A total of 113 patientswith clinically diagnosed leptospirosis (WHO,
2003) at San Lazaro Hospital inManila, Philippines,were enrolled in the
study after the August 2012 flood. Plasma, serum, and urine were collected
from each patient. A 10-mL volume of urine was centrifuged at
500 × g for 5 min, and a 1.8-mL aliquot of the supernatant was centrifuged
at 14,000 × g for 10min. The pellet thus obtainedwas resuspended
in 20-μL Tris-EDTA (TE) buffer and boiled for 10 min, yielding urine
pellet 1. The remaining 8.2 mL of supernatant was combined with the
pellet from the first centrifugation, and the mixture was centrifuged at
2095 × g for 10 min. The resultant pellet was resuspended in 20-μL TE
buffer and boiled for 10 min, yielding urine pellet 2. Serum, plasma,
and urine pellet samples were stored at −80 °C until use