Compatibility of DMSO spotting buffers with SCHOTT Nexterion Epoxy-silane coated microarray slides. Effect of using different spotting pins and DMSO concentrations.

1. November 2004DMSO Spotting BufferMicroarray Solution
SCHOTT Jenaer Glas GmbH
Final Report
DMSO Spotting Buffers on Slide E
Norman Rahnis, Kornelia Kuhn, Katrin Steinmetzer
Application Support/R&D
Microarray Solutions
Schott Jenaer Glas GmbH

2. November 2004DMSO Spotting BufferMicroarray Solution
SCHOTT Jenaer Glas GmbH
Spotting Solution
Components DMSO, SSC, phosphate buffer, SDS
Slides Nexterion Slide E
Probes 70mer (from E. coli Genome Oligo Set Version 1.0) 10 µM
Spotting QArray Mini, 22°C, 45% RH, SMP3 pin (Telechem), aQu Split
Pin K2801 (diameter 150 µm)
Target total RNA (E.coli) was used to synthesize aa-cDNA, the coupling
of NHS-ester-dyes (Nexterion Dye3 and Dye5) was performed
with a standard protocol derived from a Cy3/Cy5 labeling
protocol,
Hybridization Tecan Hybridization Station HS4800, in 5xSSC, 0.1% SDS, 0.1
mg/ml herring sperm DNA, 50% formamide, 42°C, 16 hrs
Scanning Affymetrix Array Scanner 428
Analysis IconoClust
Experimental Details

3. November 2004DMSO Spotting BufferMicroarray Solution
SCHOTT Jenaer Glas GmbH
....some remarks
the following images show results after hybridization
blue spots mean lower signal, green spots mean higher signals
suitable buffers were highlighted in red
spot diameter was only determined for good spots

4. November 2004DMSO Spotting BufferMicroarray Solution
SCHOTT Jenaer Glas GmbH
Results SMP3 Pin 50% DMSO
50% DMSO 3x SSC
50% DMSO 1.5x SSC, spot diameter 70 µm
50% DMSO 150 mM PP, 0.1% SDS
50% DMSO 150 mM PP, 0.05% SDS
50% DMSO 150 mM PP
50% DMSO

5. November 2004DMSO Spotting BufferMicroarray Solution
SCHOTT Jenaer Glas GmbH
Results SMP3 Pin 30% DMSO
30% DMSO 3x SSC
30% DMSO 1.5x SSC,
spot diameter 70 µm
30% DMSO 150 mM PP, 0.1% SDS
30% DMSO 150 mM PP, 0.05% SDS
30% DMSO 150 mM PP
30% DMSO

6. November 2004DMSO Spotting BufferMicroarray Solution
SCHOTT Jenaer Glas GmbH
Results SMP3 Pin
25% DMSO 0.005% SDS
50% DMSO 1.5x SSC, 0.005% SDS,
spot diameter 60-70 µm
25% DMSO 1.5xSSC, 0.005% SDS
Spot diameter 90 µm
50% DMSO 75 mM PP
best results were obtained when combining DMSO with SSC
and with or without SDS (but only for f.c. SDS of 0.005%, higher
concentrations lead to precipitation)
a combination of DMSO, phosphate buffer and SDS does not
work
further tests with additional combinations of DMSO, SSC and
other detergents will be tested

7. November 2004DMSO Spotting BufferMicroarray Solution
SCHOTT Jenaer Glas GmbH
Results SMP3 Pin
50% DMSO 2x SSC
best results were obtained when combining DMSO with SSC
and with or without detergents
spot sizes range between 100 and 140 µm
similar results were obtained with 30% DMSO
50% DMSO 1.5x SSC, 0.025% CTAB
50% DMSO 1.5x SSC, 0.025% Triton X100
50% DMSO 2.5x SSC
50% DMSO 1.5x SSC, 0.005% SDS

8. November 2004DMSO Spotting BufferMicroarray Solution
SCHOTT Jenaer Glas GmbH
Results aQu K2801 Pin 50% DMSO
50% DMSO 3x SSC
50% DMSO 1.5x SSC
50% DMSO 150 mM PP, 0.1% SDS
50% DMSO 150 mM PP, 0.05% SDS
50% DMSO 150 mM PP
50% DMSO
no suitable buffer combinations

9. November 2004DMSO Spotting BufferMicroarray Solution
SCHOTT Jenaer Glas GmbH
Results aQu K2801 Pin 30% DMSO
30% DMSO 3x SSC
spot diameter 360 µm
30% DMSO 1.5x SSC,
spot diameter 90 µm
30% DMSO 150 mM PP, 0.1% SDS
30% DMSO 150 mM PP, 0.05% SDS
30% DMSO 150 mM PP
30% DMSO
30% DMSO with 1.5x SSC gives good results, further SSC-
concentrations will be tested

10. November 2004DMSO Spotting BufferMicroarray Solution
SCHOTT Jenaer Glas GmbH
Results aQu K2801 Pin
25% DMSO 0.005% SDS
50% DMSO 1.5x SSC, 0.005% SDS,
spot diameter 110 µm
25% DMSO 1.5xSSC, 0.005% SDS
Spot diameter 160 µm
50% DMSO 75 mM PP
best results were obtained when combining DMSO with SSC
and with or without SDS (but only for f.c. SDS of 0.005%, higher
concentrations lead to precipitation)
a combination of DMSO, phosphate buffer and SDS does not
work
further tests with additional combinations of DMSO, SSC and
other detergents will be tested

11. November 2004DMSO Spotting BufferMicroarray Solution
SCHOTT Jenaer Glas GmbH
Results aQu K2801 Pin
50% DMSO 1.5x SSC, 0.025% CTAB
50% DMSO 1.5x SSC, 0.025% Triton X100
50% DMSO 2.5x SSC
50% DMSO 2x SSC
best results were obtained when combining DMSO with SSC
and with or without detergents
spot sizes range between 100 and 140 µm
similar results were obtained with 30% DMSO
50% DMSO 1.5x SSC, 0.005% SDS

12. November 2004DMSO Spotting BufferMicroarray Solution
SCHOTT Jenaer Glas GmbH
Summary
50 (30%) DMSO in water usually gives very small spots on Slide E
(approximately 60 µm)
addition of SSC (f.c. 1.5x to 2.5x) with or without detergent (f.c. 0.005% to
0.025%) gives good spot morphology and size
spot size can be increased by increasing SSC concentration and/or adding
detergent
50(30)% DMSO is not compatible with Next Spot or phosphate buffer (150
mM f.c. phosphate buffer), addition of detergent does not improve spot
morphology*