| AVS 55th International Symposium & Exhibition | |
| BioMEMS Topical Conference | Tuesday Sessions |
| Session BM+BI+BO+NC-TuA |
| Session: | Microfluidics/Lab-on-a-Chip |
| Presenter: | D.W. Grainger, University of Utah |
| Authors: | F. Cheng, University of Washington H. Takahashi, University of Utah M. Dubey, University of Washington K. Emoto, Acclerys Technology Corporation L.J. Gamble, University of Washington D.W. Grainger, University of Utah D.G. Castner, University of Washington |
| Correspondent: | Click to Email |
N-hydroxysuccinimide (NHS) esters are widely used to activate covalent coupling of amine-containing biomolecules onto surfaces in academic and commercial surface immobilizations in many applications. However, their intrinsic hydrolytic instability is well-known and limits this reactive surface chemistry. No methods are known to quantify this chemistry conveniently. We have used x-ray photoelectron spectroscopy (XPS) and time-of-flight secondary ion mass spectrometry (ToF-SIMS) to investigate surface hydrolysis and spatial reactivity in NHS-bearing thin films.1 Principal component analysis (PCA) of ion ToF-SIMS data correlates changes in the NHS chemistry as a function of conditions. NHS ester oligo(ethylene glycol) (NHS-OEG) monolayers on gold and commercial polymer films have been compared after surface treatments. From PCA results, multi-variate peak intensity ratios were developed to assess NHS reactivity, thin film thickness and oxidation of the monolayers during surface hydrolysis. Aging in ambient air up to seven days results in some NHS hydrolysis and thiol oxidation. Overnight film immersion under water completes hydrolysis and NHS removal. The same PCA peak intensity ratios for surface coupling of amine-terminated molecules confirmed that NHS surface regeneration methods re-establish bound NHS concentrations approximately 50% of that on freshly prepared NHS-OEG monolayers. The chemometrics were then extended to commercial poly(ethylene glycol) (PEG)-based polymer film-coated glass slides.2 Reactive NHS and methoxy-capped (MeO) regions (used for non-fouling) were co-patterned onto these slides using photolithographic methods. NHS patterns are easily imaged with ToF-SIMS/PCA, resolved at high sensitivity.3 NHS-specific protein coupling was imaged and correlated to NHS images by specific coupling of streptavidin on the surface though NHS chemistry. Specific NHS-mediated cell adhesion peptide (RGD) grafting could be imaged, and prompted fibroblasts in serum to attach and proliferate only on the NHS regions. Longer-term cell culture retains high cell-pattern fidelity correlating with chemical imaging of both the NHS and RGD patterns and also lack of cell adhesion to MeO regions. High cross-correlation between various ion-derived ToF-SIMS images is observed, providing sensitive chemical corroboration of pattern chemistry and biological reactivity in complex milieu. This method is unique with important practical impacts for application of new ToF-SIMS surface imaging tools to track and validate pattern fabrication and performance.
1X-ray photoelectron spectroscopy, time-of-flight secondary ion mass spectrometry, and principal component analysis of the hydrolysis, regeneration, and reactivity of N-hydroxysuccinimide-containing organic thin films Fang Cheng, Lara J. Gamble, David W. Grainger, David G. Castner, Anal. Chem. 2007, 79, 8781-8788
2Functionalized poly(ethylene glycol)-based bioassay surface chemistry that facilitates bio-immobilization and inhibits nonspecific protein, bacterial, and mammalian cell adhesion Gregory M. Harbers, Kazunori Emoto, Charles Greef, Steven W. Metzger, Heather N. Woodward, James J. Mascali, David W. Grainger, Michael J. Lochhead, Chem. Mater. 2007, 19, 4405-4414
3Imaging surface immobilization chemistry: correlation with cell patterning on non-adhesive hydrogel thin films Hironobu Takahashi, Kazunori Emoto, Manish Dubey, David G. Castner, David W. Grainger, Adv. Funct. Mater. In press