Two key tests of the robustness of any immobilized enzyme preparation are its long-term stability and its reusability. As a result, we investigated whether immobilized PKA preparations were stable over extended cold storage, and whether they were active through repeated rounds of assay, using VASP phosphorylation assays (Fig. ). After one-month in cold storage (4 ˚C; glycerol storage buffer), our immobilized VASP preparation retained the same level of activity as when freshly prepared (Fig. , Lanes 1 and 2). In addition, the immobilized PKA retained activity through four consecutive rounds of VASP phosphorylation (Fig. , Lanes 2 -5). VASP phosphorylation activity observed in these trails was equivalent to 300 ng of non-immobilized PKAcs, and 50% or more than that observed with 500 ng of non-immobilized PKAcs (Fig. , Lane 7-8).