NASA Bioculture System Capabilities Provides 10 individual cassettes containing hollow fiber bioreactors with media perfusion for a no shear cell culture environment for both adherent and non-adherent cell types, including 3-D tissues and microorganisms Supports automated and manual inflight inoculations for growth initiation, stimulation, sub-culturing, infection, and fixation; also supports automated and manual sample removal for inflight microscopy, analysis and/or fixation Provides programmable temperature settings, o C Includes fluid loops and a gas exchanger for CO 2 and O2 perfusion Supports design flexibility to meet investigator culture chamber, fluid circulation, and automated and manual specimen processing requirements including maintaining cultures over long durations Downlinks individual cassette data; incubation temperature, cold chamber temperature, CO 2, humidity, power, pump activity and event activation The following are examples of cells flown in the Cell Culture Module (precursor to the Bioculture System) on the Shuttle: Skeletal myoblastsHuman osteoblast-like cellsChicken Osteoblasts Human osteoblastsPrimary hamster fibroblastsHuman colon carcinoma cells Human lung fibroblastsChick embryo osteoblastsBovine aortic endothelial cells Stem cells (hematapoetic) Rat skeletal muscle cellsRat osteoblasts Adipose derived Rat glial tumor cellsMesenchymal cells (cartilage) mesenchymal stem cells Elastin heterografts

Media, sample collection, Injection fluid, and sump bags Biochamber Gas Supply Assembly Power Supply Module Cassette Assembly Bioculture System (baseline design) GoreVents Incubation Chamber Cold Chamber Insulation Cassette Base Cassette Front Plate Fan Plenum Cassette Cover Biochamber Oxygenator Heating Loop Manifolds and Valves Media Bag Media Waste Bag Sample Collection Inject Solution Incubation Chamber Cold Chamber Fluid Flow Path Schematic Pump 16 solenoid valves for up to a total of 16 sample/injection bags Solenoid

Bioculture System Specification Nominal Configuration Specimens: Cell culture, tissue culture, 3D cultures, explant cultures, and microbes Docking Station –Power Supply Module –Gas Supply Assembly –Cassette power and gas connectors –ExPRESS Rack mounted 10 Cassette Assemblies in the Docking Station Independent fluidics and environmental control except for the gas supply Each Cassette Assembly contains: –1 Biochamber –Flow Path Tubing –Pump –Up to 16 Solenoid Valves –Up to 16 sample/injection bags –~200 ml Media Bag –1 Sump waste bag –Memory card with set point programming for operational parameters –Cold Chamber –Incubation Chamber –Oxygenator –Heating Loop –Temperature and Humidity Sensors –Three levels of containment o 2 levels provided by the cover and 1 level provided by the flow path Bioculture System Specification for Experiments Environmental Control per Cassette Assembly Incubation Chamber Set Points: Ambient to +42 o C Cold Chamber Set Points: Ambient to +5 o C Temperature is experiment defined and independent for each Cassette Assembly Gas Supply – Shared between all Cassette Assemblies Nominal 5% CO 2, 20% O 2, balance Nitrogen Investigator defined gas composition Shared between all Cassette Assemblies Fluid System per Cassette Assembly Pulsatile, intermittent, and continuous modes Pump: 0 – 15 ml/ min Micro-solenoid diverter valves Biomedical grade tubing Replaceable Media bag and separate waste sump bag 16 replaceable sample/injection bags Programmable delivery schedule or continuous recirculation for medium Programmable schedule for sampling and injections Replaceable Biochamber System compatible with with most fixatives and preservatives Membrane oxygenator using silicone elastomer tubing Heating loop to pre-warm the medium Total of 3 levels of containment The fluid system can be accessed by the Crew on ISS Cassette Assemblies fit in the ISS Microgravity Sciences Glovebox

Examples of Potential ISS Supplemental Equipment for Cell, Tissue, and Microbiology Research ISS Science facilities/equipment: xY9_Yk xY9_Yk Microgravity Glove Box Light Microscopy Module for microscopic analysis Wetlab-2 RNA SmartCycler qRT-PCR System for real time gene expression data downlinked to investigator Small centrifuge for 1-g, fractional-g (e.g., European Modular Cultivation System [EMCS], Kubik), or sample processing (e.g., Cell Biology Experiment Facility [CBEF], Biolab) Refrigerators and freezers

Science program reference documents National Research Council’s 2011 Decadal Survey Report, “Recapturing a Future for Space Exploration: Life and Physical Sciences Research for a New Era” ( NASA Fundamental Space Biology Science Plan ( SBPlan-Signed_508.pdf)

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