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AAT Bioquest, Inc.社 マレイミド基定量キット 簡単にタンパク質中のマレイミド基の量を測定

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Sensitive assays of maleimide and thiol groups are required for the efficient conjugation of proteins that are expensive and available only in small amounts. A variety of crosslinking reagents with a maleimide group are widely used for crosslinking proteins to proteins or proteins to other biomolecules. There are few reagents or assay kits available for quantifying the number of maleimide groups that are introduced into the first protein. All the commercial kits have tedious protocols. Our Amplite™ Fluorimetric Maleimide Qutitation kit uses a proprietary dye that has enhanced fluorescence upon reacting with a maleimide. The kit provides a sensitive, one-step fluorimetric method to detect as little as 10 picomoles of maleimide in a 100 µL assay volume (100 nM; Figure 1). The assay can be performed in a convenient 96-well or 384-well microtiter-plate format and easily adapted to automation without a separation step. Its signal can be easily read by a fluorescence microplate reader at Ex/Em = 490/520 nm. Compared to kit 5525, this fluorometric assay is more sensitive, and has less interference from biological samples.

 

Kit Key Features

Broad Application: Can be used for quantifying maleimide group in a variety of molecules such as proteins.
Sensitive: Detect as low as 10 picomoles of maleimide.
Continuous: Easily adapted to automation without a separation step.
Convenient: Formulated to have minimal hands-on time. No wash is required.
Non-Radioactive: No special requirements for waste treatment.

 

Data Analysis

The fluorescence in blank wells (with the assay buffer only) is used as a control, and is subtracted from the values for those wells with the maleimide reactions. A maleimide standard curve is shown in Figure 1.
Note: The fluorescence background increases with time, thus it is important to subtract the fluorescence intensity value of the blank wells for each data point.

Figure 1. N-ethylmaleimide dose response was measured in a 96-well black plate with Amplite™ Fluorimetric Maleimide Quantitation Assay Kit using a NOVOstar microplate reader (BMG Labtech). As low as 0.1 µM (10 picomol/well) of maleimide can be detected with 10 minutes incubation time (n=3).

 

Product NameProduct No.PKG Size
Amplite™ Fluorimetric Maleimide Quantitation Kit 5523 200 assays

 

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