## 4 Discussion  
Regulatory guidelines for aseptic pharmaceutical production [4; 5] do not provide limits for particles >1.0 \(\upmu\)m. The ISO 14644-1 [35] gives a limit at 832,000 particles/m\({}^{3}\) for this particle size in ISO class 8. This value was not reached in any of the conducted experiments, not even in spiking phases with NCG (highest value: 594,523 particles >1.0 \(\upmu\)m/m\({}^{3}\) equivalent to 71% of the limit value). In Table 4, the statistical evaluation of all data regarding BFPC-counted particles and viables vs. CFU/m\({}^{3}\) are presented. While a weak correlation is shown for viables vs. CFUs on TSA plates, all other comparisons correlate well. Similar correlations of particles to CFUs have also been described by [PERSON] et al. [12] and [PERSON] et al. for particles to viables [20]. A good correlation between the two different MAS systems was expected, as both are based on microbial growth on agar. The different correlations between BFPC-measured viables vs. CFUs on RCS(r) strips or 90 mm plates are interesting. It is most likely that this deviation has its cause in the highly different sampling volumes per minute of the used devices. The absolute CFU counts on plates and strips must be converted to CFU/m\({}^{3}\) by division through the sampled air volume in m\({}^{3}\). For the 90 mm TSA-plates, a total volume of 0.105 m\({}^{3}\) was sampled per plate, which transforms each single colony on the plate into 9.5 CFU/m\({}^{3}\). This is a big lever with respect to the generally low numbers of CFUs found \begin{table}
\begin{tabular}{c c c c} \hline \hline \multicolumn{1}{c}{**Overall Correlation in Experimental Study**} & \multicolumn{1}{c}{**Spearman’s \(\rho\)**} & \multicolumn{1}{c}{\(p\)**} \\ \hline Particles \textgreater{} 1.0 \(\upmu\)m/m\({}^{3}\) & - & Viables/m\({}^{3}\) & 0.864 & \textless{}0.001 \\ Particles \textgreater{} 1.0 \(\upmu\)m/m\({}^{3}\) & - & CFU/m\({}^{3}\) on RCS-TSA strips & 0.778 & \textless{}0.001 \\ Particles \textgreater{} 1.0 \(\upmu\)m/m\({}^{3}\) & - & CFU/m\({}^{3}\) on 90 mm TSA plate & 0.705 & \textless{}0.001 \\ Viables/m\({}^{3}\) & - & CFU/m\({}^{3}\) on RCS-TSA strips & 0.623 & \textless{}0.001 \\ Viables/m\({}^{3}\) & - & CFU/m\({}^{3}\) on 90 mm TSA plate & 0.472 & \textless{}0.001 \\ CFU/m\({}^{3}\) on RCS-TSA strips & - & CFU/m\({}^{3}\) on 90 mm TSA plate & 0.758 & \textless{}0.001 \\ \hline \hline \end{tabular}
\end{table} 
Table 4: Correlation of particles/viables vs. CFUs. Figure 9: Particle/Viable ratio for different garments at all ACRs. on the plates, especially during the 'in operation' phase with a maximum of one CFU found on individual plates. Generally, the collected data from 18 test runs gives a good indication of airborne CFUs in an operational cleanroom at different ACRs. Still, single test run results and single plate counts for a test phase underlie a random factor whether a microbe finds its way into the probe head or not. With the RCS(r) system, the sampling volume was significantly higher. For one TSA strip, 0.683 m\({}^{3}\) of air was sampled. Hence, one CFU on the strip means 1.5 CFU/m\({}^{3}\). With respect to the generally low numbers of CFUs found on the agar plates or strips, this leads to steadier sampling with a lower chance of random effects, as also visible in Figure 6 for RCS(r) vs. IA CFU results in which the RCS(r) achieves steadier results. For the relevant 18G garment in a pharmaceutical cleanroom environment, the median value for the particle/viable ratio was 16.1. (Figure 9). Applying this value to the ISO 8 particle limit of 832,000/m\({}^{3}\), the corresponding limit value for BFPC measured viables would be 832,000/m\({}^{3}\)/16.1 = 51,700/m\({}^{3}\). This value was not reached in any of the 18 experiments. The highest measured value was reached with street clothes in test No. 17 (ACR 10/NCG) with 7880 viable particles/m\({}^{3}\). An exceedance of regulatory limits for CFUs was observed at lower MCP counts.