Hemozoin-generated vapor nanobubbles for transdermal reagent- and needle-free detection of malaria.

Successful diagnosis, screening, and elimination of malaria critically depend on rapid and sensitive detection of this dangerous infection, preferably transdermally and without sophisticated reagents or blood drawing. Such diagnostic methods are not currently available. Here we show that the high optical absorbance and nanosize of endogenous heme nanoparticles called "hemozoin," a unique component of all blood-stage malaria parasites, generates a transient vapor nanobubble around hemozoin in response to a short and safe near-infrared picosecond laser pulse. The acoustic signals of these malaria-specific nanobubbles provided transdermal noninvasive and rapid detection of a malaria infection as low as 0.00034% in animals without using any reagents or drawing blood. These on-demand transient events have no analogs among current malaria markers and probes, can detect and screen malaria in seconds, and can be realized as a compact, easy-to-use, inexpensive, and safe field technology.

In Planta Response of Arabidopsis to Photothermal Impact Mediated by Gold Nanoparticles.

Biological responses to photothermal effects of gold nanoparticles (GNPs) have been demonstrated and employed for various applications in diverse systems except for one important class - plants. Here, the uptake of GNPs through Arabidopsis thaliana roots and translocation to leaves are reported. Successful plasmonic nanobubble generation and acoustic signal detection in planta is demonstrated. Furthermore, Arabidopsis leaves harboring GNPs and exposed to continuous laser or noncoherent light show elevated temperatures across the leaf surface and induced expression of heat-shock regulated genes. Overall, these results demonstrate that Arabidopsis can readily take up GNPs through the roots and translocate the particles to leaf tissues. Once within leaves, GNPs can act as photothermal agents for on-demand remote activation of localized biological processes in plants.

Laser pulse duration is critical for the generation of plasmonic nanobubbles.

Plasmonic nanobubbles (PNBs) are transient vapor nanobubbles generated in liquid around laser-overheated plasmonic nanoparticles. Unlike plasmonic nanoparticles, PNBs' properties are still largely unknown due to their highly nonstationary nature. Here we show the influence of the duration of the optical excitation on the energy efficacy and threshold of PNB generation. The combination of picosecond pulsed excitation with the nanoparticle clustering provides the highest energy efficacy and the lowest threshold fluence, around 5 mJ cm(-2), of PNB generation. In contrast, long excitation pulses reduce the energy efficacy of PNB generation by several orders of magnitude. Ultimately, the continuous excitation has the minimal energy efficacy, nine orders of magnitude lower than that for the picosecond excitation. Thus, the duration of the optical excitation of plasmonic nanoparticles can have a stronger effect on the PNB generation than the excitation wavelength, nanoparticle size, shape, or other "stationary" properties of plasmonic nanoparticles.

Transient photothermal spectra of plasmonic nanobubbles.

The photothermal efficacy of near-infrared gold nanoparticles (NP), nanoshells, and nanorods was studied under pulsed high-energy optical excitation in plasmonic nanobubble (PNB) mode as a function of the wavelength and duration of the excitation laser pulse. PNBs, transient vapor nanobubbles, were generated around individual and clustered overheated NPs in water and living cells. Transient PNBs showed two photothermal features not previously observed for NPs: the narrowing of the spectral peaks to 1 nm and the strong dependence of the photothermal efficacy upon the duration of the laser pulse. Narrow red-shifted (relative to those of NPs) near-infrared spectral peaks were observed for 70 ps excitation laser pulses, while longer sub- and nanosecond pulses completely suppressed near-infrared peaks and blue shifted the PNB generation to the visual range. Thus, PNBs can provide superior spectral selectivity over gold NPs under specific optical excitation conditions.

Selective and self-guided micro-ablation of tissue with plasmonic nanobubbles.

BACKGROUND: The accuracy, selectivity, and safety of surgical and laser methods for tissue elimination are often limited at microscale. MATERIALS AND METHODS: We developed a novel agent, the plasmonic nanobubble (PNB), for optically guided selective elimination of the target tissue with micrometer precision. PNBs were tested in vitro in the two different models of superficial tumors and vascular plaques. RESULTS: PNBs were selectively generated around gold nanoparticles (delivered to the target tissues) with short laser pulses. Monolayers of cancerous cells and atherosclerotic plaque tissue were eliminated with PNBs with micrometer accuracy and without thermal and mechanical damage to collateral normal tissues. The effect of the PNB was dynamically controlled through the fluence of laser pulses (532 nm, duration 0.5 and 10 ns) and was guided through the optical scattering by PNB. CONCLUSIONS: Plasmonic nanobubbles were shown to provide precise, tunable, selective, and guided ablation of tissue at a microscopic level and could be employed as a new generation of surgical tools.

Short laser pulse-induced irreversible photothermal effects in red blood cells.

BACKGROUND AND OBJECTIVES: Photothermal (PT) responses of individual red blood cells (RBC) to short laser pulses may depend upon PT interactions at microscale. STUDY DESIGN/MATERIALS AND METHODS: A sequence of identical short laser pulses (0.5 and 10 nanoseconds, 532 nm) was applied to individual RBCs, and their PT properties were analyzed at microscale in real time after each single pulse. RESULTS: PT interactions in RBC were found to be localized to sub-micrometer zones associated with Hb that may be responsible for overheating and evaporation at higher optical energies. At sub-ablative energies, a single short laser pulse induced irreversible changes in the optical properties of RBC that stimulated the transition from a heating-cooling response to ablative evaporation in individual erythrocytes during their exposure to subsequent, but identical pulses. CONCLUSION: The PT response of RBCs to short laser pulses of specific energy includes localized irreversible modifications of cell structure, resulting in three different effects: thermal non-ablative response, ablative evaporation, and residual thermal response.

Hot plasmonic interactions: a new look at the photothermal efficacy of gold nanoparticles.

The photothermal (PT) outputs of individual gold nanoparticles (NP) were compared at room (cold) and high transient (hot) temperatures. High temperatures were induced in NPs by a single 0.5 ns laser pulse. All NPs with near-infrared plasmon resonances (rods, shells and bi-pyramids) exhibited a significant decrease in their photothermal output at the resonant wavelengths under high temperature, while non-resonant excitation of the same NPs provided several times higher PT efficacy of the hot NPs. This "inversion" of the PT efficacy of hot plasmonic NPs near their plasmon resonances might have been caused by damping of their resonances due to heating and surface melting. Therefore, photothermal output of plasmonic nanoparticles significantly depends upon their thermal state including the shift in excitation wavelength in hot nanoparticles. In particular, NPs with near-infrared resonances perform several times more efficiently at non-resonant excitation wavelengths rather than at the resonant ones.

Influence of transient environmental photothermal effects on optical scattering by gold nanoparticles.

Transient photothermal phenomena in the environment of light-absorbing plasmonic nanoparticles, heating and evaporation, were shown to influence the optical scattering efficacy of such nanoparticles, when they absorb and scatter the light. The heating of the environment suppresses the optical scattering, while the evaporation enhances the scattering by the nanoparticles. These opposite effects have transient, local, and thermal nature and significantly (more than 10 times) influence the optical contrast of the nanoparticles as shown for gold spheres in water.

Method for disruption and re-canalization of atherosclerotic plaques in coronary vessels with photothermal bubbles generated around gold nanoparticles.

BACKGROUND AND OBJECTIVES: Rapid and safe re-canalization of totally occluded coronary vessels, especially those with the calcified plaques, represent a challenge for cardiology. STUDY DESIGN/MATERIALS AND METHODS: We have suggested to employ photothermal microbubbles (PTMBs) that are generated around injected to plaque or thrombus gold nanoparticles with a short laser pulse for selective mechanical disruption and removal of the plaque tissue and without thermal and mechanical damage to arterial wall. PTMBs were generated in vitro around 30-250 nm gold spheres and with 10 nanoseconds laser pulse at 532 nm in three models: the layer of the living fibroblasts, the epoxy layers, and human arteries with plaques. RESULTS: In all three models, complete removal of the material was observed after 1-10 single laser pulses. The size of cleared zones (20-220 microm) was found to be 500-1,000 times bigger than the size of the nanoparticles applied. PTMB generation did not increase the temperature of the microenvironment outside PTMB and the debris size was below 2 microm. CONCLUSIONS: New proposed method for non-thermal mechanical and localized removal of plaque tissue with PTMB can provide safe and rapid re-canalization of totally occluded and calcified arteries without collateral damage.

Methods for Generation and Detection of Nonstationary Vapor Nanobubbles Around Plasmonic Nanoparticles.

Laser pulse-induced vapor nanobubbles are nonstationary nanoevents that offer a broad range of applications, especially in the biomedical field. Plasmonic (usually gold) nanoparticles have the highest energy efficacy of the generation of vapor nanobubbles and such nanobubbles were historically named as plasmonic nanobubbles. Below we review methods (protocols) for generating and detecting plasmonic nanobubbles in liquids. The biomedical applications of plasmonic nanobubbles include in vivo and in vitro detection and imaging, gene transfer, micro-surgery, drug delivery, and other diagnostic, therapeutic, and theranostic applications.

Retraction of "Tunable Plasmonic Nanoprobes for Theranostics of Prostate Cancer".

[This retracts the article on p. 3 in vol. 1, PMID: 21547151.].

All-in-one processing of heterogeneous human cell grafts for gene and cell therapy.

Current cell processing technologies for gene and cell therapies are often slow, expensive, labor intensive and are compromised by high cell losses and poor selectivity thus limiting the efficacy and availability of clinical cell therapies. We employ cell-specific on-demand mechanical intracellular impact from laser pulse-activated plasmonic nanobubbles (PNB) to process heterogeneous human cell grafts ex vivo with dual simultaneous functionality, the high cell type specificity, efficacy and processing rate for transfection of target CD3+ cells and elimination of subsets of unwanted CD25+ cells. The developed bulk flow PNB system selectively processed human cells at a rate of up to 100 million cell/minute, providing simultaneous transfection of CD3+ cells with the therapeutic gene (FKBP12(V36)-p30Caspase9) with the efficacy of 77% and viability 95% (versus 12 and 60%, respectively, for standard electroporation) and elimination of CD25+ cells with 99% efficacy. PNB flow technology can unite and replace several methodologies in an all-in-one universal ex vivo simultaneous procedure to precisely and rapidly prepare a cell graft for therapy. PNB's can process various cell systems including cord blood, stem cells, and bone marrow.

Intraoperative diagnostics and elimination of residual microtumours with plasmonic nanobubbles.

Failure of cancer surgery to intraoperatively detect and eliminate microscopic residual disease (MRD) causes lethal recurrence and metastases, and the removal of important normal tissues causes excessive morbidity. Here, we show that a plasmonic nanobubble (PNB), a non-stationary laser pulse-activated nanoevent, intraoperatively detects and eliminates MRD in the surgical bed. PNBs were generated in vivo in head and neck cancer cells by systemically targeting tumours with gold colloids and locally applying near-infrared, low-energy short laser pulses, and were simultaneously detected with an acoustic probe. In mouse models, between 3 and 30 residual cancer cells and MRD (undetectable with current methods) were non-invasively detected up to 4 mm deep in the surgical bed within 1 ms. In resectable MRD, PNB-guided surgery prevented local recurrence and delivered 100% tumour-free survival. In unresectable MRD, PNB nanosurgery improved survival twofold compared with standard surgery. Our results show that PNB-guided surgery and nanosurgery can rapidly and precisely detect and remove MRD in simple intraoperative procedures.

Rapid detection and destruction of squamous cell carcinoma of the head and neck by nano-quadrapeutics.

Survival and quality of life remain poor for patients with head and neck squamous cell carcinoma (HNSCC) that cannot be fully resected safely, and form therapy-resistant residual and recurrent tumors. We report novel cell-level technology, quadrapeutics. Quadrapeutics converts surgery, drug, and radiation therapies into on-demand microtreatment that unites the diagnosis and treatment in 1 rapid procedure by using 4 standard components: (1) targeted gold colloids; (2) liposomal drugs; (3) a laser pulse; and (4) radiation, all at safe doses. The therapeutic strength of quadrapeutics increases with cancer aggressiveness. In animal models of a primary and microscopic residual HNSCC, quadrapeutics increased the efficacy of standard chemoradiation therapy by more than 17-fold by using only 3% to 6% of clinical doses of drug and radiation, did not cause side effects, and detected residual microtumors in vivo intraoperatively. Quadrapeutics can be applied to detect and eradicate HNSCC and similar microtumors in a safe and rapid theranostic procedure.

Safety and efficacy of quadrapeutics versus chemoradiation in head and neck carcinoma xenograft model.

Chemoradiation is the strongest anti-tumor therapy but in resistant unresectable cancers it often lacks safety and efficacy. We compared our recently developed cell-level combination approach, quadrapeutics, to chemoradiation therapy to establish pre-clinical data for its biodistribution, safety and efficacy in head and neck squamous cell carcinoma (HNSCC), as a clinically challenging aggressive and resistant cancer. In vitro and in vivo models of four carcinomas were treated with standard chemoradiation and quadrapeutics using identical drug and radiation doses. We applied liposomal cisplatin or doxorubicin, colloidal gold, near-infrared laser pulses and radiation, all at low safe doses. The final evaluation used a xenograft model of HNSCC. Quadrapeutics enhanced standard chemoradiation in vitro by reducing head and neck cancer cell proliferation by 1000-fold, inhibiting tumor growth in vivo by 34-fold and improving animal survival by 5-fold, and reducing the side effects to a negligible level. In quadrapeutics, we observed an "inversion" of the drug efficacy of two standard drugs: doxorubicin, a low efficacy drug for the cancers studied, was two times more efficient than cisplatin, the first choice drug in clinic for HNSCC. The radical therapeutic gain of quadrapeutics resulted from the intracellular synergy of the four components employed which we administered in a specific sequence, while the reduction in the toxicity was due to the low doses of all four components. The biodistribution, safety and efficacy data for quadrapeutics in HNSCC ensure its high translational potential and justify the possibility of clinical trials.

Malaria theranostics using hemozoin-generated vapor nanobubbles.

Malaria remains a widespread and deadly infectious human disease, with increasing diagnostic and therapeutic challenges due to the drug resistance and aggressiveness of malaria infection. Early detection and innovative approaches for parasite destruction are needed. The high optical absorbance and nano-size of hemozoin crystals have been exploited to detect and mechanically destroy the malaria parasite in a single theranostic procedure. Transient vapor nanobubbles are generated around hemozoin crystals in malaria parasites in infected erythrocytes in response to a single short laser pulse. Optical scattering signals of the nanobubble report the presence of the malaria parasite. The mechanical impact of the same nanobubble physically destroys the parasite in nanoseconds in a drug-free manner. Laser-induced nanobubble treatment of human blood in vitro results in destruction of up to 95% of parasites after a single procedure, and delivers an 8-fold better parasiticidal efficacy compared to standard chloroquine drug treatment. The mechanism of destruction is highly selective for malaria infected red cells and does not harm neighboring, uninfected erythrocytes. Thus, laser pulse-induced vapor nanobubble generation around hemozoin supports both rapid and highly specific detection and destruction of malaria parasites in one theranostic procedure.

On-demand intracellular amplification of chemoradiation with cancer-specific plasmonic nanobubbles.

Chemoradiation-resistant cancers limit treatment efficacy and safety. We show here the cancer cell-specific, on-demand intracellular amplification of chemotherapy and chemoradiation therapy via gold nanoparticle- and laser pulse-induced mechanical intracellular impact. Cancer aggressiveness promotes the clustering of drug nanocarriers and gold nanoparticles in cancer cells. This cluster, upon exposure to a laser pulse, generates a plasmonic nanobubble, the mechanical explosion that destroys the host cancer cell or ejects the drug into its cytoplasm by disrupting the liposome and endosome. The same cluster locally amplifies external X-rays. Intracellular synergy of the mechanical impact of plasmonic nanobubble, ejected drug and amplified X-rays improves the efficacy of standard chemoradiation in resistant and aggressive head and neck cancer by 100-fold in vitro and 17-fold in vivo, reduces the effective entry doses of drugs and X-rays to 2-6% of their clinical doses and efficiently spares normal cells. The developed quadrapeutics technology combines four clinically validated components and transforms a standard macrotherapy into an intracellular on-demand theranostic microtreatment with radically amplified therapeutic efficacy and specificity.

Transient enhancement and spectral narrowing of the photothermal effect of plasmonic nanoparticles under pulsed excitation.

The transient 100-fold enhancement and spectral narrowing to 2 nm of the photothermal conversion by solid gold nanospheres under near-infrared excitation with a short laser pulse is reported. This non-stationary effect was observed for a wide range of optical fluences starting from 10 mJ cm(-2) for single nanospheres, their ensembles and aggregated clusters in water, in vitro and in vivo.

Cell-specific multifunctional processing of heterogeneous cell systems in a single laser pulse treatment.

Current methods of cell processing for gene and cell therapies use several separate procedures for gene transfer and cell separation or elimination, because no current technology can offer simultaneous multifunctional processing of specific cell subsets in highly heterogeneous cell systems. Using the cell-specific generation of plasmonic nanobubbles of different sizes around cell-targeted gold nanoshells and nanospheres, we achieved simultaneous multifunctional cell-specific processing in a rapid single 70 ps laser pulse bulk treatment of heterogeneous cell suspension. This method supported the detection of cells, delivery of external molecular cargo to one type of cells and the concomitant destruction of another type of cells without damaging other cells in suspension, and real-time guidance of the above two cellular effects.