Samples containing subcellular fractions from bacteria or the purified protein were subjected to electrophoresis on the 10 % polyacrylamide gel and transferred onto a polyvinylidene fluoride (PVDF) membrane as reported [22 (link), 47 (link)]. The membranes were then exposed overnight at 4 °C to the chicken polyclonal antibodies against CYP2S1 prepared as described previously [25 (link)] and the antigen–antibody complex was visualized with an alkaline phosphatase-conjugated goat anti-rabbit IgG antibody (1:1428, Sigma-Aldrich, USA) and 5-bromo-4-chloro-3-indolylphosphate/nitrobluetetrazolium as chromogenic substrate [48 (link)].
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