title: Group determination of 14-membered macrolide antibiotics and azithromycin using antibodies against common epitopes.
abstract: Erythromycin (ERY), clarithromycin (CLA), roxithromycin (ROX), and azithromycin (AZI) are macrolide antibiotics widely used in livestock and human medicine. Therefore, they are frequently found as pollutants in environmental water. A method based on indirect competitive enzyme-linked immunosorbent assay (ELISA) for group determination of these macrolides in foodstuffs, human biofluids, and water was developed. Carboxymethyloxime of clarithromycin (CMO-CLA) was synthesized and conjugated to bovine serum albumin (BSA) and gelatin to prepare immunogen and coating antigen with advantageous presentation of target epitopes, l-cladinose and d-desosamine, common for these analytes. Antibodies generated in rabbits were capable of recognizing ERY, CLA, and ROX as a group (100-150%), and AZI (12%) and did not cross-react with ERY degradants, which lack antibiotic activity. Assay displayed sensitivity of determination of 14-membered macrolides (IC50=0.13-0.2ng/ml) and low limit of detection (LOD) that was achieved at 0.02 to 0.03ng/ml. It allowed performing analysis of milk, muscle, eggs, bovine serum, water, human serum and urine, and avoiding matrix effect without special pretreatment using simple dilution with assay buffer. For 15-membered macrolide AZI, the corresponding characteristics were IC50=1.6ng/ml and LOD=0.14ng/ml. The recoveries of veterinary and human medicine macrolides from corresponding matrices were validated and found to be satisfactory.
authors.affiliation: Department of Hybridomas, Mechnikov Research Institute for Vaccines and Sera, Russian Academy of Medical Sciences, Moscow 105064, Russia.
authors.forename: Inna
authors.initials: I
authors.lastname: Galvidis
authors.affiliation: Laboratory of Chemical Transformation of Antibiotics, Gause Institute of New Antibiotics, Russian Academy of Medical Sciences, Moscow 119021, Russia.
authors.forename: Gennady
authors.initials: G
authors.lastname: Lapa
authors.affiliation: Department of Hybridomas, Mechnikov Research Institute for Vaccines and Sera, Russian Academy of Medical Sciences, Moscow 105064, Russia. Electronic address: burma68@yandex.ru.
authors.forename: Maksim
authors.initials: M
authors.lastname: Burkin
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000032:analysis* / Q000737:chemistry / Q000276:immunology; D000906:Antibodies; D017963:Azithromycin / Q000032:analysis* / Q000737:chemistry / Q000276:immunology; D001826:Body Fluids / Q000737:chemistry; D002417:Cattle; D017291:Clarithromycin / Q000032:analysis / Q000737:chemistry / Q000276:immunology; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D000939:Epitopes / Q000737:chemistry; D004917:Erythromycin / Q000032:analysis / Q000737:chemistry / Q000276:immunology; D005506:Food Contamination / Q000032:analysis; D006801:Humans; D057230:Limit of Detection; D018942:Macrolides / Q000032:analysis* / Q000737:chemistry / Q000276:immunology; D011817:Rabbits; D015575:Roxithromycin / Q000032:analysis / Q000737:chemistry / Q000276:immunology; D014874:Water Pollutants, Chemical / Q000032:analysis / Q000737:chemistry / Q000276:immunology
chemical_list: D000900:Anti-Bacterial Agents; D000906:Antibodies; D000939:Epitopes; D018942:Macrolides; D014874:Water Pollutants, Chemical; D015575:Roxithromycin; D004917:Erythromycin; D017963:Azithromycin; D017291:Clarithromycin
keywords: Azithromycin; Clarithromycin; Erythromycin; Group-specific immunoassay; Roxithromycin