1 POIWG #38, July 2015 POIWG #38, July 2015 BIOTESC USOC, Switzerland Martin Haag & Jeannine Winkler BIOTESC

2

3 ESA experiment to be performed in Biolab The scientific aim is to study how mammalian cell cultures in vitro react specifically to microgravity Two different cell lines, both of human origin, will be tested; a fibroblast cell line (synthesizes the extracellular matrix and collagen) and an osteoblast cell line (synthesizes bone) Computer-assisted image analysis of PFA-fixed human skin fibroblasts Background

4 Hardware: 6 different Integrated Advanced Experiment Containers (EC) are uploaded at ambient temperatures. Three EC are to be fixated with RNAlater and three with PFA. EC

5 Hardware: Upload Fixative Reservoir Single Sample Reservoir EC Parts of the hardware are uploaded already mounted inside the ECs

6 Hardware: Upload Fixative Reservoir Single Sample Reservoir Med Reservoir/ Fixative Reservoir Double Culture Chamber Type 1 EC EC The other parts of the hardware are uploaded at -80°C inside Type 1 ECs and need to be integrated into the EC on orbit

7 Hardware: Assembly Culture Chamber EC The assembly of one EC is expected to take up to 1 hour to complete Fixative Reservoir Single Sample Reservoir Med Reservoir/ Fixative Reservoir Double

8 Hardware: Disassembly Culture Chamber EC Only the ECs fixed with RNAlater are disassembled and downloaded at -80°C inside Type 1 ECs. The ECs fixed with PFA are downloaded in one piece at +4°C Sample Reservoir

9 Experiment Overview SpX docking Transfer and storage on ISS Experiment start zero time control run 0g run 1g run Recovery period: 37°C, 1 g 38 ± 2 h Fixation/Medium Replacement 37°C, 1 g 38 ± 2 h 37°C, 1 g 38 ± 2 h Disclaimer: Only two ECs fit on the Biolab Rotor at the same time and the experiment will be run sequentially!  Three runs!

10 Experiment Overview SpX docking Transfer and storage on ISS Experiment start zero time control run 0g run 1g run Recovery period: 37°C, 1 g 38 ± 2 h Fixation/Medium Replacement Fixation Active phase: 37°C, 1 g 38 ± 2 h 37°C, 1 g 38 ± 2 h 37°C, 0 g 34 ± 1 h 37°C, 1 g 34 ± 1 h

11 Key points Upload with SpX: -80°C: 56 Culture Chambers (in 28 Type 1 EC), 10 Med Reservoirs (in 10 Type 1 EC) Ambient: 1 Zero EC PFA, 1 Zero EC RNA, 2 Nominal EC PFA, 2 Nominal EC RNA After Docking: Transfer to cold stowage (MELFI -80°C and BIOLAB TCU -20°C) 6 Experiment Containers, processed sequentially in 3 runs: 11 days of experiment! Download with SpX: -80°C: 2 Sample Reservoirs (in 2 Type 1 EC) -20°C: 28 Culture Chambers (in 14 Type 1 EC) +4°C: 1 Zero EC PFA, 2 Nominal EC PFA (all PFA ECs are downloaded fully assembled) Responsibilities NASA: upload/download, transfer to COLUMBUS/SpX, Transfer to MELFI MUSC: for all ground activities, Biolab facility related crew questions BIOTESC: experiment related activities, scientific questions

12 Challenges Very demanding timeline, few break points to pause Once the cells are thawed extremely strict margins to fulfill Planning: days with dense timetable for crew

13 BIOTESC PoC: SPHEROIDS

14 SPHEROIDS – Experiment Overview Scientific Aims: comparing the effects of real microgravity to those induced by mimicked weightlessness on the RPM evaluating the protective effect of VEGF against potential apoptosis post-flight analysis of three dimensional cell assembly, differentiation and apoptosis H/W upload: SpX-7: KUBIK Interface Plates (KIPs) (lost) SpX-8: SPHEROIDS Experiment Containers 2x 8x

15 Impact of SpX-7 Loss and Spheroids Flight Readiness 2 KIPs can be replaced by 1 spare FM and upgrade of 1 QM (upgrade feasible in 3-5 months) 2 KIPs need to be uploaded before the Experiment H/W KUBIK 6 is at the moment not operational due to expired batteries in centrifuge eBox 2 KUBIK eBoxes were lost on SpX-7 Refurbished eBoxes from KUBIK 3 will have to replace the lost ones and be integrated in KUBIK 6 before the experiment

16 KUBIK Overview D1D2 KUBIK Interface Plates (KIPs) – installed at least 1 day before experiment start KUBIK 5 – 39 hours incubation at 37˚C => MELFI cold stowage KUBIK 6 – 8.5 days incubation at 37˚C => change of temperature to 6˚C and cold stowage in KUBIK up to 30 days

17 Experiment Activities -Day 0 Big picture words KUBIK 5 setup & KIP installation, KUBIK 6 setup & KIP instalation -Day 1 Experiment installation in KUBIK 5 and KUBIK 6 -Day 3 Experiment removal from KUBIK 5 & transfer to MELFI -Day 3 – 9 KUBIK 5 datacopy Regular checks of KUBIK 6 temperature (37 ˚C ) -Day 10 KUBIK 6 datacopy Setting KUBIK 6 to 6 ˚C -Day 10 - download Regular checks of KUBIK 6 temperature (6 ˚C ) - Download day Experiment removal from KUBIK 6 and MELFI and packing -1 day after download KUBIK 6 datacopy 2 SPHEROIDS- KUB5-SU SPHEROIDS- KUB6-SU KUB5-KIP- INST KUB6-KIP- INST SPHEROIDS-KUB-PREP SPHEROIDS-EC-INST SPHEROIDS- KUB5-DEINST SPHEROIDS- KUB5- DATACOPY SPHEROIDS- KUB6- DATACOPY INCUBATION at 37˚C 39 hrs INCUBATION at 37˚C 207 hrs SPHEROIDS- COOLTEMP SPHEROIDS- KUB6-DEINST SPHEROIDS-EC-PACK SPHEROIDS- KUB6- DATACOPY 2

Challenges / Watch Outs i.Time constraints are given by the automatic, pre-programmed timeline in the EC: i.Removal of EC before end of experiment run will lead to complete loss of science (no fixation) ii.In case of KUBIK power loss the timeline is interrupted by the duration of the power loss –> de-installation activity (for ECs in KUBIK 5) and/or setting of KUBIK to 6°C activity (ECs in KUBIK 6) to be shifted in the timeline accordingly iii.The experiment run between EC installation and removal/cold stowage activity is 39hrs/207hrs: EC installation and EC removal/cold stowage activity need to be coordinated such that they fit in the crew day ii.In case of KUBIK anomaly the recovery action shall be started as soon as possible: gradual loss of sample viability and complete loss of science after 12 hours

19 BIOTESC PoC: EPO Peake

20 Objectives of EPO Peake To strengthen and to encourage the teaching of science and thus stimulate the curiosity of students and motivate them towards further study of STEM (Science, Technology, Engineering, and Mathematics) and computing and coding subjects To primarily make the students aware that conditions on the ISS are different to those on earth and that in weightlessness, effects can be detected that are very hard to detect on Earth.

21 Overview of EPO Peake Educational Program Operations of Timothy Peake Joint project between UK Space Agency (UKSA) and ESA Four different projects  Seeds in Space o One activity: selfies of Timothy Peake with seeds (during day pass) o Post flight growth difference analysis of seeds that were in space and on ground done by students o Location: Cupola o Inc46 crew time: 30min prime o Task List or hard scheduled

22 Overview of EPO Peake  Classroom Videos Demonstrations (CVD) o One activity o Five demonstrations on physical laws (describing motions and interactions of physical entities on orbit) o Location: Columbus o Inc46 crew time: 2hrs reserve o Task List

23 Overview of EPO Peake  Astro Pi o Credit card sized Raspberry Pi computer which will execute codes programmed by students o Two Astro Pis will be uploaded: one with a standard and one with an infrared camera o Insertion of SD card in Astro Pi to start the computer and execute the students codes o Download of output files from Astro Pi activities stored on SD card, distribution to schools o Locations: Columbus, Cupola, ev. US LAB (three deployments required) o Upload: 45S o Inc46 crew time: 50 min o Hard scheduled prime and reserve  More details in splinter, July 23 rd at 13:00

24 Astro Pi FlowChart  Execution of the same sequence for each deployment

25 Overview of EPO Peake  Generic Videos o 13 videos (script recording) o To provide scientific information, the astronaut’s views on earth or linking experiment activities via visual and audio media o Locations: Cupola, Columbus, US LAB, JEM o Inc46 crew time: 1hr 10 min prime, 6hrs reserve o Task List (gap fillers)

26 Challenges Loss of CVD Kit and Seeds with SpX-7 Re-launch possibilities: - 44S (Prime path), 61P ( ), Orb4

27 Thank you for your attention

28 Located in Hergiswil, Switzerland KUBIK Facility Responsible Center (KUBIK in Columbus since Inc. 23) BIOLAB Facility Support Center Responsible USOC for ESA Thermal Container, EPO Experiments, NightPod BIOTESC Biotechnology Space Support Center