Dataset: bacteria_FCM_lab
Deployment: MV1008

Samples were collected from the CTD-Rosette PVC Niskin bottles, then preserved with paraformaldehyde (0.5% final concentration). Preserved samples were then flash frozen in liquid nitrogen, and subsequently stored at -80°C until analysis on-shore in batches. Upon thawing, samples were stained with Hoechst 33342 DNA stain (1 ug/mL final concentration), incubated at room temperature in the dark for 1 h, then analyzed using a Beckman-Coulter Altra flow cytometer, equipped with one 488 nm 1W laser and one UV 200 mM laser for excitation. Samples (100 ul) were delivered quantitatively to the instrument using a Harvard Apparatus syringe pump, at a rate of 50 ul per minute. Signals from the DNA detector (450±40 nm) and the forward and side scatter detectors were used to delineate non-pigmented bacteria, in conjunction with the Chlorophyll detector (680±20 nm) and Phycoerythrin detector (575±20 nm) to eliminate chlorophyll and/or phycoerythrin-bearing cells that overlapped (e.g., Prochlorococcus and Synechoccoccus). Calibration beads (0.5 uµm and 1.0 um yellow-green and 0.5 um UV) were used to normalize cellular fluorescence values and assure optimal instrument settings. Raw data files (listmode) were processed in FlowJo software (Treestar Inc.).