PBMC Isolation, Freezing, and Shipping Protocol

Approximately 10 ml of blood was collected at the field clinic by technicians proficient in blood collection. Detailed procedures [24 (link)] were followed for PBMC isolation, freezing, and storage. Dry shippers (Cryoport, Irvine, CA) that maintain temperature at or below -150°C were used to ship samples from Bangladesh to the United States. Upon arrival to our lab samples were stored in liquid nitrogen until thawed for testing. All samples were analyzed at the end of the study and researchers were blinded to the exposure. As described previously [15 (link)], samples were thawed quickly in a 37°C water bath, washed with warmed media, resuspended in 3 ml fresh media and counted. Cell counts and viabilities were acquired with a Nexcelom Cellometer Auto 2000 Cell Viability Counter using acridine orange and propidium iodide (AO/PI; Nexcelom Bioscience, CS2-0106) according to manufacturer’s directions. Cells with viabilities exceeding 80% were used for testing.