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Can the β-Galactosidase Assay System be used with Promega’s five different cell lysis buffers?

We have used the β-Galactosidase Assay System (Cat.# E2000) to compare the activity of β-galactosidase in the presence of five different cell lysis buffers available with Promega’s reporter systems: Reporter Lysis Buffer (RLB), Cell Culture Lysis Reagent (CCLR), Passive Lysis Buffer (PLB), Glo Lysis Buffer (GLB), and Renilla Luciferase Assay Lysis Buffer (RLALB).

Figures 1 and 2 compare β-galactosidase activity in the presence of the five different lysis buffers using either 1M Sodium Carbonate (Figure 1) or 1M Tris base, pH 10 (Figure 2) as a stop solution. All of the lysis buffers worked well in the β-Galactosidase Assay System, but CCLR and GLB required the use of the 1M Tris base, pH 10 stop solution, as a precipitate formed when Sodium Carbonate was used (notice high background values for CCLR and GLB at 0 milliunits of β-galactosidase in Figure 1).

Figure 1. Comparison of β-galactosidase activity in the presence of five different lysis buffers using Sodium Carbonate as a stop solution. The β-Galactosidase Standard provided in the β-Galactosidase Assay System was diluted in 1X RLB to 1 or 0.2 milliunits/µl. An aliquot of 1X lysis buffer (45µl) was then added to triplicate wells of a clear flat-bottomed 96-well plate. An aliquot (5µl) of each standard or 1X RLB alone (background) was then added to the lysis buffers. An equal volume (50µl) of 2X Assay Buffer was then added to each well and the plate was incubated for 45 minutes at 37°C. Sodium Carbonate (150µl) was added and the plate read at 420nm in a SpectraMax^® 250 plate reader. The results are presented as the average of triplicates.

Figure 2. Comparison of β-galactosidase activity in the presence of five different lysis buffers using Tris base, pH 10 as a stop solution. β-Galactosidase reactions were performed as described in Figure 1, except 150µl of 1M Tris base, pH 10 (150µl) was used as the stop solution.

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