | Shandon Lipshaw #121 | <= Each chamber holds 350 mL |
ddH2O | ~5 L | ||
NaOH | 70 g | ||
95% Ethanol | 420 mL | ||
Poly-L-lysine | 70 mL | Sigma #P 8920 | |
Tissue culture PBS | 70 mL | ||
Vacuum oven (45C) | |||
Slide box (plastic only) | 1 | VWR #48443-806 |
1. | Place slides in slide racks. Place racks in chambers. |
2. | Prepare CLEANING SOLUTION: |
Dissolve 70 g NaOH in 280 mL ddH2O. | |
Add 420 mL 95% ethanol. Total volume is 700 mL (= 2 X 350 mL); stir until completely mixed. | |
If solution remains cloudy, add ddH2O until clear. | |
3. | Pour solution into chambers with slides; cover chambers with glass lids. Mix on orbital shaker for 2 hr. |
Once slides are clean, they should be exposed to air as little as possible. Dust particles will interfere with coating and printing. | |
4. | Quickly transfer racks to fresh chambers filled with ddH2O. Rinse vigorously by plunging racks up and down. |
Repeat rinses 4X with fresh ddH2O each time . It is critical to remove all traces of NaOH-ethanol. | |
5. | Prepare POLYLYSINE SOLUTION: |
70 mL poly-L-lysine + 70 mL tissue culture PBS in 560 mL water. | |
Use plastic graduated cylinder and beaker. | |
6. | Transfer slides to polylysine solution and shake 15 min. - 1 hr. |
7. | Transfer rack to fresh chambers filled with ddH2O. Plunge up and down 5X to rinse. |
8. | Centrifuge slides on microtiter plate carriers (place paper towels below rack to absorb liquid) for 5 min. @ 500 rpm. |
Transfer slide racks to empty chambers with covers for transport to vacuum oven. | |
9. | Dry slide racks in 45C vacuum oven for 10 min. (Vacuum is optional.) |
10. | Store slides in closed slide box (plastic only, without rubber mat bottom) |
11. | BEFORE PRINTING ARRAYS: |
Check that polylysine coating is not opaque. | |
Test print, hyb and scan sample slides to determine slide batch quality. |