Although photoacoustic computed tomography (PACT) operates with high spatial resolution in biological tissues deeper than other optical modalities, light scattering is a limiting factor. The use of longer near infrared wavelengths reduces scattering. Recently, the rational design of a stable phosphorus phthalocyanine (P-Pc) with a long wavelength absorption band beyond 1000 nm has been reported. Here, we show that when dissolved in liquid surfactants, P-Pc can give rise to formulations with absorbance of greater than 1000 (calculated for a 1 cm path length) at wavelengths beyond 1000 nm. Using the broadly accessible Nd:YAG pulse laser emission output of 1064 nm, P-Pc could be imaged through 11.6 cm of chicken breast with PACT. P-Pc accumulated passively in tumors following intravenous injection in mice as observed by PACT. Following oral administration, P-Pc passed through the intestine harmlessly, and PACT could be used to non-invasively observe intestine function. When the contrast agent placed under the arm of a healthy adult human, a PACT transducer on the top of the arm could readily detect P-Pc through the entire 5 cm limb. Thus, the approach of using contrast media with extreme absorption at 1064 nm readily enables high quality optical imaging in vitro and in vivo in humans at exceptional depths.

Here, we demonstrate that a simple nanoparticle can be used as a contrast agent for biomedical imaging in six different modalities. Near infrared (NIR) fluorescence (FL), NIR-to-NIR upconversion (UC) luminescence, photoacoustic (PA) imaging, Cerenkov luminescence (CL), X-ray computed tomography (CT) and positron emission tomography (PET) were compared in phantom studies and then used for lymphatic mapping in mice. The nanoparticle is self-assembled from just two active imaging components: porphyrin-phospholipid, which coats a core-shell upconversion nanosphere. The porphyrin-phospholipid provides strong absorption for PA, allows for self-assembly-responsive FL, and enables seamless post-labeling with 64Cu for PET and CL. The core-shell provides UC that is not quenched by the porphyrin coating as well as electron density for CT.

Advances in biomedical imaging have spurred the development of integrated multimodal scanners, usually capable of two simultaneous imaging modes. The long-term vision of higher-order multimodality is to improve diagnostics or guidance through analysis of complementary, data-rich, co-registered images. Synergies achieved through combined modalities could enable researchers to better track diverse physiological and structural events, analyze biodistribution and treatment efficacy, and compare established and emerging modalities. Higher-order multimodal approaches stand to benefit from molecular imaging probes and in recent years, contrast agents that have hypermodal characteristics have increasingly been reported in preclinical studies. Given the chemical requirements for contrast agents representing various modalities to be integrated into a single entity, higher-order multimodal agents reported so far tend to be of nanoparticulate form. To date, the majority of reported nanoparticles have included components that are active for magnetic resonance. Herein, we review recent progress in higher-order multimodal imaging agents, which span a range of material and structural classes, that have demonstrated utility in three (or more) imaging modalities.

Pfs25 is a malaria transmission-blocking vaccine antigen candidate, but its apparently limited immunogenicity in humans has hindered clinical development. Here, we show that recombinant, his-tagged Pfs25 can be mixed at the time of immunization with pre-formed liposomes containing cobalt-porphyrin-phospholipid (CoPoP), resulting in spontaneous nanoliposome antigen particleization (SNAP). Antigens are stably presented in uniformly-oriented display via his-tag insertion in the CoPoP bilayer, without covalent modification or disruption of antigen conformation. SNAP immunization of mice and rabbits is well tolerated with minimal local reactogenicity and results in orders-of-magnitude higher functional antibody generation compared to other “mix-and-inject” adjuvants. Serum-stable antigen-binding during transit to draining lymph nodes leads to enhanced antigen uptake by phagocytic antigen presenting cells, with subsequent generation of long-lived, antigen-specific plasma cells. Seamless multiplexing with four additional his-tagged Plasmodium falciparum polypeptides induces strong and balanced antibody production, illustrating the simplicity of developing multi-stage particulate vaccines with SNAP immunization.

Phototherapies involve the irradiation of target tissues with light. To further enhance selectivity and potency, numerous molecularly targeted photosensitizers and photoactive nanoparticles have been developed. Active targeting typically involves harnessing the affinity between a ligand and a cell surface receptor for improved accumulation in the targeted tissue. Targeting ligands including peptides, proteins, aptamers and small molecules have been explored for phototherapy. In this review, recent examples of targeted nanomaterials used in phototherapy are summarized.

Injectable hydrophobic drugs are typically dissolved in surfactants and non-aqueous solvents which can induce negative side-effects. Alternatives like ‘top-down' fine milling of excipient-free injectable drug suspensions are not yet clinically viable and ‘bottom-up' self-assembled delivery systems usually substitute one solubilizing excipient for another, bringing new issues to consider. Here, we show that Pluronic (Poloxamer) block copolymers are amenable to low-temperature processing to strip away all free and loosely bound surfactant, leaving behind concentrated, kinetically frozen drug micelles containing minimal solubilizing excipient. This approach was validated for phylloquinone, cyclosporine, testosterone undecanoate, cabazitaxel and seven other bioactive molecules, achieving sizes between 45 and 160 nm and drug to solubilizer molar ratios 2–3 orders of magnitude higher than current formulations. Hypertonic saline or co-loaded cargo was found to prevent aggregation in some cases. Use of surfactant-stripped micelles avoided potential risks associated with other injectable formulations. Mechanistic insights are elucidated and therapeutic dose responses are demonstrated.

Surfactant-stripped micelles are formed from a commercially available cyanine fluroalkylphosphate salt dye (CyFaP) and used for high contrast photoacoustic imaging in the second near infrared window (NIR-II). The co-loading of Coenzyme Q10 into surfactantstripped CyFaP (ss-CyFaP) micelles improves yield, storage stability and results in a peak absorption wavelength in the NIR-II window close to the 1064 nm output of Nd-YAG lasers used for photoacoustic imaging. Aqueous ss-CyFaP dispersions exhibit intense NIR-II optical absorption, calculated to be greater than 500 at 1064 nm. ss-CyFaP is detected through 12 cm of chicken breast tissue with PAI. In preclinical animal models, ss-CyFaP is visualized in

A gut imaging contrast agent would benefit from the following: 1) being edible and generally regarded as safe; 2) transiting stably through the intestine; and 3) activity in multiple modalities. We found that Chlorophyll-a (Chl), a plant pigment that is ubiquitous in human diets, was amenable to formulation in surfactant-stripped induced frozen micelles (ss-InFroMs). However, Chl ss-InFroM stability was limited in gastric conditions in vitro and in vivo. By demetallating Chl to generate pheophytin-a (Pheo), more stable ss-InFroms were obtained which exhibited complete fecal excretion following oral gavage without systemic absorption. Because ss-InFroms could be concentrated to high concentration during the low-temperature stripping of the loose Pluronic surfactant, Pheo ss-InFroMs were used for non-invasive photoacoustic computed tomography of the gut of mice. Unexpectedly, Pheo (but not Chl) ss-InFroms exhibited strong near infrared fluorescence, further enabling non-invasive fluorescence gut imaging. Since Pheo (but not Chl) could be chelated with exogenous metals, Pheo ss-InFroMs could be seamlessly post-labeled with 64Cu for whole body positron emission tomography. Taken together, these results underscore the potential for Pheo, as an edible material, to be re-formulated for higher-order multimodal gut imaging.