Rapid quantification of melamine in milk using competitive 1,1'-oxalyldiimidazole chemiluminescent enzyme immunoassay.
A novel competitive 1,1'-oxalyldiimidazole (ODI) chemiluminescent enzyme immunoassay (CLEIA) was developed as a method for rapid and simple screening of melamine in milk. Fat existing in milk acts as an inhibitor in the competitive binding interaction of melamine and anti-melamine in the presence of melamine-conjugated horseradish peroxidase. Thus, the calibration curve and sensitivity of competitive ODI CLEIA for the quantification of melamine in fat free milk were wider and better than those in milk containing fat. However, a centrifuge is not a good method for removing the inhibitor because a portion of the melamine is also removed with the fat. The incubation time (20 min) for the competitive binding interaction of anti-melamine and melamine in 20% milk diluted with PBS buffer of pH 7.4 was longer than that (10 min) in 100% milk even though the sensitivity of the former was better than latter. The limit of detection (1.12 ppb) determined in rapid ODI CLEIA (dynamic range: 3.8-125 ppb) for the quantification of melamine in 20% milk not containing fat was lower than those (6.3 and 9.0 ppb) calculated in relatively time-consuming luminol CLEIA and enzyme-linked immunosorbent assay (ELISA). Also, we expect that ODI-CLEIA (dynamic range: 62.5-2000 ppb) capable of directly quantifying melamine in 100% milk without any pretreatment can be applied as a new and simple method for rapid screening of melamine in milk.
Choi
JooHee
J
Langley High School, 6520 Georgetown Pike, McLean, VA 22101, USA.
Kim
Young-Teck
YT
Lee
Ji Hoon
JH
D000818:Animals; D000906:Antibodies / Q000276:immunology; D002417:Cattle; D006735:Horseradish Peroxidase / Q000737:chemistry / Q000378:metabolism; D007093:Imidazoles / Q000737:chemistry*; D007124:Immunoenzyme Techniques / Q000379:methods*; D008163:Luminescent Measurements / Q000379:methods*; D008892:Milk / Q000737:chemistry*; D014227:Triazines / Q000032:analysis*
C554599:1,1'-oxalyldiimidazole; D000906:Antibodies; D007093:Imidazoles; D014227:Triazines; D006735:Horseradish Peroxidase; C011907:melamine