Colorimetric ELISA

Radioimmunoassays can detect ricin in blood at concentrations as low as 50-100 pg/mL (Godal et al., 1981). Colorimetric ELISA methods can detect ricin spiked into ex vivo human serum or urine samples accurately at levels as low as 1 ng/mL (100 pg/well) with acceptable inter- and intraassay variation (Poli et al., 1994). Chemical luminescence-based ELISA technology improves the detection limit to 0.1-1 ng/mL ricin, but the coefficients of variation may be as high as 50% (Poli et al., 1994). Combination of a sensitive immunological assay with PCR amplification may achieve a limit of detection for ricin at levels as low as 10 fg/mL (Lubelli et al., 2006). 

EHEK-Tek (Organon Teknika) 0157 and H7 Colorimetric ELISA Microplate Health Canada Method MRLP-91... 

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The rate of DNA synthesis can be measured by [ HJthymidine incorporation into the DNA of cells, or in a non-radioactive manner by bromodeoxyuridine (BrdU) incorporation using either a BrdU antibody in a colorimetric ELISA setting or by using FITC-conjugated BrdU and flow cytometry. 

Mickova et al. (2004) have described an enzyme-linked immunosorbent assays (ELISA) with chemiluminescent detection for the determination of carbaryl, carbo-furan and methiocarb. These compounds can also be detected by colorimetric-ELISA methods. The authors have reported very low detection levels of 0.03, 0.007 and... 

The CCD Camera-Based Chemiliimmescence System, luminometer bas Chemiluminescence ELISA and microtiter plate based Colorimetric ELISA were performed as described earlier (27). 

A comparison of the colorimetric and the chemiluminometric assay indicated that the sensitivity of the immunoassay was highly dependent on the antibody used. A prehminaiy comparison revealed that the detection hmit of the tube based chemiluminescent assay was 0.1 ng mL 2,4-D while the detection limit of the colorimetric assay was 22 ng mL This indicated the microformat imaging ELISA to be 20 times more sensitive than the luminometric assay and 4x10 times more sensitive than the colorimetric ELISA. The precision of the luminometric assay was 6% with a workable range between 0.1 and 10 ng mL Similarly for the colorimetric assay the precision was 33% with a workable range between 10 10 ng mL". ... 

Guglielmo-Viret, V, ThulHer, R, 2007. Comparison of an electrochemilumines-cence assay in plate format over a colorimetric ELISA, for the detection of ricin B chain. J. Immunol. Methods 328,70-78. 

Hormones Androgens Testosterone Testosterone EIA kit Salivary assay test Colorimetric EIA kit Testosterone ELISA kit Immunometric Ltd. Salim etric Assay Designs Inc. Oxford Biomedical Research http //www.oxfordbiomed.com/... 

Finally, there are custom two-step quantitation methods such as chromatography or ELISA that require a capture step for isolating the protein and then a quantitation step based on a standard curve of the purified target protein. The preliminary capture step may also concentrate the protein for increased sensitivity. These techniques are typically not available in a commercial kit form and may require extensive method development. They are more labor intensive and complex than the colorimetric or absorbance-based assays. In addition, recovery of the protein from and reproducibility of the capture step complicate validation. Despite these disadvantages, the custom two-step quantitation methods are essential in situations requiring protein specificity. 

Enzyme-linked immunosorbent assay (ELISA) is based on the specific reaction between an antibody and an antigen. One of the reagents in the reaction is labeled with an enzyme that generates a colorimetric product that can be measured with a spectrophotometric device. The color intensity correlates with the concentration of specific antibody and the respective antigen. The reaction can be formatted in various ways in a multiwell plate (microtiter plate) with the common formats being the sandwich assay, the competitive assay, and the direct assay. (See Figure 11.1.)... 

Fig. 12. Effect of spontaneous apoptosis on the activation of caspase-3 in human eosinophils. Eosinophils (2 x 10 /ml) were cultured for 3 h. Cells were lysed and caspase-3 activity was measured by caspase-3 colorimetric assay kit (R D Systems). Enzymatic products were measured at 405 nm with BIOTEK EL340 ELISA microplate reader (BIO-TEK Instrument Inc., Vermont). Human recombinant caspase-3 (5 U) (Calbiochem, California) was used as a positive control. The stimulation index was determined by direct comparison to the level of the normal control. Background readings from cell lysates and buffers were substracted from the readings of both induced and uninduced samples samples before calculating the stimulation index in caspase-3 activity. The differences between control and treated groups were assessed by Student s i-test. P < 0.05 P < 0.001 (Z2).

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Gluten in Foods Colorimetric Monoclonal Antibody ELISA Method Medical Innovations Ltd. Cortecs Diagnostics Transia SA 991.19 132... 

Among the high number of immunoassay techniques, the enzyme-linked immunosorbent assays (ELISAs) combined with a colorimetric end point measurement are the most widely used. These techniques have also been introduced on the market as PCBs ELISA kits by many companies (see Table 25.2). 

Unlike ELISA, which can detect and quantitate host-cell proteins as a group, Western blots detect single protein impurities. Western blot analysis starts with an SDS-PAGE protocol but does not include a final colorimetric staining step. Instead after electrophoresis the proteins are electrotransferred (blotted) from the gel onto a thin membrane. Membranes made of nitrocellulose or PVDF are most often used. Once the transfer is complete, the membrane is incubated with a nonspecific protein solution to saturate and to... 

Table 17-1 Soluble Colorimetric Enzyme/Substrate Systems for Detection of Antigens in the ELISA...

The ability to measure minute amounts of substances such as insulin represented a key breakthrough in the field of endocrinology, and development of RIA was key to the later development of ELISAs. Indeed, ELISA has superseded many RIA measures with its inherent advantages associated with the avoidance of the use of radioactivity and ability to conduct automated analyses with colorimetric signals. 

ELISA is a colorimetric immunoassay which may generally he classified as direct, indirect, sandwich (douhle-antihody) or competitive. 

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