Format microplate

Allow to react in the wells of 96-format microplates at room temperature in an incubator (shaking) for 24 h. 

The microplate ELISA testis conducted in standard 96-well microplates. A microplate consists of a 12 X 8 grid of wells for test solutions. The three most widely used ELISA formats are immobilized antigen competitive immunoassay, immobilized antibody competitive immunoassay and sandwich immunoassay. " ... 

The number and configuration of wells on a microplate. The most widely used formats are arrays of 96 wells (8x12), or 384 wells (16x24), or 1,536 wells (32x48). 

A very versatile piece of equipment that is affordable for individual laboratories is the microplate reader. This allows multiple samples to be analyzed at once, commonly in a 96-well format, although 384- and 1536-well formats are available. Typical measurements that can be performed include UV-Vis absorbance, fluorescence, or luminescence, allowing a range of assays to be performed, such as cell growth, enzyme kinetics, enzyme stability, or enzyme-linked immunosorbent assay [60-62]. Functionality can be increased by the use of liquid dispensing systems or automatic plate handling. 

Unfortunately, there are no universal methods to detect all types of protein oxidation, because the products formed can be so diverse in nature. However, some forms of protein oxidation can be assayed using chemical modification (Davies et al., 1999 Shacter, 2000). In particular, the formation of carbonyl groups on proteins can be targeted using the reagent 2,4-dinitrophenyl-hydrazine (DNPH). This compound reacts with aldehydes to form 2,4-dinitrophenylhydrazone derivatives, which create chromogenic modifications that can be detected at high sensitivity in microplate assays or Western blot analysis (Buss et al., 1997 Winterbourn et al., 1999). 

To a set of labeled tubes, add 2 ml of OPA reagent (Thermo Fisher) and 200 pi of the appropriate standard or sample. Mix well. If using a microplate format, scale back these quantities 10-fold to fit in the microwells. 

With today s instruments the customer may specify an injector or an automatic reagent dispenser, and a choice of sample formats is available test tube, vial, microplate, Petri dish. The volumes of sample and reagent required to carry out a determination steadily decrease and are typically a few microliters. Sample temperature during the assays is generally controlled. 

Other methods employ a microplate format followed by fast HPLC. Some researchers approach the determination from a different perspective. For example, an alternative method for ionizable substances is the pSol determination based on an acid-base titration.25 26 Kinetic solubility determinations involve determining the concentration of the compound in the buffer of interest when an induced precipitate first appears. 

Chandler, G.T., Cary, T.L., Volz, D.C., Walse, S.S., Ferry, J.L., and Klosterhaus, S.L. Fibronil effects on estuarine copepod (Amphiascus tenuiremid) development, fertility, and reproduction a rapid life cycle assay in 96-well microplate format. Environ. Toxicol Chem., 24(1) 117-124, 2004. 

There are two yeast expression hosts that have an established track record for high-level production of heterologous proteins, namely Saccharomyces cerevisiae and Pichia pastoris. HTP expression screening using microplate formats has been reported for both these yeasts by Lang and coworkers (Holz et ah, 2002, 2003 Boettner et ah, 2002). In both cases standard protocols have been miniaturized with cells cultured in either 1.5 ml cultures in 96-deep-well plates for S. cerevisiae or 2 ml cultures in 24-deep-well plates for P. pastoris. Soluble... 

With a format in which four compounds are tested per 96-well microplate, with robust host cells and rapid assay time, it is possible to assess up to 160 compounds/ week, even without automation. Beyond this throughput, compound supply and data collection become the rate-limiting factors. 

For this purpose, Bruiser has already coupled the microplate stacking device Twister 1 to its microplate reader [22]. In this combination, which is controlled by OPUS software, 40 IR microplates can be measured automatically. To load samples with high throughput, the Microlab 4000 SP autosampler can be used. Both formats (96 and 384) of the Bruker silicon microplates are suitable for automatic loading of various types of samples (proteins, cells, culture media). 

O-Phthaldialdehyde (OPA) is an amine detection reagent that reacts in the presence of 2-mercaptoethanol to generate a fluorescent product (for preparation, see Section 4.1, 2-mercaptoethanol) (Fig. 91). The resultant fluorophore has an excitation wavelength of 360 nm and an emission point at 455 nm. OPA can be used as a sensitive detection reagent for the HPLC separation of amino acids, peptides, and proteins (Fried et al., 1985). It is also possible to measure the amine content in proteins and other molecules using a test tube or microplate format assay with OPA. Detection limits are typically in the microgram per milliliter range for proteins. 

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