Macromolecule-small molecule interactions: A synthetic polymer with greater affinity than serum albumin for small molecules

Abstract

The chemiluminescent detection methods described in this chapter have been successfully applied to the detection of plasmid DNA and genomic DNA in Southern and sequencing protocols. Chemiluminescent organic reactions can be classified in several ways. The majority of these reactions require a critical step that involves the oxidation of a substrate with molecular oxygen or its synthetic equivalent. The oxidation of luminol is a classic example of a chemiluminescent reaction in which the key oxidative step involves hydrogen peroxide and aminophthalhydrazide in the presence of suitable catalysts. The high sensitivity and the simplicity of alkaline phosphatase-conjugated probe (AMPPD) are instrumental in making the chemiluminescent detection of DNA successful in hybridization assays. This detection technique has also been used to detect DNA in dot blots and in situ hybridization experiments along with the protein in enzyme-linked immunosorbent assays (ELISAs) and Western blots. All of the DNA hybridization techniques described in this chapter involve the use of neutral nylon membranes as a solid support onto which the target DNA is immobilized. Subsequently, a complementary probe, labeled with alkaline phosphatase or biotin, is hybridized to the immobilized DNA on the membrane.