I have 11 mg of commercial PME from orange peel. I would like to use this as a positive control for two different assays which I am currently standardizing. The assays and the buffers used for both are very different. The details for the assays are below.
1. Gel diffusion assay / Pectoplate assay - citrate phosphate buffer (pH 6.8) - is commonly used buffer, no other buffer substitute has been reported. PME diffuses and de-esterifies pectin in the gel and results in increased binding of ruthenium red dye to it during staining procedure. This results in a deepened pink zone around the well in the gel.
2. Bromothymol blue assay - pH -7.5 . In this assay a drop in pH indicates PME activity. Bromothymol changes colour from blue at pH 7.5 to yellow at pH 6.0 This assay uses a buffer with a low buffering capacity since we are monitoring a pH change caused by PME activity on pectin
I would like to use a buffer for the positive control that works for both the above pHs and assays.
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