Table 4.

Evaluation of suitability of Inflammation assays for SbD hazard testing.

	Submerged Cell Models		ALI Cell Models
Performance Criteria	Submerged Cytokine Release Mono-Culture	Submerged Cytokine Release Co-Culture	ALI Cytokine Release Mono-Cultures	ALI Cytokine Release Co-Cultures
Simplicity and cost	Simple and cost effective.	Simple and cost effective, however creating a co-culture requires more effort and experience than a mono-culture.	Requires specialized exposure equipment and a certain level of expertise.	Requires specialized exposure equipment and a certain level of expertise; creating a co-culture requires more effort and experience than a mono-culture.
Predictivity (Sensitivity and Specificity)	Good correlation with in vivo found [190]. Found to be more sensitive than ALI in several studies [214,216]. Accurate ranking found [208,209].	Combination of immune cell and epithelial cell more predictive than epithelial cell alone [94]. Accurate ranking found [213].	Generally good for primary cells. Lower predictivity of epithelial cell lines. ALI exposures found more sensitive than submerged in several studies [210,211,212].	Co-cultures perform better than two cell types separately [94]. BMDL of this model comes closer to the in vivo BMDL compared to submerged [211].
Robustness	Large inter-laboratory variability for THP-1 cells [100,108], no inter-laboratory data on other cell types.	Not assessed for NMs specifically.	Low reproducibility but similar trends between labs [195]. Low reproducibility improved after protocol optimizations [196].	Low reproducibility but similar trends between labs [195].
Compatibility	NMs may interfere with ELISA [99,118].	NMs may interfere with ELISA [99,118].	Compatible with a wide range of materials, including hydrophobic and low-density NMs. However, NMs may interfere with ELISA [99,118].	Compatible with a wide range of materials, as exposures do not necessarily require a dispersion. However, NMs may interfere with ELISA. Might be more suitable for NMs released form NEPs.
Readiness	No NM-specific standardized protocol available.	No NM-specific standardized protocol available.	No NM-specific standardized protocol available.	No NM-specific standardized protocol available.