1. Remove the hybridization cocktail from the array into a new RNAse-free tube and store it at -20°C. The same hybridization cocktail can be used again (denature the cocktail at 99°C for 5 min before each use).

2. Fill the probe array manually with 250 |l of non-stringent wash buffer. The probe array can be stored up to 3 h at 4°C in the dark before proceeding with the washing and staining.

3. Turn on the power for the hardware (note the order recommended by the manufacturer). Open scanner-operating software (e.g. GCOS).

4. Prepare staining cocktails.

SAPE solution (1200 |l per array)

600 |l 2 x stain buffer 540 |l RNAse-free water 48 |l acetylated BSA (50 mg/ml) 12 |l SAPE (1mg/ml)

Mix well and divide into two aliquots of 600 |l each.

Antibody solution (600 |l per array)

300 | l 2 x stain buffer

266.4 |l RNAse-free water

24 |l acetylated BSA (50 mg/ml)

3.6 |l biotinylated antibody (0.5 mg/ml)

Mix well.

Affymetrix GeneChip analyses - the impact of RNA quality

0 0

Post a comment