Innovative Tissue Array Technology for High-Throughput Screening of Gene,,,Expression ( were rehydrated and washed in PBS imm...)

Innovative Tissue Array Technology for High-Throughput Screening of Gene,,,Expression

were rehydrated and washed in PBS, immersed in 0.1 M glycine, and washed three times in PBS. Then, the slides were incubated with proteinase K and one was subsequently treated with RNase. To stop the reactions, slides were immersed in 4% paraformaldehyde in PBS followed by a PBS rinse. The slides were then incubated with acetic anhydride in triethanolamine and prehybridized in formamide (50% v/v) in 4X SSC at 37C for 30 minutes, then hybridized with 20 ml of poly d(T)-FITC incubated at 42C for 12 to 18 hours. This incubation was followed by a series of washes with 4X SSC. Each slide was examined with an Olympus BX60 fluorescent microscope.

Immunohistochemistry: Paraffin was removed from the array tissue by treating the slide as described in the product instructions. The slide was rehydrated, then incubated in PBS for 5 minutes and placed in a humidity chamber. The diluted primary antibody was added to the slide and incubated for 30 to 60 minutes at 37C. After washing with PBS, the diluted secondary antibody was added to the slide and incubated for 30 to 60 minutes at 37C. Following two additional PBS washes, a coverglass was placed over the slide, and the slide was examined using an Olympus BX60 fluorescent microscope.

REFERENCES

Battifora, H. (1986) Lab. Invest. 55(2): 244-248.
Battifora, H. and Mehta, P. (1990) Lab. Invest. 63(5): 722-724.
Kononen, J., et al. (1998). Nature Med. 4(7): 844-847.
Bubendorf, L., et al. (1999) Cancer Res. 59: 803-806.

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( were rehydrated and washed in PBS imm...)Innovative Tissue Array Technology for High-Throughput Screening of Gene,,,Expression

REFERENCES