Biotechnology and Bioengineering, Vol.60, No.2, 137-146, 1998
Detection of functional groups and antibodies on microfabricated surfaces by confocal microscopy
Fluorescence confocal microscopy was used to characterize micron-sized microfabricated silicon particles and planar oxide surfaces after silanization and immobilization of IgG antibody. Surfaces treated with amino- and mercaptosilanes were tested for the presence of amine and sulfhydryl groups by labeling with specific fluorescein probes. In addition, human antibody (IgG) was immobilized to the thiol-coated microparticles using the heterobifunctional crosstinker succinimidyl 4-(N-maleimidolmethyl)-cyclohexane-1-carboxylate. Estimates of the surface density of IgG were consistent with 8.3% of a monolayer of covalently-bound antibody. Confocal images confirmed uniform layers of both silanes and a nti bod ies on the microparticles. The sensitivity limit for the confocal measurements was determined to be as low as 1.5 x 10(-5) fluors per nm(2).