β-Galactosidase Assay Kit

From： macgene Update： 2008-08-28

b-Galactosidase Assay Kit beta－检测试剂盒

Catalogue：MPK006 Storage: 4－8 ºC

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

Ø Remove the media from the tissue culture plate wells.

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

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

Ø 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

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

Ø Incubation at 37 ºC for 20-60 minutes till yellow shade is observed.

Ø Add 100 ml stop solution (1 M Na2CO3).

Ø Measure O.D. at 420 nm.

Ø 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

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