Imprinted proteins for determination of ovalbumin

A strategy to design imprinted proteins (IPs) able to detect a glycoprotein (ovalbumin, OVA) was proposed. Glucose oxidase (GOx) was used as a matrix for obtaining the binding cavities with high specificity towards the template protein. The effective method to purify the obtained IPs from the template molecules was developed based on a combination of dialysis and gel filtration. HRP was chosen as a model template to discover the optimal production conditions, and the optimal template concentration (100 µg⋅L^-1) was chosen. The obtained imprinted proteins were characterized by the high adsorption selectivity towards the target protein (the imprinting factor towards OVA and HRP is 4.7). The developed method was transferred to the synthesis of the anti-OVA IPs. The binding properties of these IPs were estimated using the OVA conjugates with low- (FITC) and high- (HRP) molecular weight label molecules. The ability of the synthesized anti-OVA IPs as recognition elements in immunoassay was studied. Under the optimized experimental conditions, the proposed imprinted proteins exhibited a good linear response to OVA in the concentration range of 10-2000 ng⋅mL^-1 with a detection limit of 6 ng⋅mL^-1. The obtained recognition elements were tested for OVA determination in real samples of chicken egg white, and a sample of OVA-free cake spiked by OVA.