Urea

An equal number of cells was lysed in 1 ml of guanidinium lysis buffer
(6M guanidinium-HCl, 0.1M Na₂HPO4 (Sigma)/NaH₂PO4 (Fisher)
pH 8.0, 0.01 M Tris pH 8.0, 5mM imidazole, 10mM β-mercaptoethanol
(Fisher)). Lysates were incubated with 45 μl of Ni2+-NTA-agarose
beads (Thermo Scientific) for 4 hrs at room temperature by end-over-end
rotation. The beads were washed with the following buffers: guanidinium buffer
(6M guanidinium-HCl, 0.1 M Na₂HPO4/NaH₂PO4 pH 8.0, 0.01 M
Tris pH 8.0, 10 mM β-mercaptoethanol); urea pH 8 buffer (8M urea
(Fisher), 0.1 M Na₂HPO4/NaH₂PO4 pH 8.0, 0.01 M Tris pH
8.0, 10 mM β-mercaptoethanol); buffer A (8M urea, 0.1 M
Na₂HPO4/NaH₂PO4 pH 6.3, 0.01 M Tris pH 6.3, 10 mM
β-mercaptoethanol) plus 0.2% Triton-X 100 (Acros); buffer A;
buffer A plus 0.1% Triton X 100. Proteins were eluted with 200 mM
imidazole in 5% SDS (Fisher), 0.15 M Tris pH 6.7, 30% glycerol
(Fisher), 0.72M β-mercaptoethanol. Samples were then subjected to
SDS-PAGE and Western blotting.

3-mercaptophenylboronic acid (3-MPBA) (≥95%), 3-hydroxythiophenol (3-HTP) (≥96%), D-(+)-glucose (≥99.5% GC grade), D-(−)-fructose (≥99%), L-glutamine (≥99%), phorbol 12-myristate 13-acetate (PMA) (≥99%) and sodium hypochlorite (5% w/w) are purchased from Sigma-Aldrich. D-(+)-galactose (≥98%), D-(+)-mannose (≥99%), urea (≥99%), sodium bromide (≥99%), sodium nitroprusside (≥98%), hydrogen peroxide (35% w/w), tert-butyl hydroperoxide (TBHP) (70% w/w), calcium sulfate dehydrate (≥98%) and sodium dihydrogen phosphate (≥98%) are purchased from Alfa Aesar. D-(+)-sucrose (≥99%), creatinine (≥99%) is purchased from TCI. Glucose oxidase from aspergillus and sodium chloride (≥99%) are purchased from VWR. Iron (II) chloride tetrahydrate (≥99%) is purchased from JT Baker. Magnesium sulfate (≥98%) is purchased from EMD Millipore. Potassium chloride (≥99%) is purchased from Acros Organics. Ultrapure water is obtained from a Millipore water system. Citrate-capped gold nanoparticles (60 nm, 5.2×10⁻¹¹ M) are purchased from Nanopartz. Normal human serum is purchased from UTAK Laboratories. Artificial urine is made from 55 mM sodium chloride, 67 mM Potassium chloride, 2.6 mM calcium sulfate, 29.6 mM sodium sulfate, 3.2 mM Magnesium sulfate, 19.8 mM sodium dihydrogen phosphate, 9.8 mM creatinine and 310 mM urea.