H37rv

Immunomacroarray analysis was conducted as follows. Purified LAM from the Mycobacterium tuberculosis strain H37Rv was obtained from BEI Resources (catalog #NR-14848). Anti-LAM antibody was obtained from BEI Resources (NR-13811 LAM mAb clone CS-35). One milliliter of human urine was incubated with 100 μl of nanocage suspension (5 mg/ml, dry weight). Cages were separated from urine by centrifugation, washed with DI water, mixed with 10 μl of Novex 2× Tris-Glycine SDS Sample Buffer (Thermo Fisher Scientific) containing 10% (v/v) 2-mercaptoethanol, and incubated at 100°C for 2 min. The cage suspension was centrifuged (16,100 rcf for 10 min at 25°C), and the supernatant was saved and subjected to detergent removal (HiPPR Detergent Removal Resin Column Kit, Thermo Fisher Scientific) according to the vendor’s instruction and using 100 μl of bead suspension. Aliquots (4 μl) of the resulting purified elution were manually spotted on polyvinylidene difluoride membranes previously activated with methanol and rinsed with DI water. Membranes were allowed to dry at room temperature and then stained using anti-LAM CS-35, horseradish peroxidase–labeled anti-mouse antibody, and enhanced chemiluminescence system (Super-Signal West Dura, Thermo Fisher Scientific).