Optimization of in situ protein purification. (A) His-tagged FolE and (B) His-tagged Lac repressor. Relative purities of in situ purified proteins were determined using the ratio of fluorescence signals of recombinant proteins versus the fluorescence signals of total protein, as shown in Equation 1 (see text). White and gray bars represent Cu2+/IDA/PEG slides and non-PEG coated Ni2+/NTA slides, respectively. A series of spotting buffers were examined: PBST [10 mM phosphate buffer, 2.7 mM KCl, 140 mM NaCl, 0.05% Tween 20, pH 7.4], PBS [10 mM phosphate buffer, 154 mM NaCl, pH 7.4], TS [50 mM Tris, 300 mM NaCl, 10 mM imidazole, pH 7.8] and SB [100 mM sodium borate, pH 7.8].