Endotoxin contamination of nanoparticle formulations: A concern in vaccine adjuvant mechanistic studies

The increasing awareness of endotoxin contamination has raised important questions during the study of the mechanism of action of the vaccine adjuvants. The endotoxins or lipopolysaccharides (LPS) can con-taminate vaccine formulations contributing to result misinterpretations of the in vitro and in vivo studies. In this short communication, we considered the suitability of the Limulus amebocyte lysate (LAL) assay to quantify chitosan (Chit) nanoparticle (NP) endotoxin contamination to use them in a comparative in vitro immunotoxicology study using both LPS-free (LF) and non-LF Chit NPs. It was shown that chit NPs had a masking effect on endotoxin levels, hampering a reliable conclusion about the effect of their contamina-tion. Neither non-LF nor LF Chit NPs induced the production of ROS in RAW 264.7 cells or IL-6 and TNF-a in PBMCs. The lack of effect of non-LF NPs was not expected and likely due to the NPs masking effect, more evident for higher deacetylation degree Chit. Overall, to prevent questionable results, nanomateri-als should be produced under endotoxin-free conditions.(c) 2023 The Author(s). Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

Automated summary

The increasing awareness of endotoxin contamination has raised important questions during the study of the mechanism of action of the vaccine adjuvants. This short communication discusses the suitability of the Limulus amebocyte lysate assay to quantify chitosan nanoparticle endotoxin contamination and its impact on in vitro immunotoxicology studies. The study found that chit nanoparticles had a masking effect on endotoxin levels, making it difficult to draw reliable conclusions about their contamination effects. Non-LF and LF Chit NPs did not induce the production of certain immune response markers. Overall, nanomaterials should be produced under endotoxin-free conditions to prevent questionable results.