Alternative ELISA Using a RNA Aptamer against Calf Intestinal Alkaline Phosphatase

Alkaline Phosphatase (AP) is extensively used in enzyme-linked immunosorbent assays(ELISA), most requiring expensive and time-intensive reagents to generate (such as AP-conjugated antibodies). In addition, these antibodies may have reduced immunoreactivity when conjugated with AP. We seek to address these problems by developing an alternative AP ligand in the form of an aptamer (nucleic acid binding species) against Calf Intestinal AP (CIAP). This CIAP-aptamer will lead to the development of an alternative reagent that is readily amendable to conjugation, such as conjugation to a primary antibody or perhaps another aptamer. Anti-CIAP aptamers were isolated using Systematic Evolution of Ligands by Exponential Enrichment (SELEX). A diversified pool of RNA (“R50” pool) was used and iterative rounds of selection were performed to wash away the non-binding RNA species and enrich RNA species specifically bound to CIAP. After nine rounds of selection, a reoccurring 14-mer RNA motif (named VDH2.14 and identified ...