Dot‐Elisa for the rapid detection of gentamicin in milk

Abstract

A dipstick dot-ELISA for the detection of gentamicin in milk of dairy cattle is reported for the first time. The test is based on a sandwich ELISA using high affinity monoclonal antibodies to gentamicin. Antibodies were adsorbed to nitrocellulose filters, blocked, dried, and stored for several weeks before use. The dipstick ELISA detected gentamicin at a concentration of 0.1 microgram/ml and produced strongly positive results at 0.2 microgram-0.3 microgram/ml. This ELISA is highly specific and no false positives were detected when tested against various aminoglycoside analogs including streptomycin, kanamycin, bekanamycin, amikacin, neomycin, and tobramycin. Further, the elimination in cow milk of gentamicin residues following intramammary administration of the drug was studied in two dairy cattle using dot-ELISA. Milk gentamicin levels were detected at post injection hours up to 120 hr in each of the two dairy cattle. It therefore, appears that gentamicin residues can still be detected in milk after 5 days using dot-ELISA. Based on the simplicity of performance and the economical nature of the test system, dipstick is recommended as a suitable method for wide scale use in field studies and diagnostic laboratories.

A rapid on-filter immunoassay screen for dexamethasone in equine urine

“…It is of interest to note that the on-filter assay is considerably faster than the dot-ELISA for gentamicin in milk which employs a sandwich format and takes 270 min. 13 The assay is reasonably precise with RSD values below 11%. The spots thus obtained on filters are stable for many months.…”

“…11 In dot-ELISA only a coloured product is formed and positive reactions can be visually assessed against the almost white background of filter disc which also act as a control for nonspecific antibody binding 12 thus expensive ELISA photometers are not necessary. A dot-ELISA with a limit of detection (LOD) of 0.1 µg ml 21 for detection of gentamicin in milk has been described by Ara et al 13 Another versatile dot-ELISA for detecting small peptides has also been described by Sithigorngul et al 14 The conventional dot-ELISAs employ a sandwich format in which strips of cellulose nitrate are impregnated with dots of capture antibody.…”

Sensitive immunochemical approaches for quantitative (FPIA) and qualitative (lateral flow tests) determination of gentamicin in milk

“…Some commercial ELISA kits (Green Spring, cut-off 0.2 ppb) together with a few lateral flow tests are procurable. A dip-stick dot-ELISA for visual detection of gentamicin in milk with a detection limit of 100 mg L À 1 was described by Ara et al [16]. Although the device is reported to be used in the field, the analysis procedure is far to be rapid, requiring several 1 h incubations, together with few manual sample handling and washing steps.…”

Gentamicin release from two‐solution and powder–liquid poly(methyl methacrylate)‐based bone cements by using novel pH method

“…In the last three decades scientists have used a variety of techniques to measure the amount of gentamicin release and the release rate from different kinds of bone cements. These methods include fluorescence polarization immunoassay (FPIA),4, 5 enzyme‐linked immunosorbent assay (ELISA, dot‐ELISA),6 enzyme‐multiplied immunoassay technique (EMIT),7, 8 microbiological assay,9, 10 high performance liquid chromatography with fluorescence detection,11 and spectrophotometric methods 11–13. Radioenzymatic methods (ELISA, EMIT) and radioimmunoassays (FPIA) are reliable but very expensive, whereas chromatographic methods are time consuming 14.…”