Slide pretreatment

NOTE 1: This section should be performed simultaneously with the purification of the labeling. NOTE 2: Use aminosilane-coated slide only.

1. Prepare the following buffers and preheat at the right temperature:

Ethanolamine (120 ml; preheat at 50°C):

360 ml ethanolamine 12 ml Tris pH 9.0 1.2 ml 10% SDS 108 ml water

Blockwash buffer (120 ml; preheat at 50°C):

Prehybridization buffer (120 ml; preheat at 42°C):

30 ml 20 x SSC 90 ml water 1.2 g BSA 1.2 ml 10% SDS

2. Place the slide in a coplin jar or slide container and pour prewarmed ethanolamine blocking buffer onto the slide, and incubate the slide for 60 min at 50°C.

3. Wash the slide 5-7 times with distilled water.

4. Pour prewarmed Blockwash buffer onto the slide and incubate for 60 min at 50°C.

5. Wash the slide 5-7 times with distilled water.

6. Place the slide into a vertical slide holder (prevent drying of the slide).

7. Place the slide in boiling distilled water for 3 min for denaturation of DNA on the array.

8. Wash the slide two times with distilled water.

9. Place the slide back into the coplin jar or slide-container and pour prewarmed prehybridization buffer and incubate for 45 min at 42°C.

10. Wash the slide five times with distilled water.

11. Dry the slide as quickly as possible by centrifugation (5 min, 216 g)

12. Scan the slide to check for autofluorescence. The scan should now show a very low autofluorescence signal, if any. If not, repeat pretreatment of the slide.

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