标题：【提供】b－半乳糖苷酶检测试剂盒

b－半乳糖苷酶检测试剂盒

Catalogue：MPK006

Storage: 4－8 ordm;C

Kit Components:

Kit Components

 Chemicals

 Size

 Cat #

I            Extraction Buffer                    10 mL               MC045

 II           Assay Buffer (b-gal)                  60 mL               MC048

 III           Substrate Solution (ONPG)            10 mL               MC049

 IV           Stop Solution                       10 mL               MC007.1

 V            pCMV-b-gal                       10 mg                MN008

 Procedures:

Preparation of Cell Lysates

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Remove the media from the tissue culture plate wells.

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Add 100 μl Extraction Buffer into each well to cover the cells.

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Incubate the plate on shaker platform at room temperature for 5-10 minutes.

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Place the plates on ice and proceed to the next step.

Preparation of Reaction Solution

Calculate how much Reaction Solution you need (200ml per well of 96-well plate). Below is the table for preparation of 100 assays.

Materials

 Quantity

 Note: The kit contains sufficient reagents for 250 assays performed in 96-well tissue culture plate.

* b-Mercaptoethanol was not provided in the kit

 Assay Buffer                           20 mL

 ONPG                                 4 mL

 b-Mercaptoethanol*                     100 mL

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For each well, add 200 ml assay buffer and 5-10 ml of cell lysates.

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Incubation at 37 ordm;C for 20-60 minutes till yellow shade is observed.

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Add 100 ml stop solution (1 M Na2CO3).

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Measure O.D. at 420 nm.

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Calculate the units of b-galactosidase activity using the formula: b-gal (unit) = (O.D.420 nm X 380 X 10) / (minutes of incubation X volume (ml) of cell extract)

Reference:

Herbomel et al. Cell 39:653, 1984

The assay has to be finished in 30 minutes after mixing the working solution and samples.

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