Colorimetric kits

As part of SW-846, the EPA has validated and approved many immunoassay and colorimetric screening methods for a wide range of contaminants, such as petroleum fuels, pesticides, herbicides, PCBs, and explosives. Immunoassay technology uses the property of antibodies to bind to specific classes of environmental pollutants allowing fast and sensitive semiquantitative or qualitative detection. Colorimetric kits are based on the use of chemical reactions that indicate the presence of target analytes by a change in color. Table 3.9 presents a summary of EPA-approved screening methods and their detection capabilities. 

In addition to test kits used in EPA-approved screening methods, a variety of other test kits are available from several manufacturers, for example, immunoassay test kits for BTEX in soil and water and for chlorinated solvents in water colorimetric kits for the detection of lead kits for a wide range of water quality parameter manufactured by Hach Company. 

PBS and gently blotted to remove blood and tissue fluids, then suspended over the lip of a small (250 pi) microcentrifuge tube and punctured with a needle to allow the bile to drain into the tube. Store frozen until assay. There is usually enough material to measure lipid composition (bile acids, cholesterol, phospholipids) with standard colorimetric kits (<1 pi needed for each assay). In addition to biliary cholesterol levels, it is important to take note of bile salt concentrations, since these are the detergents which suspend dietary lipids in micelles and deliver them to the intestinal epithelium for absorption by enterocytes. Differences in bile salt concentration alone could lead to differences in cholesterol absorption. 

Measure bile acids with a standard colorimetric kit using 5-20 xl of the solution from 12. Volumes may need to be adjusted due to interference of color or turbidity in the samples or precipitates that may form when reagents and sample are mixed. 

There are available also several kits for the assay of calcium, in 10 or 20 microliter samples by chelate formation colorimetrically or fluorimetrically. (Pierce Chem. Co., Rockford, 111.). These are read either with the spectrophotometer or by spectrofluorometry. In our experience, while these systems can be used for approximate results, the plot of concentration versus reading curves are rather flat and only an approximation of the values can be obtained. This may be very important late at night or at times when the atomic absorption machine is down, but if the atomic absorption instrument is available it should be used in preference to these procedures. 

The diabetic rats were treated with 18 IU of bovine insulin imbibed into polyacid resins b.i.d. orally using 1 cc syringes and gavage tubes. After 14 days of treatment the rats were sacrificed about 1.5 hours after the last dose. Blood samples were taken and assayed for immunoactive insulin activity (Amersham-Searie RIA kit) and serum glucose levels (glucose oxidase colorimetric assay, Sigma 510 Glucose Kit). 

Normal Rabbits. Six male, white rabbits (2.5 - 3.0 kg) were housed individually. Animals were fasted overnight for 16 hours (with access to water) prior to each experiment to reduce the gastrointestinal content and absorption variability. After treatment with either a control dose or experimental insulin in poly(acrylic acid) resin dose, a one week washout period was required before the next experiment. The protocol called for blood samples to be taken from an indwelling ear catheter at -1, -.5, -.25, +.5, +1, +1.5, +2, +3, +4, +5 and +6 hours. Serum glucose levels were determined by an oxidase colorimetric method using the Sigma 510 Glucose Kit. 

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. 

Colorimetric field tests for TATP and HMTD were described in Section 5 dealing with peroxide-based explosives. This group contains Keinan s PEX [85] (E. Keinan, Personal Communication, February 2006) and the kit developed by Schulte-Ladbeck et al., which involves also a preliminary stage to avoid falsepositive responses by non-explosive peroxides [86]. The color change of molybdenum hydrogen bronze suspension upon reaction with TATP was recommended also as a field test. Exposure of filter paper strips which were soaked in butanol suspension of the molybdenum compound to TATP or hydrogen peroxide vapors rapidly bleaches the blue color [87, 88]. 

C. T. Yuen, P. R. N. Kind, R. G. Price, P. F. G. Praill, and A. C. Richardson, Colorimetric assay for N-acetyl-P-D-glucosaminidase (NAG) in pathological urine using the oo-nitrostyryl substrate the development of a kit and the comparison of the manual procedure with the automated fluorimetric method, Ann. Clin. Biochem., 21 (1984) 295-300. 

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).

SafTest System A set of kits for colorimetric determination of various oxidation indices (POV, ACV, MDA, HNE) with undisclosed reagents . ... 

Calibrate your micropipet by measuring the mass of water it delivers [B. Kratochvil and N. Motkosky, Precision and Accuracy of Mechanical-Action Micropipets, Anal. Chem. 1987, 59, 1064]. A colorimetric calibration kit is available from Artel, Inc., Westbrook, ME 207-854-0860. 

Bachman and Wilcox (1976) found an average cholesterol content of 15.2 mg/100 ml in 356 samples (fat content 3.69%). After separation, 16.9% of the cholesterol was found in the skim milk phase. Patton et al. (1980) did not find any increase in the cholesterol content of skim milk obtained by 24-hr aging of milk at 2-4°C. Cholesterol was determined by nonspecific colorimetric methods in both investigations, which is acceptable since almost all of the sterols are cholesterol. Gen-tner and Haasemen (1979) have analyzed cholesterol in milk enzymatically with a commercially available kit, finding 13 mg/100 ml. The method is very sensitive and is more specific than colorimetric determination, but is not as good as by GLC. Determination of /3-sitosterol by GLC is used to detect adulteration of butter with vegetable oils. 

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). 

Excavation guidance colorimetric field kits correlation, confirm at a... 

Colorimetric bile acid assay kit (Trinity Biotech PLC)... 

Esterases. Acetyl esterase (EC 3.1.1.6) removes acetyl esters from acetylated xylose and short-chain xylo-oligomers. It s polymeracting counterpart, acetyl xylan esterase (EC 3.1.1.72), has a similar activity, but prefers polymeric xylan.244 In addition to acetate-specific enzyme detection kits, HPLC or GC analysis of acetate release from native extracted xylan and chemically acetylated xylan, colorimetric substrates, such as p-nitrophenol acetate and /3-napthyl acetate, or the fluorometric substrate, 4-methylumbelliferyl acetate are also used to assay acetyl esterases.244,253 The third esterase, ferulic acid esterase (EC 3.1.1.73), hydrolyzes the ester bond between ferulic acid or coumaric acid and the arabinose side chain of arabinoxylan. Assays for this activity are usually carried out using starch-free wheat bran or cellulase-treated gramineous biomass as a substrate and monitoring ferulic or coumaric acid released by HPLC or TLC. When preparing enzyme-treated substrates, care must be taken to employ phenolic-acid-esterase-free cellulases.244 Other substrates include methyl and ethyl esters of the phenolic acids, as well as finely ground plant biomass.240,254,255... 

Some of these reactions can also be used for non-radioactive labelling (Isaac 1994) since Tag-polymerase, DNA polymerase I and also terminal transferase can accept compounds such as digoxygenin-dUTP (DIG-dUTP) and the dideoxy derivative DIG-ddUTP as substrates. DIG-labelled DNA can be detected colorimetrically by antibodies coupled to peroxidase using chromogenic substrates. Similar procedures can also be used to label RNA using DIG-UTP with T7- or SP6-RNA polymerases. Digoxygenin labelling kits are commercially available. 

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