ArrayNL platform^©

ArrayNL - second meeting

Contributed by Peter Groot and Klaas Kok

Praktical questions with regards to the making and hybridization of micro-arrays

One of the goals of the "ArrayNL-platform" is to share our practical experiences. For this purpose, at the next ArrayNL-microarray meeting (Utrecht, 23 March) we will organize a practical workshop. The goal of this workshop will be to exchange practical information, based on actual experiments. For this, we need your assistance.

This mail is a request to let us know of your experience (mail to Peter Groot and Klaas Kok), based on real experimental data. We will try to summarize this and present it at the workshop as well as on the ArrayNL website. We urge you to indicate if and what your experience is with the issues listed below (both positive and negative). Those who can illustrate specific points with experimental data are invited to do this in short (5-10 min) presentations during the workshop. The aim of all this is to help each other in developing good, robust protocols for the production, hybridization and analyses of micro-arrays and to prevent us from reinventing the wheel ever so many times.

Please indicate what your experience is with the issues below (remember that negative experiences may be equally useful as positive ones, especially if they prevent others from making the same "errors").

1. procedures used to clean and coat glass slides
   • type of glass-slides used, manufacturers
   • how long is it stable?
2. procedures to covalently bind DNA
   (UV, "baking", aging)
3. quality control glass slides
   • amount of DNA bound
   • background fluorescence
4. commerciale available glass slides, e.g.
   • GAP-coated (Corning)
   • FAST Slides (Schleicher & Schuell)
   • other?
5. spotting of DNA
   • influence of humidity and temperature of the room
6. solvents used to spot the DNA
   • 3x SSC / 5x SSC / 3x SSC + 50 % DMSO / other
7. different hybridization-solutions
   • SSC
   • SSC / formamide
   • Ultrahyb (Ambion)
   • effect of BSA
   • other
8. commercially vs home-made hybridization chambers
   • perspex boxes (as used for standard in situ's)
   • Corning hybridization chambers
   • other?
9. preparation of the DNAs to spot (the probe)
   • PCR-protocols (yields suffiecient for spotting?)
   • use of aminolinkers:
     • significant increase of fluorscent signal?
     • stripping of slides possible?
   • purification-protocols (precipitation)
10. different arrayers
    • pin-ring system / solid tip / inktjet systems
11. different scanners
12. microtiterplates used
13. different methods for fluorescent labeling of target DNA
    • enzymatic
    • chemical labeling (ULS-system, Kreatech)
    • fluorochromes used (Cy3, Cy5 or other)
14. analysis software
    • Sscanalyze
    • other
15. other points of interest which are not listed here

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Email or send your reactions to:

Peter Groot (P.C.Groot@pharm.uu.nl) or	Klaas Kok (K.Kok@medgen.AZG.nl)
Dept. Pharmacol, Fac. Pharmacy		Dept of Medical Genetics
Universiteit Utrecht			University Groningen 
Postbus 80082				Antonius Deusinglaan 4
3508 TB Utrecht				9713 AW Groningen
Tel: 030 2537355			Tel: 050-3632925 (2251)
Fax: 2537420				Fax: 050-3632947
Fax to E-mail: 020-8831442
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