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COMPANY PRESENTATION      PRODUCT GROUP NEXTERIONSCHOTT Nexterion®The Microarray Substrate Company                        ...
COMPANY PRESENTATION                      PRODUCT GROUP NEXTERION      Approaches to increase signal or sensitivityObvious...
COMPANY PRESENTATION                     PRODUCT GROUP NEXTERION      SCHOTT’s solution is a fluorescence enhancing       ...
COMPANY PRESENTATION                             PRODUCT GROUP NEXTERION          How does the optical coating work?      ...
COMPANY PRESENTATION                               PRODUCT GROUP NEXTERION          How does the optical coating work?    ...
COMPANY PRESENTATION                   PRODUCT GROUP NEXTERION       Benchmark study on Nexterion® HiSens E               ...
COMPANY PRESENTATION                 PRODUCT GROUP NEXTERION                       Objectives of the study     Comparing S...
COMPANY PRESENTATION                          PRODUCT GROUP NEXTERION                        Experimental DetailsSlides   ...
COMPANY PRESENTATION                              PRODUCT GROUP NEXTERION        Higher Signals with Nexterion® HiSens E  ...
COMPANY PRESENTATION                                                                             PRODUCT GROUP NEXTERION  ...
COMPANY PRESENTATION                                                                             PRODUCT GROUP NEXTERION  ...
COMPANY PRESENTATION                                                               PRODUCT GROUP NEXTERION    8 fold highe...
COMPANY PRESENTATION                   PRODUCT GROUP NEXTERIONAmbion Inc.: Slide E versus HiSens E for miRNA arraysmiRNA f...
COMPANY PRESENTATION         PRODUCT GROUP NEXTERION      10 times less target and better signals!20µg totalRNA equival. o...
COMPANY PRESENTATION                                        PRODUCT GROUP NEXTERION          Quantification of signal-back...
COMPANY PRESENTATION                                                     PRODUCT GROUP NEXTERION                      Corr...
COMPANY PRESENTATION                        PRODUCT GROUP NEXTERION     Dye swap experiments of Nexterion® HiSens E at    ...
COMPANY PRESENTATION                                                                              PRODUCT GROUP NEXTERION ...
COMPANY PRESENTATION               PRODUCT GROUP NEXTERION       Different coatings and formats available....             ...
COMPANY PRESENTATION                    PRODUCT GROUP NEXTERION                       Conclusions       • On average 8 fol...
COMPANY PRESENTATION      PRODUCT GROUP NEXTERIONSCHOTT Nexterion®The Microarray Substrate Company                        ...
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SCHOTT NEXTERION® HiSens optical coating for high sensitive microarray analysis

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The reflective layers on Nexterion® HiSens significantly enhance sensitivity and signal response. The characteristics of the reflective layers have been optimized for the fluorescent wavelengths most commonly used in microarray experiments, and will simultaneously improve the performance in both the Cy3™ and Cy5™ channels. The slide is produced according to industry standard slide dimensions and is available with SCHOTT’s standard high quality functional coatings for DNA and protein microarraying. This means that the Nexterion® HiSens Slide is fully compatible with all microarray printing technologies and most slide processing protocols, allowing customers a smooth transition from industry-standard transparent microarray slides.

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SCHOTT NEXTERION® HiSens optical coating for high sensitive microarray analysis

  1. 1. COMPANY PRESENTATION PRODUCT GROUP NEXTERIONSCHOTT Nexterion®The Microarray Substrate Company SCHOTT HiSens Slides: A Cure for the Sensitivity Blues – Achieve Higher Signals with Less Target
  2. 2. COMPANY PRESENTATION PRODUCT GROUP NEXTERION Approaches to increase signal or sensitivityObvious options• Selection of appropriate slide surface• Optimized handling to achieve optimal signal/background ratios• Increase the dye load per target moleculeMore sophisticated options• Increase the dye load by for instance dendrimer technology or nanoparticles• Use of scanners with low intrinsic background levels• Use of optical coatings on slides
  3. 3. COMPANY PRESENTATION PRODUCT GROUP NEXTERION SCHOTT’s solution is a fluorescence enhancing slide coating Nexterion® HiSens SlidesØ Launched January 2006Ø Innovative optical coating specifically developed for microarray substratesØ Fully compatable with any attachement chemistry and arraying hardwareØ Unique coating developed using SCHOTT expertise in optical glass technology
  4. 4. COMPANY PRESENTATION PRODUCT GROUP NEXTERION How does the optical coating work? 1. Increase of excitation Reflecting wave overlaps incoming wave Standing wave pattern ⇒ Organic coating Thin-film dielectric coatingCoating is optimized to give the maximum E-field field strengthin the vicinity of the surface ⇒ strongest fluorescence excitation
  5. 5. COMPANY PRESENTATION PRODUCT GROUP NEXTERION How does the optical coating work? 2. Maximize reflection towards the detector λex Fluorescence emission λex Fluorescence emission Spots with labeled DNA or oligos Organic coating Thin-film dielectric glass substrate coating (borofloat) glass substrate (borofloat) within ~ 50 nm fluorescence is enhanced Coating is optimized to give the maximum reflection towards the detectorBoth effects together ⇒ theoretically 12-foldenhancement of slide performance
  6. 6. COMPANY PRESENTATION PRODUCT GROUP NEXTERION Benchmark study on Nexterion® HiSens E Which improvements can be obtained with Nexterion® HiSens E slides?
  7. 7. COMPANY PRESENTATION PRODUCT GROUP NEXTERION Objectives of the study Comparing Slide E against Slide HiSens E against the following parameters: • Signal intensity • Target concentrations used • Number of genes detectable • Data correlation
  8. 8. COMPANY PRESENTATION PRODUCT GROUP NEXTERION Experimental DetailsSlides Nexterion® Slide E, Nexterion® HiSens ESlide Processing standard protocolsProbes 70mer oligos, human AROS V4 (1 plate) + control plateSpotting QArray Mini (Genetix), SMP3 pins (ArrayIt)Target Dye 3 labeled cDNA derived from human brain/liver total RNA, Dye 5 labeled cDNA derived from human liver/brain total RNAHybridization 2h - 8h, Tecan Hyb station HS4800Scanning Affymetrix 428 and Tecan LS400 reloadedAnalysis IconoClust (Clondiag)
  9. 9. COMPANY PRESENTATION PRODUCT GROUP NEXTERION Higher Signals with Nexterion® HiSens E AROSAROS Control Oligos Oligos Slide E Slide HiSens E Intensity Lo Hi
  10. 10. COMPANY PRESENTATION PRODUCT GROUP NEXTERION Higher Signals with Nexterion® HiSens E Comparison of Signal Intensity Nexterion® Slide E vs. Slide HiSens E (Gain: 30 dB, Cy3) Slide E 1 Slide HiSens E 0.9 0.8 0.7 Intensity 0.6 0.5 0.4 0.3 0.2 0.1 0 Background 1AA01 1AB01 1AC01 1AD01 1AE01 1AF01 1AG01 1AH01 1BA01 1BB01 1BC01 1BD01 1BE01 1BF01 1BG01 1BH01 Oligo/Gene
  11. 11. COMPANY PRESENTATION PRODUCT GROUP NEXTERION More detectable signals with Nexterion® HiSens E Comparison of Signal Intensity Nexterion® Slide E vs. Slide HiSens E (Gain: 30 dB, Cy3) Slide E 1 Slide HiSens E 0.9 0.8 0.7 Intensity 0.6 0.5 0.4 0.3 0.2 0.1 0 Background 1AA01 1AB01 1AC01 1AD01 1AE01 1AF01 1AG01 1AH01 1BA01 1BB01 1BC01 1BD01 1BE01 1BF01 1BG01 1BH0125-35 % additional low expressor genes (30 of 96 testet onthis slide) are detectable with HiSens E only Oligo/Gene
  12. 12. COMPANY PRESENTATION PRODUCT GROUP NEXTERION 8 fold higher performance with Nexterion® HiSensE Signal Enhancement Factor Nexterion ® Slide E vs. Slide HiSens E (Gain: 30 dB, Cy3) 17 16 15 14 13 12 11 10Factor 9 8 Mean 7 6 5 4 3 2 1 0 1AA01 1AB01 1AC01 1AD01 1AE01 1AF01 1AG01 1AH01 1BA01 1BB01 1BC01 1BD01 1BE01 1BF01 1BG01 1BH01 Signals on average Oligo/Gene amplified approx. 8-fold
  13. 13. COMPANY PRESENTATION PRODUCT GROUP NEXTERIONAmbion Inc.: Slide E versus HiSens E for miRNA arraysmiRNA fractions from lung and bladder were hybridized against printed oligos20µg totalRNA equivalents on Slide E on HiSens E
  14. 14. COMPANY PRESENTATION PRODUCT GROUP NEXTERION 10 times less target and better signals!20µg totalRNA equival. on E 2µg totalRNA equival. on HiSens E
  15. 15. COMPANY PRESENTATION PRODUCT GROUP NEXTERION Quantification of signal-background 1. median signal – local background 2. normalization to ratios of slide E 20 µg. eq Sum Intensity 100000 90000 80000 70000 60000 RFU 50000 40000 30000 20000 10000 0 Oligo avg E avg HiSens • On average 7.8 fold increase of net signal • 50% more signals detectable
  16. 16. COMPANY PRESENTATION PRODUCT GROUP NEXTERION Correlation of Nexterion® HiSens E and Slide E data HiSens vs Slide E Optimized Scanning 6 80000 Correlation SlideE 20 µg & Slide E 5 µg = 0.991834 Slide E 20 µg & Slide E 2 µg = 0.958447 70000 4 Slide E 20 µg & HiSens 20 µg = 0.983246 Slide E 20 µg & HiSens 5 µg = 0.969035 Slide E 20 µg & HiSens 2 µg = 0.971587 60000 HiSens 20 µg & HiSens 5 µg = 0.990909 2 HiSens 20 µg & HiSens 2 µg = 0.973892 50000Log(base2)R/G 0 40000 30000 -2 20000 -4 10000 -6 0 psID SlideE 20ug avg SlideE 5ug avg SlideE 2 ug avg HiSens 20ug avg HiSens 5ug avg HiSens 2ug avg AVERAGE sum med. Int. Ø High data correlation between slide E and HiSens E Ø High data correlation between different target concentrations
  17. 17. COMPANY PRESENTATION PRODUCT GROUP NEXTERION Dye swap experiments of Nexterion® HiSens E at City of Hope National Medical CentermiRNA array with Ambion‘s mirVana probes dissolved in low evaporation buffer Hu.Thymus 555 Hu.Thymus 555 Hu.Brain 647 Hu.Brain 647
  18. 18. COMPANY PRESENTATION PRODUCT GROUP NEXTERION Data correlation Hu.Thymus 555 Hu.Thymus 555 Hu.Brain 647 Hu.Brain 647 20000 hsa_mi hsa_mir r_16 hsa_mir _26a 10000 hsa_mi ambi_mir hsa_mi _124a r_205 hsa_mi _7029 r_494 hsa_mir r_26a _494 F532 Median - B532F532 Median - B532 hsa_mir 2000 _16 1000 ambi_mir _7029 hsa_mi r_124a hsa_mir _205 100 200 2000 20000 200 2000 20000 F635 Median - B635 F635 Median - B635 Slide 31 Slide 30 Ø Data show excellent correlation with dye swap on HiSens E
  19. 19. COMPANY PRESENTATION PRODUCT GROUP NEXTERION Different coatings and formats available.... Nexterion® Slide E Nexterion® HiSens Epoxy chemistry for E, A and H chemistry Nexterion® Slide A Amino chemistry MPX format for all functional coatings Nexterion® Slide A+ Amino chemistry MTP format for E and A chemistry Nexterion® Slide AL Aldehyde chemistry Nexterion® Slide H NHS-ester chemistry
  20. 20. COMPANY PRESENTATION PRODUCT GROUP NEXTERION Conclusions • On average 8 fold higher net signals • 3-4 fold increased signal/noise ratio • Identification of weakly expressed genes 25%-50% more genes detectable! • Less target/probe required • Protocols identical to conventional slides • Compatible with all microarray equipment (except scanners that image through the rear face of the slide)
  21. 21. COMPANY PRESENTATION PRODUCT GROUP NEXTERIONSCHOTT Nexterion®The Microarray Substrate Company SCHOTT HiSens Slides: A Cure for the Sensitivity Blues – Achieve Higher Signals with Less Target

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