MAKING AN ARRAY

(Starting with a gold slide)

1. Soak in 1 mM MUAM for at least 2 hr.

• rinse with ethanol first, then water; dry

2. Protect with Fmoc

• need to make up fresh Fmoc solution each time, as NHS ester is very reactive
• Fmoc-NHS is in dessicator in the fridge
• 3 mM Fmoc (0.5 mg) in 50% DMSO (300 uL) / 50% 0.1 M TEA pH 7.0 (200 uL)

a) dissolve the Fmoc in the DMSO, vortex, then add the TEA

b) cover surface of slide with this solution for 1 hour.

c) rinse with water

d) soak briefly in DMSO to remove excess Fmoc

3. Photopattern

• use pattern with the 750 um square holes

a) place slide in holder, gold side up

b) place pattern on slide, copper side down

c) photopattern for 1 hour at 400 Watts

d) rinse with EtOH

4. Soak in 1mM MUAM for 2 hour

5. Spot on SSMCC

• 0.4 mg of SSMCC (1mM) (in freezer) (in 0.1 M TEA pH 7.0)

a) let sit 30 min.

b) rinse with water, dry

6. Spot on thiol-DNA

• 1mM thiol-DNA

a) let sit in humid environment for at least 2 hours; can let sit overnight

b) rinse with water

7. Deprotect

a) submerge slide in 1M TAEA [tris (2-aminoethyl) amine] for 15 min.

• 900 uL TAEA in 5.1 mL DMF (both found in hood in rm. 3365)

b) rinse with water

8. "PEG"ylate

a) cover surface with 4mM PEG-NHS for 30 - 60 min

• 2mg PEG-NHS (in freezer) in 250 uL 0.1M TEA pH 8.0
• make up immediately before use, as NHS ester is very reactive

b) rinse with water

9. Store sample in phosphate buffer (20mM, pH 7.4) until ready to do imaging experiment

Copyright 1999 Corn Group

REFERENCE: J.M. Brockman et al. J. Am. Chem. Soc., 1999, 121, 8044-8051.