## 2 Materials and Methods
### Mission-Specific Growth Conditions and Procedures  
#### 2.3.1 Micro-6 (and the Reflight Micro-8), Space X CRS-1 and -4, Respectively  
The hardware utilized for these payloads consisted of FPAs, organized eight per Group Activation Pack (GAP) (BioServe Space Technologies). See Figure 1A for a graphic representation of the FPA hardware and Figure 1C for an overview of the experimental design. Each FPA was divided into 2 or 3 chambers, indicated as chamber A, B and C, which were loaded sequentially and compartmentalized by rubber septa. Chamber A was loaded with 2.7 mL 1.2\(\times\) YPD (Sigma Aldrich, St. Louis, MO, USA), containing the antifungal agent AmB (Sigma Aldrich, St. Louis, MO, USA) where indicated. Chamber B was loaded with 0.5 mL _C. albicans_ diluted in water (Low inoculum = 100 cells/mL, High inoculum = 1 \(\times\) 10\({}^{5}\) cells/mL, and are referred to as Low and High, respectively). Chamber C contained 2.7 mL termination medium, consisting of either fixative (8.4% \(v/v\) paraformaldehyde, Polysciences, Inc, Warrington, PA, USA) or fresh medium (1x YPD for  
\begin{table}
\begin{tabular}{c c c c c}  & **SpX CRS-1** & **SpX CRS-4** & **SpX CRS-16** & **SpX CRS-17** \\ \hline  & Micro-6 & Micro-8 & Micro-14/FPA & Micro-14/FEP \\ \hline Scrub delay (days) & 0 & 1 & 1 & 4 \\ Launch date & 8 October 2012 & 21 September 2014 & 5 December 2018 & 4 May 2019 \\ Launch to dock (h) & 70.5 & 79.5 & 69.5 & 42 \\ Dock to sample access (h) \({}^{\ast}\) & 79 & 25 & 30 & 18 \\ Length of mission (days, launch to & 21 & 35 & 39 & 31 \\ splashdown) & 48 & 27 & 120 \({}^{\ast}\) & 264 \({}^{\ast}\) \\ Splashdown to home lab (h) & 70 & 68 & 84.5 & 66 \\ \hline \end{tabular} \({}^{\ast}\) Transition to ISS crew and power resources; * Weather related unberth delay; * Late experiments not requested.  
\end{table}
Table 1: Mission-specific timeline details for four separate experiments.  
samples returned live). The GAPs were maintained at a cool temperature, specifically 4 \({}^{\circ}\)C pre-flight, ambient during launch, and 12 \({}^{\circ}\)C on the ISS, until the scheduled activation at 30 \({}^{\circ}\)C. Following the growth period (44 h for Low inoculum and 30 h for High inoculum), GAPs were chilled to 4 \({}^{\circ}\)C and maintained at that temperature through the remainder of the mission. There were two operational windows during the mission. Samples that were scheduled to be fixed were activated early in the mission (referred to as Early) to avoid any potential decreases in viability due to on-orbit storage. Conversely, cells that were scheduled to be returned live were activated as late as possible in the mission (referred to as Late) so as to retain viability following the growth period and during the transition back to Earth and the home laboratory. During transport back to Earth (splashdown in the Pacific Ocean) and then to the California-based handover location, samples were at ambient temperature (Micro-6) or 4 \({}^{\circ}\)C (Micro-8). Following payload handover to the Implementation Partner (BioServe Technologies) and research team, the samples were transitioned to storage on ice until they arrived at the home laboratory. Viable samples were isolated from the de-integrated hardware and processed the day of arrival. HOBO temperature sensors were included in representative GAPs to retrospectively analyze the temperature profiles and consistency between flight and terrestrial control samples (Figure 2).