On-line coupling of sol–gel-generated immunoaffinity columns with high-performance liquid chromatography

The paper demonstrates the possibility to use sol–gel-generated immunoaffinity columns as selective sample preparation step in on-line combination with HPLC. In the past sol–gel-generated immunoaffinity columns have only been included in off-line sample preparation schemes. Compared with conventional RP-materials on-line coupling of sol–gel-generated silica matrices with a pore structure designed to retain antibodies poses additional problems caused by their lower pressure tolerance and by the necessity to match the mobile phases not only to take into account the chromatographic properties but also the conformational stability of the antibodies. These problems have been overcome by an on-line system which can be regarded as a prototype for similar systems which exploit the selectivity of sol–gel immunoaffinity columns. The system consists of a sol–gel-generated immunoaffinity column coupled to an RP enrichment column and an analytical column. The practicality of such systems is demonstrated using the example of anti-pyrene immunoaffinity columns applied for the determination of pyrene in aqueous solutions.