Aptamers are short single-stranded DNA or RNA oligo-
nucleotides that can bind to a wide range of target
molecules with high affinity and specificity. As nucleic
acids, aptamers can undergo denaturation, but the pro-
cess is reversible. As a result of this stability and the
possibility of automated selection of aptamers, these
oligonucleotides are highly promising capture molecules
in microarray formats. In this study, his-tagged proteins
and an aptamer directed against the his-tag were chosen
as a model system. Different factors affect the activity of
aptamers immobilized on a solid support like a microarray
surface. The orientation of the immobilized aptamer plays
an important role in correct aptamer folding and, thus,
in effective binding of the corresponding target. Other
important parameters identified in this work are the
microarrays’ surface charge as well as the length of the
spacer between aptamer and solid support. These param-
eters were investigated systematically, resulting in the
development of an aptamer-based microarray for detection
of his-tagged proteins. The general applicability of the
developed immobilization strategy was demonstrated by
utilization of three different aptamers.