Spatiotemporal Three-Dimensional Quantitative Visualization of Macrophage Phagocytosis of Adjuvants In Vivo Using Optical-Resolution Photoacoustic Microscopy

As a crucial component of vaccines, adjuvants rely on the ability of macrophages to phagocytose for immune activation. It is extremely valuable to capture this complex process accurately. In this study, we constructed an optical-resolution photoacoustic microscopy (OR-PAM) system with lateral and axial resolutions of 630 nm and 2.52 μm, respectively, enabling high-resolution three-dimensional visualization and quantitative analysis of the adjuvant phagocytosis process of macrophages. OR-PAM results showed that compared with undifferentiated monocyte macrophages (M0), classically activated macrophages (M1) had profound phagocytic ability to label CpG oligodeoxynucleotides with gold nanoparticles (AuNPs) (CpG–Au) and aluminum adjuvant labeled with lumogallion (Alum–Ga), consistent with the phagocytic observations of AuNPs. In addition, the TNF-α, IL-6, and IL-1β released by M1 were significantly higher than those released by M0 within 2 h after CpG–Au and Alum–Ga stimulation. These results indicated that the size of the adjuvant can affect the phagocytosis efficiency of the macrophages. The findings provide a better understanding of the adjuvant-induced macrophage response and help to develop more efficient and specific vaccines.