title: Development of indirect competitive ELISA using egg yolk-derived immunoglobulin (IgY) for the detection of Gentamicin residues.
abstract: Gentamicin (Gent) is an aminoglycoside antibiotic being used in livestock sector. Gent residues could cause some genetic disorders by nonsense mutations. This study aimed to develop IgY-based ELISA for the detection of Gent in animal products. Gent was conjugated with Bovine serum albumin (BSA) by carbodiimide method for further immunization in the laying chickens. PEG-6000 extraction method was employed to extract IgY from the egg yolk. The titer of anti-Gent-IgY attained the peak of 1:256,000 after the 5(th) booster immunization. Checkerboard titration confirmed that, anti-Gent IgY in 1:2,000 dilution could give an Optical Density (OD) 1.0 at 2 µg mL(-1) of Gent-OVA coating concentration. IgY-based indirect competitive ELISA (Ic-ELISA) showed that, the IC50 value of anti-Gent IgY was 2.69 ng mL(-1) and regression curve equation was y = -16.27x + 56.97 (R(2) = 0.95, n = 3), confirming that, the detection limit (LOD, IC10 value) was 0.01 ng mL(-1). Recoveries from fresh milk, pork and chicken samples were ranged from 69.82% to 94.32%, with relative standard deviation lower than 10.88%. Our results suggested that generated anti-Gent IgY antibodies can be used in routine screening analysis of Gent residues in food samples.
authors.affiliation: a College of Animal Science, Tarim University , Alaer, Aksu Xinjiang , China.|b College of Veterinary Medicine, Northwest A&F University , Yangling, Shaanxi , China.
authors.forename: Jinxin
authors.initials: J
authors.lastname: He
authors.affiliation: a College of Animal Science, Tarim University , Alaer, Aksu Xinjiang , China.
authors.forename: Jianjun
authors.initials: J
authors.lastname: Hu
authors.affiliation: b College of Veterinary Medicine, Northwest A&F University , Yangling, Shaanxi , China.
authors.forename: Diraviyam
authors.initials: D
authors.lastname: Thirumalai
authors.affiliation: b College of Veterinary Medicine, Northwest A&F University , Yangling, Shaanxi , China.
authors.forename: Ruediger
authors.initials: R
authors.lastname: Schade
authors.affiliation: b College of Veterinary Medicine, Northwest A&F University , Yangling, Shaanxi , China.
authors.forename: Enqi
authors.initials: E
authors.lastname: Du
authors.affiliation: a College of Animal Science, Tarim University , Alaer, Aksu Xinjiang , China.|b College of Veterinary Medicine, Northwest A&F University , Yangling, Shaanxi , China.
authors.forename: Xiaoying
authors.initials: X
authors.lastname: Zhang
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000032:analysis*; D002645:Chickens; D004350:Drug Residues / Q000032:analysis*; D004530:Egg Yolk / Q000737:chemistry; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005506:Food Contamination / Q000032:analysis*; D005839:Gentamicins / Q000032:analysis*; D007136:Immunoglobulins / Q000737:chemistry*
chemical_list: D000900:Anti-Bacterial Agents; D005839:Gentamicins; C016512:IgY; D007136:Immunoglobulins
keywords: Drug residues; ELISA; gentamycin; yolk immunoglobulin (IgY)