Development of a Sensitive Assay to Screen Nanoparticles in vitro for Complement Activation

Area of Science:

• Immunology
• Nanotechnology
• Biomedical Engineering

Background:

• Nanomedicines trigger innate immune responses, causing complement activation, leading to reduced bioavailability and potential adverse effects.
• Current in vitro methods for assessing complement response lack sensitivity and fail to accurately mimic in vivo conditions due to inter-species variations and differing biological responses.
• A critical need exists for reliable in vitro tools to evaluate nanomedicine safety during early development.

Purpose of the Study:

• To develop highly sensitive in vitro assay conditions and sample preparation techniques for assessing complement-mediated responses to nanomedicines.
• To establish an in vitro model that accurately mimics in vivo complement activation relevant to nanomedicine administration.
• To validate the developed assay by correlating in vitro findings with known in vivo observations, specifically regarding zeta potential's impact.

Main Methods:

• Optimized incubation time, nanoparticle dosage, anticoagulant choice, and donor species (blood and blood components) for complement assays.
• Screened various parameters to enhance assay sensitivity and reproducibility for nanomedicine evaluation.
• Validated the developed assay by comparing in vitro complement activation with nanoparticles of varying zeta potentials to established in vivo data.

Main Results:

• Developed optimized assay conditions and sample preparation techniques demonstrating high sensitivity for detecting complement-mediated responses.
• Successfully replicated the in vivo effect of zeta potential on complement activation in vitro.
• Neutral zeta potential nanoparticles showed the least change in complement biomarkers, consistent with in vivo findings, validating the assay's predictive capability.

Conclusions:

• The developed in vitro assay conditions provide a sensitive and reliable tool for assessing nanomedicine-induced complement activation.
• This assay effectively mimics in vivo complement response, enabling accurate safety evaluations in the early stages of nanomedicine development.
• The optimized assay is vital for predicting the safety of intravenously administered nanomedicines, mitigating potential adverse immune reactions.