Incubate the array in the hybridization oven for at least 30 minutes.

8. Hybridization of Cy3 + Cy5 probe to glass array (NON-Automated version).

Denature the probe at 95oC for 3-5 minutes in PCR machine.

Transfer immediately to 65C waterbath.

Remove the pre-hybridization solution from array slide cover chamber using P-1000 pipette.

Add:

15 ul of probe to 500 ul (pre-warmed at 65C) hybridization solution.(Clonetech, ExpressHyb)

Mix well and Add directly to array cover chamber.

Wipe any spill gently with a Kimwipe.

Re-Seal the chamber.

Incubate at 65C, overnight in hybridization oven.

9. Preparation of Dendrimer Cy3 and Cy5.

Place 3DNA dendrimer hybridization buffer at 65oC to incubate until use (about 10 minutes).

Prepare a mixture of Cy3 dendrimer and Cy5 dendrimer in a 1:1 ratio.

(a stock solution of 36 ul (18ul each) can be made and kept in a light block tube at +4oC)

Incubate the dendrimer mixture at 65oC for 10 minutes.

Vortex the mixture again for 30-60 seconds to break up aggregates.

Jeffrey D. Randall, Takumi Yoshida, Ryoji Kojima, Steven Gullans ?Page 6 9/6/2001

Jeffrey D. Randall, Takumi Yoshida, Ryoji Kojima, Steven Gullans ?Page 6 9/6/2001

Incubate at 65oC until use (Should be about 10 more minutes).