Minimum Information for Conducting and Reporting In Vitro Intracellular Infection Assays

Abstract

Bacterial pathogens are constantly evolving to outsmart the host immune system and antibiotics developed to eradicate them. One key strategy involves the ability of bacteria to survive and replicate within host cells, thereby causing intracellular infections. To address this unmet clinical need, researchers are adopting new approaches, such as the development of novel molecules that can penetrate host cells, thus exerting their antimicrobial activity intracellularly, or repurposing existing antibiotics using nanocarriers (i.e., nanoantibiotics) for site-specific delivery. However, inconsistency in information reported across published studies makes it challenging for scientific comparison and judgment of experiments for future direction by researchers. Together with the lack of reproducibility of experiments, these inconsistencies limit the translation of experimental results beyond pre-clinical evaluation. Minimum information guidelines have been instrumental in addressing such challenges in other fields of biomedical research. Guidelines and recommendations provided herein have been designed for researchers as essential parameters to be disclosed when publishing their methodology and results, divided into four main categories: (i) experimental design, (ii) establishing an in vitro model, (iii) assessment of efficacy of novel therapeutics, and (iv) statistical assessment. These guidelines have been designed with the intention to improve the reproducibility and rigor of future studies while enabling quantitative comparisons of published studies, ultimately facilitating translation of emerging antimicrobial technologies into clinically viable therapies that safely and effectively treat intracellular infections.

Cellular uptake and in vitro antibacterial activity of lipid-based nanoantibiotics are influenced by protein corona

“…The cellular viability of RAW264.7 (American Type Cell Culture, ATCC TIB-71™) murine macrophages and A549 cell lines (ATCC CCL-185™) (passage number 5 to 18) in presence of nanoparticles were determined via MTT assay. 21 RAW264.7 were grown in Dulbecco's Modified Eagle Medium (DMEM) while A549 were cultured in F-12K Medium (ThermoFisher Scientific, Scoresby, VIC, Australia), both supplemented with 10% v/v fetal bovine serum (FBS) and 1% v/v penicillin–streptomycin at 37 °C in 5% CO 2 . Cells were seeded in a 96 well plate at a seeding density of 1 × 10 4 cells per well and incubated for 24 h. Following incubation, cells were washed twice and treated with nanoparticles diluted in either media only ( i.e.…”