title: Stanford Human cDNA SHEW
status: Public on Nov 16 2007
contact.city: Boston
contact.first: Bao Jian
contact.last: Fan
contact.email: baojian_fan@meei.harvard.edu
contact.state: MA
contact.address: MEEI, 243 Charles Street
contact.department: Ophthalmology
contact.country: USA
contact.institute: Harvard Medical School
contact.phone: 617-573-6448
organism: Homo sapiens
manufacture_protocol: Preparation of Slides to be used to make microarrays.
  Last updated: October 6, 1999
  Materials
  Qty
  Order info
  Notes
  Glass microscope slides
  60
  Gold Seal #3010
  Slide rack
  2
  Shandon Lipshaw #121 (800-245-6212)
  Each rack holds 30 slides
  Slide chamber
  6
  Shandon Lipshaw #121
  Each chamber holds 350 mL
  ddH2O
  ~5 L
  NaOH
  70 g
  95% Ethanol
  420 mL
  Poly-L-lysine
  70 mL
  Sigma #P 8920
  Tissue culture PBS
  70 mL
  Vacuum oven (45C)
  Slide box (plastic only)
  1
  VWR #48443-806
  1. Place slides in slide racks. Place racks in chambers.
  2. Prepare CLEANING SOLUTION:
  a. Dissolve 70 g NaOH in 280 mL ddH2O.
  b. Add 420 mL 95% ethanol. Total volume is 700 mL (= 2 X 350 mL); stir until completely mixed.
  c. If solution remains cloudy, add ddH2O until clear.
  3. Pour solution into chambers with slides; cover chambers with glass lids. Mix on orbital shaker for 2 hr. Once slides are clean, they should be exposed to air as little as possible. Dust particles will interfere with coating and printing.
  4. Quickly transfer racks to fresh chambers filled with ddH2O. Rinse vigorously by plunging racks up and down. Repeat rinses 4X with fresh ddH2O each time. It is critical to remove all traces of NaOH-ethanol.
  5. Prepare POLYLYSINE SOLUTION:
  a. 70 mL poly-L-lysine + 70 mL tissue culture PBS in 560 mL water.
  b. Use plastic graduated cylinder and beaker.
  6. Transfer slides to polylysine solution and shake 15 min. - 1 hr.
  7. Transfer rack to fresh chambers filled with ddH2O. Plunge up and down 5X to rinse.
  8. Centrifuge slides on microtiter plate carriers (place paper towels below rack to absorb liquid) for 5 min. @ 500 rpm. Transfer slide racks to empty chambers with covers for transport to vacuum oven.
  9. Dry slide racks in 45C vacuum oven for 10 min. (Vacuum is optional.)
  10. Store slides in closed slide box (plastic only, without rubber mat bottom)
  11. BEFORE PRINTING ARRAYS:
  a. Check that polylysine coating is not opaque.