2.3f rapid transit microcatheter

PBMCs were purified from whole blood or leukapheresis products via standard density gradient centrifugation and cryopreserved at −140°C. Lymph node mononuclear cells were isolated via mechanical disruption and cryopreserved at −140°C. Non-lymphoid mononuclear cells were isolated using a combination of mechanical disruption and collagenase treatment. TDL was accessed as described previously (Nadolski and Itkin, 2012 (link)). Briefly, a 25-gauge spinal needle was inserted under ultrasound guidance into an inguinal LN on each side of the body, and the oil-based contrast agent ethiodol (Savage Laboratories) was injected under fluoroscopic guidance into each LN. After opacification of the cisterna chyli, access was gained via an anterior transabdominal approach using a 21-gauge or a 22-gauge Chiba needle (Cook Medical Inc.), and a V-18 control guidewire (Boston Scientific) was inserted into the thoracic duct and manipulated cephalad, followed by a 60-cm 2.3F Rapid Transit Microcatheter (Cordis Corp.), which was advanced further into the thoracic duct to aspirate TDL. Aspirated TDL was collected in heparin tubes, and the contrast agent was removed via standard density gradient centrifugation. TDL samples were used directly in flow cytometry experiments or cryopreserved at −140°C.