Journal: Foods

Article Title: A Novel Method for Antibiotic Detection in Milk Based on Competitive Magnetic Immunodetection

doi: 10.3390/foods9121773

Figure Lengend Snippet: Affinity determination of ( A ) anti-penicillin G specific mAb against penicillin G-BSA conjugate and ( B ) anti-kanamycin specific mAb against kanamycin-BSA conjugate. For both, a microtiter plate was coated with respective antibiotic-BSA conjugate and remaining binding sites were blocked. Subsequently, antibody dilutions in the range from 9.76 ng·mL −1 to 1250 ng·mL −1 were applied and afterward labelled with mouse IgG-specific HRPO-conjugated secondary antibody. Absorbance was measured after 10 min of ABTS substrate incubation. n = 1.

Article Snippet: Anti-kanamycin monoclonal antibody (article number CSB-MA000511I0m) was purchased from Cusabio, Wuhan, China.

Techniques: Binding Assay, Incubation

Journal: Foods

Article Title: A Novel Method for Antibiotic Detection in Milk Based on Competitive Magnetic Immunodetection

doi: 10.3390/foods9121773

Figure Lengend Snippet: Schematic overview of competitive magnetic immunodetection assay procedure for detection of antibiotics in milk. ( A ) Biotinylated monoclonal antibodies are applied into the sample (either sample buffer or whole fat milk (WFM). Throughout pre-incubation, mAbs bind soluble antibiotic molecules. Afterward, the sample is applied onto antibiotic-conjugate coated and blocked polyethylene matrix of immunofiltration columns (IFCs). Here, a competitive binding reaction of monoclonal antibodies between coated antibiotic-conjugate and soluble antigen results in a retention of non-saturated antibodies within the IFC matrix. Subsequently, retained mAbs are magnetically labeled with streptavidin-functionalized magnetic particles (SA-magnetic particle). Finally, retained SA-magnetic particles can be detected and quantified by means of frequency mixing magnetic detection (FMMD). ( B ) Overview about assay steps with incubation times. Light grey arrows represent washing steps with 2 min of duration.

Article Snippet: Anti-kanamycin monoclonal antibody (article number CSB-MA000511I0m) was purchased from Cusabio, Wuhan, China.

Techniques: Immunodetection, Bioprocessing, Incubation, Binding Assay, Labeling

Journal: Foods

Article Title: A Novel Method for Antibiotic Detection in Milk Based on Competitive Magnetic Immunodetection

doi: 10.3390/foods9121773

Figure Lengend Snippet: cMID calibration curves for detection of antibiotics in PBS sample buffer. cMID calibration curves for detection of antibiotics in PBS sample buffer. ( A ) Penicillin cMID calibration curve and ( B ) kanamycin cMID calibration curve. For both, columns were coated with respective antibiotic-BSA conjugate and subsequently blocked. After application of each antibiotic diluted from 0.011 ng·mL −1 up to 1500 ng·mL −1 pre-incubated with 1.2 µg·mL −1 of respective mAb, 60 µg·mL −1 streptavidin-functionalized superparamagnetic particles were applied and rinsed through the column. Readout was done using frequency mixing magnetic detection (FMMD). Each data point represents mean ± SD ( n = 2). Limit of detection and, if possible, maximum of detection are indicated by green square. IC 50 (half maximal inhibitory concentration) is symbolized by red triangle.

Article Snippet: Anti-kanamycin monoclonal antibody (article number CSB-MA000511I0m) was purchased from Cusabio, Wuhan, China.

Techniques: Incubation, Concentration Assay

Journal: Foods

Article Title: A Novel Method for Antibiotic Detection in Milk Based on Competitive Magnetic Immunodetection

doi: 10.3390/foods9121773

Figure Lengend Snippet: cMID calibration curves for detection of antibiotics spiked in whole fat milk. ( A ) Penicillin G calibration curve and ( B ) kanamycin calibration curve. Both antibiotics were diluted from 0.011 ng·mL −1 up to 1500 ng·mL −1 in whole fat milk and were pre-incubated with 1.2 µg·mL −1 of respective biotinylated monoclonal antibodies (mAb). Afterward, the samples were applied onto 3.5 µg·mL −1 antibiotic-BSA conjugate coated and blocked IFC. Finally, 60 µg·mL −1 superparamagnetic particles functionalized with streptavidin were applied and rinsed through the column. Readout was done using FMMD. Each data point represents mean ± SD. For ( A ) penicillin, data points were averaged from two independent calibration curve experiments ( n = 4) and for ( B ) kanamycin each data point represents the mean ± SD of n = 2. Limit of detection and maximum of detection are indicated by green square. IC 50 (half maximal inhibitory concentration) is symbolized by red triangle.

Article Snippet: Anti-kanamycin monoclonal antibody (article number CSB-MA000511I0m) was purchased from Cusabio, Wuhan, China.

Techniques: Incubation, Bioprocessing, Concentration Assay