Patent Application
20150252259
The present invention broadly relates to methods, systems and/or particles for enhancing upconversion luminescence, preferably in particles doped with rare-earth metals.
Upconversion nanocrystals converting, for example, infrared radiation to higher-energy visible luminescence hold a significant promise for applications in bio-detection, bio-imaging, solar cells and 3-D display technologies. Lanthanide-doped upconversion nanocrystals are typically doped with ytterbium Yb3+ sensitiser ions which absorb infrared radiation and non-radiatively transfer sequential excitations to activator ions, such as Erbium (Er3+), Thulium (Tm3+) or Holmium (Ho3+). Traditionally, Er3+ ions which are resonant with Yb3+ ions and have quantum yield of 0.3% for upconversion luminescence, have been intensively investigated for biolabeling and background free imaging. Under low-irradiance excitation Tm3+ as an activator is not as bright as Er3+, however the infrared emissions of Tm3+ at about 802 nm lie in the “biological tissue transparency window”.
In an example upconversion system, luminescent lanthanide ions act as activators (also called emitters) but have a relatively small absorption cross-section to directly absorb incident infrared irradiation. As such, a sensitizer ion with much larger absorption cross-section at infrared (such as Yb) is employed as a type of antenna, which acts to transfer energy non-radiatively to the activators.
Although recent advances in synthesis have led to precise control of upconversion nanocrystal morphology, crystal phase and emission colours, it has remained difficult to achieve strong upconversion luminescence. Attempts to overcome this problem include using noble metal nanostructures to enhance the energy transfer rate by surface plasmons. A fundamental limitation is the concentration of sensitisers and activators cannot be increased beyond a relatively low threshold because this induces a significant decrease in luminescence which is known as “concentration quenching”. The optimised dopant concentrations in NaYF4 host lattices have been determined to be in the range of 0.2˜0.5 mol % for Tm3+ and 20˜40 mol % for Yb3+. These values were established at low irradiance below 100 W/cm2.
The present inventors have developed an understanding of the factors that contribute to concentration quenching in rare-earth doped particles, and have developed methods, systems and/or particles which enable concentration quenching to be minimised or avoided, so that, for example, more than thousands of emitters (and sensitizers) can be embedded into the upconversion nanocrystals, which gives amplified and exceptional brightness.
In various forms, the present invention provides a method, system and/or particles, such as nanocrystals and microcrystals (considered as bulk materials), for enhanced upconversion luminescence, preferably using particles doped with rare-earth elements or metals.
In a first aspect the present invention provides a method for enhancing upconversion luminescence of rare-earth doped particles comprising a host material, a sensitiser and/or an activator, the method comprising subjecting the particles to increased irradiance or a minimum level of irradiance. In a particular example, the activator is present at high concentration and the sensitiser is present at sufficient concentration matching to the activator concentration.
The increased irradiance or the minimum level of irradiance is higher than presently used relatively low irradiance levels of below 100 W/cm2.
Preferably, enhancing upconversion luminescence involves enhancing luminescence intensity and/or brightness and/or upconversion efficiency.
The method may comprise subjecting the particles to an irradiance which is sufficient to overcome or reverse concentration quenching of upconversion luminescence.
The method may comprise subjecting the particles to an irradiance which is sufficient to cause population of an upconversion energy state of the activator.
Preferably, the activator has an intermediate meta stable energy level which accepts resonance energy from the sensitiser excited state level.
The intermediate meta stable energy level may be below the sensitiser excited state level. Alternatively, the intermediate meta stable energy level may be above the sensitiser excited state level.
The particles may be configured to reduce, minimize or exclude quenchers from between the sensitiser and the activator.
The particles may be core-shell particles, wherein the core comprises the host material, highly-doped sensitiser and the activator, and the shell at least partially comprises, or consists of, one or more materials which prevent, retard or inhibit surface quenching.
The method may comprise subjecting the particles to an irradiance (i.e. an increased irradiance or a minimum level of irradiance) of at least about 102 W/cm2, or at least about 103 W/cm2, or at least about 104 W/cm2, or at least about 105 W/cm2, or at least about 106 W/cm2, or at least about 107 W/cm2, or at least about 108 W/cm2, or at least about 109 W/cm2, or at least about 1010 W/cm2, or at least about 1011 W/cm2, or at least about 1012 W/cm2.
The method may comprise subjecting the particles to an irradiance (i.e. an increased irradiance or a minimum level of irradiance) of between about 1×104 and 5×106 W/cm2, or between about 1.6×104 and 2.5×106 W/cm2.
The irradiance may be infrared (or near infrared) irradiance.
The particles may be nanoparticles, microparticles or bulk materials. In some embodiments the particles are nanocrystals or microcystals.
The particles may have an increased or enriched activator concentration. The particles may have an activator concentration of at least about 0.5 mol %, or at least about 1 mol %, or at least about 2 mol %, or at least about 3 mol %, or at least about 4 mol %, or at least about 5 mol %, or at least about 6 mol %, or at least about 7 mol %, or at least about 8 mol %, or at least about 10 mol %, or at least about 12 mol %, or at least about 14 mol %, or at least about 16 mol %, or at least about 18 mol % or at least about 20 mol %.
The activator may be Er3+, Tm3+, Sm3+, Dy3+, Ho+, Eu3+, Tb+, Pr3+ or any other rare-earth metal ion, including combinations thereof. In one embodiment the activator is Tm3+.
The particles may have an increased or enriched sensitiser concentration. The particles may have a sensitiser concentration in the range of about 10 mol % to about 95 mol %, or about 20 mol % to 90 mol %, or about 20 mol % to 80 mol %, or about 30 mol % to 80 mol %, or about 40 mol % to 80 mol %, or about 20 mol % to 40 mol %. In various embodiments the sensitiser is Yb3+, Nd3+ or Gd3+, or a combination thereof.
In the case of a quencher-free system, the concentration level of sensitizers can be increased from the currently used level of 20% to 30% or above, 40% or above, 50% or above, 60% or above, 70% or above, 80% or above, 90% or above.
Where the sensitiser is Yb3+ and the activator is Tm3+, the method may comprise subjecting the particles to an irradiance which is sufficient to cause at least partial population of the 3H4 energy level and/or higher energy levels including the 1G4 and 1D2 energy levels of the Tm3+.
The host material may be, or may comprise, a lanthanide based material, an alkali fluoride, such as for example, NaYF4, NaLuF4, LiLuF4, or KMnF3 or an oxide, such as for example Y2O3, or oxysulfide, such as Gd2O2S.
In one embodiment, there is provided a method for enhancing upconversion luminescence of rare-earth doped particles comprising a host material, a sensitiser and an activator, wherein the particles have an activator concentration of at least about 1 mol %, and the method comprising subjecting the particles to an irradiance of at least about 103 W/cm2.
In another embodiment, there is provided a method for enhancing upconversion is luminescence of rare-earth doped particles comprising a host material, a sensitiser and an activator, wherein the particles have an activator concentration between about 1 mol % and 15 mol %, or between about 2 mol % and 10 mol %, the method comprising subjecting the particles to an irradiance of at least about 103 W/cm2, at least about 104 W/cm2, or at least about 105 W/cm2.
In another embodiment, there is provided a method for enhancing upconversion luminescence of rare-earth doped particles comprising a host material, a sensitiser and an activator, wherein the particles have an activator concentration between about 1 mol % and 20 mol %, or between about 2 mol % and 10 mol %, the method comprising subjecting the particles to an irradiance of at least about 106 W/cm2.
In another embodiment, there is provided a method for enhancing upconversion luminescence of rare-earth doped particles comprising a host material, a sensitiser which is Yb3+ present in a concentration between about 10 mol % and 99 mol %, or between about 20 mol % and 80 mol %, and an activator which is Tm3+ present in a concentration between about 1 mol % and 20 mol %, or between about 1 mol % and 10 mol %, the method comprising subjecting the particles to an irradiance of at least about 105 W/cm2, or at least about 106 W/cm2.
In another embodiment, there is provided a method for enhancing upconversion luminescence of rare-earth doped particles comprising a host material, a sensitiser which is Yb3+ present in a concentration between about 20 mol % and 60 mol %, or between about 20 mol % and 40 mol %, and an activator which is Tm3+ present in a concentration between about 1 mol % and 20 mol %, or between about 4 mol % and 10 mol %, the method comprising subjecting the particles to an irradiance of at least about 106 W/cm2.
In another embodiment, there is provided a method for enhancing upconversion luminescence of rare-earth doped particles comprising a host material, a sensitiser which is Yb3+ present in a concentration between about 20 mol % and 50 mol %, or between about 20 mol % and 40 mol %, and an activator which is Tm3+ present in a concentration between about 1 mol % and 20 mol %; or between about 2 mol % and 10 mol %, the method comprising subjecting the particles to an irradiance of at least about 105 W/cm2, or at least about 106 W/cm2.
In a second aspect, the present invention provides a system comprising rare-earth doped particles comprising a host material, a sensitiser and an activator, and a source of irradiance for subjecting the particles to increased irradiance or a minimum level of irradiance.
In another embodiment, there is provided a system for enhancing upconversion luminescence comprising: rare-earth doped particles comprising a host material, a sensitiser and an activator, wherein the particles have an activator concentration of at least about 1 mol %; and a source of irradiance for subjecting the particles to an irradiance of at least about 103 W/cm2.
The particles may be as defined in the first aspect.
The particles may be subjected to increased irradiance by, and/or in accordance with, the methods of the first aspect.
In a third aspect, the present invention provides rare-earth doped particles comprising a host material, a sensitiser and an activator, wherein the sensitiser is present in a concentration of at least about 20 mol %, and wherein the activator is present in a concentration of at least about 1 mol %.
The host material, activator and sensitiser may be as defined in the first aspect.
In some embodiments the sensitiser is Yb3+ and the activator is Tm3+.
The particles may be nanoparticles, microparticles or bulk materials. In some embodiments the particles are nanocrystals, microcrystals or bulk crystals.
In some embodiments, the sensitiser is present in a concentration of at least about 25 mol %, or at least about 30 mol %, or at least about 40 mol %, or at least about 50 mol %, or at least about 60 mol %, or at least about 70 mol %, or at least about 80 mol %, or at least about 90 mol %, and/or the activator is present in a concentration of at least about 4 mol %, at least about 5 mol %, at least about 10 mol %, at least about 15 mol %, at least about 20 mol %, at least. about 25 mol %, or at least about 30 mol %. Any combinations of the above noted concentrations are contemplated.
The following statements apply to the first, second and third aspects.
The particles may be present in a fibre, for example a suspended-core fibre.
The method, system and particles may find use in detection, sensing, imaging, flow cytometry, photo-dynamic therapy, nanomedicines, solar cell or display applications, fibre amplifier and optical communication, or security printings.
The sensing application may be, for example, a fibre sensing method, such as a fibre dip sensing method. Display applications include TV's and monitors. Nanomedicine applications include drug-carriers and drug-release activators.
In a fourth aspect, the present invention provides a system for capturing upconversion luminescence comprising: a suspended-core optical fibre comprising particles, the particles comprising a host material, an activator and a sensitiser, a laser beam for exciting the particles to produce upconversion luminescence, and a spectrometer for capturing the luminescence.
In another embodiment, there is provided a system for capturing or observing upconversion luminescence comprising: a suspended-core optical fibre including rare-earth doped particles, the particles comprising a host material, a sensitiser and an activator, wherein the particles have an activator concentration of at least about 1 mol %; at least one laser beam as a source of irradiance for subjecting the particles to an irradiance of at least about 103 W/cm2, thereby exciting the particles to produce upconversion luminescence; and a spectrometer for capturing or observing the luminescence.
The particles may be as defined in the first, second or third aspects.
A preferred embodiment of the present invention will now be described, by way of example only, with reference to the accompanying drawings wherein:
Decomposition of the spectra into individual Gaussian peaks. Integrated intensities are given by Iλ where λ is the peak wavelength. Different transitions are indicated in the energy-level scheme (a). For example, the shaded area represents the 3H4
Accordingly, the sensitized photons travel rapidly over large distances within the crystal, thereby significantly increasing the probability of encountering quenchers.
The following are some definitions that may be helpful in understanding the description of the present invention. These are intended as general definitions and should in no way limit the scope of the present invention to those terms alone, but are put forth for a better understanding of the following description.
Throughout this specification and the claims which follow, unless the context requires otherwise, the word “comprise”, and variations such as “comprised”, “comprises” or “comprising”, will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps.
The articles “a” and “an” are used herein to refer to one or to more than one (i.e. to at least one) of the grammatical object of the article. By way of example, “an element” means one element or more than one element.
In the context of this specification, the term “about” is understood to refer to a range of numbers that a person of skill in the art would consider equivalent to the recited value in the context of achieving the same function or result.
In the context of this specification, the terms “rare-earth”, “rare-earth metal”, “rare-earth element” and the like are understood to refer to the following elements and ions thereof: Lanthanum, Cerium, Praseodymium, Neodymium, Promethium, Samarium, Europium, Gadolinium, Terbium, Dysprosium, Holmium, Erbium, Thulium, Ytterbium, Lutetium, Scandium and Yttrium. The ions may be present in the +3 oxidation state, or other oxidation states.
In the context of this specification, the term “sensitiser” is understood to mean an entity that absorbs energy (such as infrared energy) and transfers this energy non-radiatively to the activator.
In the context of this specification, the term “activator” (i.e. emitter) is understood to mean an entity which receives energy from the sensitiser and as a consequence thereof emits upconversion luminescence.
The present inventors have developed an understanding of the factors that contribute to concentration quenching in rare-earth doped particles, and developed methods, systems and particles which enable concentration quenching to be minimised or avoided such that increased activator and sensitiser concentrations may be utilised to optimise luminescence intensity/brightness.
Concentration quenching occurs as a result of the following phenomena. 1) A lack of available sensitised photons per activator ion which inactivates the upconversion luminescence process because statistically most of the activator ions remain in a lower “dark” energy level. 2) Back energy transfer occurring between excited activator ions and sensitiser ions which leads to photons travelling efficiently between sensitiser ions and activator ions thereby rapidly encountering quenchers located at the crystal surface or within the crystal lattice (i.e. crystal defects, this typically happens in high phonon-energy host materials such as glass, or high quenching crystal host, such as the cubic-phase crystals, therefore hexagonal phase fluoride crystals are typically the best host materials). 3) The increased occurrence of sensitised photons encountering quenchers located at the crystal surface or within the crystal lattice at high sensitiser concentrations (for example above 30 mol %). The contribution of these phenomena results in the activator concentration, or the activator and sensitiser concentration, “quenching” upconversion luminescence at relatively low irradiation power.
It has been discovered by the present inventors that upconversion luminescence, by way of example specifically in NaYF4:Yb/Tm nanocrystals, can be significantly enhanced at increased activator concentrations by subjecting the nanocrystals to increased irradiance.
The inventors have surprisingly found that high excitation irradiance can alleviate concentration quenching in upconversion luminescence when combined with higher activator concentration. For example, this allows activator concentration to be increased well above the known level of 0.5 mol % Tm3+ in NaYF4. This leads to significantly enhanced luminescence signals, in one example by up to a factor of about seventy. By using such bright nanocrystals, remote tracking of a single nanocrystal can be achieved, as demonstrated with a microstructured optical-fibre dip sensor by way of illustrative example. This achievement represents a sensitivity improvement of three orders of magnitude over benchmark nanocrystals such as quantum dots.
Without wishing to be bound by theory the inventors postulate that in the case of NaYF4:Yb/Tm nanocrystals elevated irradiance using a 980 nm diode laser beam induces neighbouring Yb3+ sensitisers to transfer sufficient excitation to Tm3+ activators so that each Tm3+ ion receives at least two sequential 980 nm photons. At increased activator concentrations the additional photons sequentially pump the increased Tm3+ present from the 3F4 level (dark state) to the 3H4 energy level or higher energy levels, including the 1G4 and 1D2 levels (visible luminescent states). In addition, back energy transfer from excited Tm3+ ions to Yb3+ sensitisers is avoided because Tm3+ has an intermediate meta stable energy level below the excited state level of Yb3+. Concentration quenching is therefore reversed leading to significantly enhanced upconversion luminescence by virtue of both increased activator concentration and accelerated sensitiser-activator energy transfer rate as a result of a decreased average minimum distance between the sensitisers and activators.
By virtue of overcoming the phenomenon of concentration quenching the present invention enables the use of increased activator and sensitiser concentrations to optimise luminescence intensity/brightness.
As described herein the inventors have observed that at an irradiance power of 2.5×106 W/cm2 nanocrystals comprising 8 mol % Tm3+ resulted in an increase in the integrated upconversion signal by a factor of 1105 compared to the integrated upconversion signal at an irradiance power of 1.6×104 W/cm2. Conveniently, excitation irradiance powers in the range of 104 to 106 W/cm2 are within the normal operating range of various microscopes. In this regard, 104 W/cm2 corresponds to 1 mW over a 10 μm2 cross-sectional area, which is achievable in wide-field microscopy illumination, while 105 W/cm2 corresponds to 1 mW in a 1 μm2 cross-sectional area, which is consistent with laser scanning confocal microscopy.
In one embodiment there is provided a method for enhancing upconversion luminescence of rare-earth doped particles comprising a host material, an enriched concentration of sensitiser and a sufficient concentration level of activator, the method comprising subjecting the particles to increased irradiance or a minimum level of irradiance. The increased or minimum level of irradiance is higher than presently used relatively low irradiance levels. Enhancing upconversion luminescence involves enhancing luminescence intensity and/or brightness and/or upconversion efficiency. The particles are preferably subjected to an irradiance power density which is sufficient to overcome or reverse concentration quenching of upconversion luminescence. The activator preferably has an intermediate meta stable energy level which exists accepting resonance energy from the sensitiser excited state level. In another form, the particles are configured to or designed to reduce, minimize or exclude one or more quenchers from the upconversion system between the sensitizer and the activator. For example, a core-shell particle or system can be provided wherein the core comprises the host material, sensitiser and the activator, and the shell comprises a material which prevents, retards or inhibits surface quenching.
In one embodiment, the particles are subjected to an irradiance, i.e. an increased irradiance or a minimum level of irradiance, which is sufficient to overcome or reverse concentration quenching of upconversion luminescence. In another embodiment, the particles are subjected to an irradiance which is sufficient to cause population of an upconversion energy state of the activator.
In alternative embodiments, where the sensitiser is Yb3+and where the activator is Tm3+ the particles may be subjected to an irradiance which is sufficient to cause population of the 3H4 energy level and/or higher energy levels including the 1G4 and 1D2 energy levels, of Tm3+.
In other embodiments, the particles may be subjected to an irradiance (i.e. an increased irradiance or a minimum level of irradiance) of at least about 102 W/cm2, or at least about 103 W/cm2, or at least about 104 W/cm2, or at least about 105 W/cm2, or at least about 106 W/cm2, or at least about 107 W/cm2, or at least about 108 W/cm2, or at least about 109 W/cm2, or at least about 1010 W/cm2. In some embodiments, the particles may be subjected to an irradiance of at least about 1.6×104 W/cm2, or an irradiance between about 1.0×104 W/cm2 and 5.0×106 W/cm2, or an irradiance between about 1.6×104 W/cm2 and 2.5×106 W/cm2, or an irradiance of about 2.5×106 W/cm2.
Based on the information herein, those skilled in the art will be able to select an appropriate irradiance value for a given activator concentration so as to overcome or reverse concentration quenching. Likewise, those skilled in the art will be able to select an appropriate activator concentration for a given irradiance value so as to overcome or reverse concentration quenching.
The particles described herein are comprised of an inert host material doped with sensitiser(s) and activator(s), and may be referred to as “upconversion particles”, “upconversion nanoparticles” or “upconversion nanocrystals”. The sensitiser and the activator are typically in the form of ions (for example but not necessarily the 3+ oxidation state), and may comprise combinations of different activators and/or combinations of different sensitisers. At least one of the sensitiser(s) and activator(s) is a rare-earth metal, and hence the particles are referred to herein as “rare-earth doped particles”. Typically, both the activator(s) and sensitiser(s) are rare-earth metals.
In various aspects, the activator may be present in a concentration of at least about 0.5 mol %, at least about 1 mol %, at least about 1.5 mol %, at least about 2 mol %, at least about 2.5 mol %, at least about 3 mol %, at least about 3.5 mol %, at least about 4 mol %, at least about 4.5 mol %, at least about 5 mol %, at least about 5.5 mol %, at least about 6 mol %, at least about 6.5 mol %, at least about 7 mol %, at least about 7.5 mol %, least about 8 mol %, at least about 10 mol %, at least about 12 mol %, at least about 14 mol %, at least about 16 mol %, at least about 18 mol %, or at least about 20 mol %.
In some embodiments the activator is present in a concentration between about 1 mol % and 30 mol %, or between about 1 mol % and 25 mol %, or between about 1 mol % and 20 mol %, or between about 1 mol % and 15 mol %, or between about 2 mol % and 30 mol %, or between about 2 mol % and 25 mol %, or between about 2 mol % and 20 mol %, or between about 2 mol % and 15 mol %, or between about 4 mol % and 30 mol %, or between about 4 mol % and 25 mol %, or between about 4 mol % and 20 mol %, or between about 4 mol % and 15 mol %, or between about 4 mol % and 8 mol %.
In other various aspects the activator may be present in a concentration of at least about 2 mol %, at least about 2.5 mol %, at least about 3 mol %, at least about 3.5 mol %, at least about 4 mol %, at least about 4.5 mol %, at least about 5 mol %, at least about 5.5 mol %, at least about 6 mol %, at least about 6.5 mol %, at least about 7 mol %, at least about 7.5 mol %, at least about 8 mol %, at least about 10 mol %, at least about 12 mol %, at least about 14 mol %, at least about 16 mol %, at least about 18 mol %, or at least about 20 mol %. In some embodiments the activator is present in a concentration between about 2 mol % and 30 mol %, or between about 2 mol % and 20 mol %, or between about 2 mol % and 15 mol %, or between about 2 mol % and 8 mol %, or between about 4 mol % and 8 mol %.
Activators that may be used in the particles will be well known to those skilled in the art and include any rare-earth metal ions and combinations thereof, for example Er3+, Tm3+, Ho3+, Dy3+, Eu3+, Tb3+, Sm3+ and Pr3+.
In other various aspects, the sensitiser may be present in a concentration between about 10 mol % and 95 mol %, or between about 15 mol % and 90 mol %, or between about 20 mol % and 90 mol %, or between about 25 mol % and 90 mol %, or between about 15 mol % and 30 mol %, or between about 15 mol % and 25 mol %, or about 20 mol %.
In other various aspects the sensitiser may be present in a concentration between about 20 mol % and 95 mol %, or between about 20 mol % and 80 mol %, or between about 30 mol % and 90 mol %, or between about 35 mol % and 90 mol %, or between about 40 mol % and 90 mol %, or between about 20 mol % and 40 mol %, or between about 50 mol % and 90 mol %, or between about 60 mol % and 90 mol %, or about 20 mol %, or about 40 mol %, or about 60 mol %, or about 80 mol %.
Suitable sensitisers include any rare-earth metal ions and combinations thereof. In one embodiment the sensitiser is Yb3+. In other embodiments the sensitiser could be Gd3+, Nd3+ or Ce3+, or combinations of the sensitisers. For example, the Nd3+ sensitiser can be used as a sensitizer to absorb 800 nm excitation, and the Gd3+ sensitiser can be a sensitizer to absorb UV excitation.
The ratio of the sensitiser to the activator may be between about 1:1 and 40:1, or between about 1:1 and 30:1, or between about 1:1 and 20:1, or between about 1:1 and 10:1, or between about 1:1 and 5:1, or between about 1:1 and 4:1, or between about 1:1 and 3:1.
In embodiments of the invention the particles may be nanoparticles or nanocrystals. In other embodiments of the invention the particles may be microparticles or microcrystals. In other embodiments of the invention the particles may be, or may form, a bulk material.
In some embodiments the particles may comprise increased or enriched amounts of activators and also sensitisers. For example, in various aspects the activator may be present in a concentration of at least about 0.5 mol %, at least about 1 mol %, at least about 1.5 mol %, at least about 2 mol %, at least about 2.5 mol %, at least about 3 mol %, at least about 3.5 mol %, at least about 4 mol %, at least about 4.5 mol %, at least about 5 mol %, or at least about 10 mol %, or at least about 12 mol %, or at least about 14 mol %, or at least about 16 mol %, or at least about 18 mol %, or at least about 20 mol %, or at least about 22 mol %, or at least about 24 mol %, or at least about 26 mol %, or at least about 28 mol %, or at least about 30 mol %, or at least about 35 mol %, or at least about 40 mol %, or at least about 45 mol % or at least about 50 mol %, and/or the sensitiser may be present in a concentration of at least about 20 mol %, or at least about 25 mol %, or at least about 30 mol %, or at least about 35 mol %, or at least about 40 mol %, or at least about 45 mol %, or at least about 50 mol %, or at least about 55 mol %, or at least about 60 mol %, or at least about 65 mol %, or at least about 70 mol %, or at least about 75 mol %, or at least about 80 mol %, or at least about 85 mol % or at least about 90 mol %. Any combinations of the above noted concentrations are contemplated.
In some embodiments the activator may be present in a concentration between about 1 mol % and 30 mol %, or between about 1 mol % and 25 mol %, or between about 1 mol % and 20 mol %, or between about 1 mol % and 15 mol %, or between about 2 mol % and 30 mol %, or between about 2 mol % and 25 mol %, or between about 2 mol % and 20 mol %, or between about 2 mol % and 15 mol %, or between about 4 mol % and 30 mol %, or between about 4 mol % and 25 mol %, or between about 4 mol % and 20 mol %, or between about 4 mol % and 15 mol %, or between about 4 mol % and 8 mol %, and/or the sensitiser may be present in a concentration between about 10 mol % and 95 mol %, or between about 15 mol % and 90 mol %, or between about 20 mol % and 90 mol %, or between about 25 mol % and 90 mol %, or between about 15 mol % and 30 mol %, or between about 15 mol % and 25 mol %, or about 20 mol %. Any combinations of the above noted concentrations are is contemplated.
In other various aspects, the activator may be present in a concentration of at least about 2 mol %, or at least about 6 mol %, or at least about 10 mol %, or at least about 15 mol %, or at least about 20 mol %, or at least about 25 mol %, or at least about 30 mol %, or at least about 35 mol %, or at least about 40 mol %, or at least about 45 mol %, or at least about 50 mol % or at least about 55 mol %, and/or the sensitiser may be present in a concentration of at least about 20 mol %, or at least about 25 mol %, or at least about 30 mol %, or at least about 35 mol %, or at least about 40 mol %, or at least about 45 mol %, or at least about 50 mol %, or at least about 55 mol %, or at least about 60 mol %, or at least about 65 mol %, or at least about 70 mol %, or at least about 75 mol %, or at least about 80 mol %, or at least about 85 mol % or at least about 90 mol %. Any combinations of the above noted concentrations are contemplated.
In other embodiments the activator is present in a concentration between about 2 mol % and 30 mol %, or between about 2 mol % and 15 mol %, or between about 2 mol % and 8 mol %, or between about 4 mol % and 8 mol %, and/or the sensitiser is present in a concentration between about 20 mol % and 95 mol %, or between about 20 mol % and 80 mol %, or between about 30 mol % and 90 mol %, or between about 35 mol % and 90 mol %, or between about 40 mol % and 90 mol %, or between about 20 mol % and 40 mol %, or between about 50 mol % and 90 mol %, or between about 60 mol % and 90 mol %, or about 20 mol %, or about 40 mol %, or about 60 mol %, or about 80 mol %. Any combinations of the above noted concentrations are contemplated.
Suitable host materials will be familiar to those skilled in the art and include any materials having a low phonon energy level and minimal internal quenchers. For example, the host material preferably has a phonon energy level below about 750 cm−1, or below about 500 cm−1, or below about 400 cm−1, or below about 370 cm−1.
Suitable host materials include, but are not limited to, alkali fluorides, such as NaGdF4, NaYF4, LiYF4, NaLuF4 and LiLuF4, KMnF3, and oxides, such as Y2O3. Mixtures of these materials are also contemplated. In one embodiment, the host material is NaYF4. Where the particles are crystalline the NaYF4 may be hexagonal phase, or any other crystal phase.
Once sensitised by the sensitiser, photons are primarily transferred to either activators or neighbouring sensitisers. Consequently, photons will either be transferred to an activator leading to upconversion and resultant luminescence emission, or alternatively encounter a quencher. In some examples, quenchers are populated primarily on the crystal surface due to the large surface to volume ratio, but also exist internally in the form of crystal defects which are dependent on phonon energy levels. Where the sensitiser concentration exceeds 30 mol % for example, the chance of sensitised photons encountering quenchers is significantly increased thereby contributing to concentration quenching. A further contribution to concentration quenching occurs via back energy transfer, which is possible when the activator has an excited meta stable state that is above, or equal to, the sensitiser excited state level (see
Accordingly, in one embodiment the combination of activator and sensitiser is chosen such that a meta stable energy level of the activator exists below the sensitiser excited state level so that back energy transfer from the activator to the sensitiser is minimised or prevented from occurring. Such activators may be referred to as “trap activators” in the sense that sensitised photons cannot undergo back energy transfer to the sensitiser, and are in effect “trapped” by the activator. Because such photons travel only within a limited space in the particle (i.e. from a sensitiser to an activator), the chance of encountering a quencher is minimised (see
In other embodiments the sensitiser, activator and host material are protected against surface quenchers by a shell, such that the particles are core-shell particles wherein the core comprises the activator, the sensitiser and the host material, and the shell comprises, or consists of, a material which prevents, retards or inhibits surface quenching. The shell may partially or completely encapsulate the core. Preferably, the shell comprises or consists of the same material as the host material, but without the rare-earth metal dopants. In the case of crystals, this avoids the need for phase matching.
The presence of a protective shell permits the use of “relay activators” in the particles, i.e. those activators having a meta stable energy level of the activator that is equal to, below, or approximately the same as the sensitiser excited state level. An example of core-shell particles of this type are particles having a core comprising NaYF4Yb:Er and a NaYF4 shell.
A protective shell may also be employed where a meta stable energy level of the activator exists below the sensitiser excited state level. An example of a core-shell particle of this type is depicted in
In embodiments of the invention the activator concentration of the particles and the irradiance may be chosen depending on the particular application, such as the type of emission desired (see
The luminescence decay lifetimes of the particles may be modulated by varying the concentrations of the activator and the sensitiser. The method, system and particles described herein may therefore find application in time-domain multiplexing coding and decoding.
The methods described herein for optimisation of upconversion luminescence make it possible to significantly extend the detection limit of the particles in advanced imaging and sensing applications, such as for example fibre dip sensors. The detection limits of fluorescent quantum dots in such fibres are in the range of about 10 pM and Er3+ upconversion nanocrystals are in the range of about 660 fM due to the competing autofluorescence background from the fibre itself. The inventors have found that by using 4 mol % Tm3+ upconversion nanocrystals it is possible to enhance the upconversion signal via increased activator concentration and to avoid the fibre autofluorescence problem by monitoring several distinct emission peaks of Tm3+ as shown in
In another embodiment there is provided a system for capturing upconversion luminescence comprising: a suspended-core optical fibre comprising particles, the particles comprising a host material, an activator and a sensitiser, a laser beam for exciting the is particles to produce upconversion luminescence, and a spectrometer for capturing the luminescence. The laser beam may subject the particles to an irradiance value or values as defined in accordance with the first aspect. The particles may be as defined in accordance with the first, second or third aspects.
A system in accordance with one embodiment is shown in
The invention will now be described in more detail, by way of illustration only, with respect to the following examples. The examples are intended to serve to illustrate this invention and should in no way be construed as limiting the generality of the disclosure of the description throughout this specification.
Hexagonal-phase NaYF4 nanocrystals with Tm3+ concentrations in the range 0.2-8 mol % and co-doped with 20 mol % Yb3+ were synthesised (see
Upconversion NaYF4:Yb,Tm nanocrystals were synthesized using organometallic methods described previously (see Liu, Y. S. et al. A Strategy to Achieve Efficient Dual-Mode Luminescence of Eu3+ in Lanthanides Doped Multifunctional NaGdF4 Nanocrystals. Adv Mater 22, 3266 (2010); and Wang, F. et al. Simultaneous phase and size control of upconversion nanocrystals through lanthanide doping. Nature 463, 1061-1065, (2010)). Briefly, 5 ml of a methanolic solution of LnCl3 (1.0 mmol, Ln=Y, Yb, Tm/Er) was magnetically mixed with 6 ml OA and 15 ml ODE in a three-neck round-bottom flask. The resulting mixture was heated at 150 ° C. under argon flow for 30 min to form a clear light yellow solution. After cooling down to 50 ° C., 10 mL of a methanolic solution containing 0.16 g NH4F and 0.10 g NaOH was added with vigorous stirring for 30 min. Then, the slurry was slowly heated and kept at 110 ° C. for 30 min to remove methanol and residual water. Next, the reaction mixture was protected with an argon atmosphere, quickly heated to 305° C. and maintained for 1.5 h. The products were isolated by adding ethanol and centrifugation without size-selective fractionation. On occasions the final NaYF4:Yb,Tm nanocrystals were redispersed in cyclohexane with 5 mg/ml concentration after washing with cyclohexane/ethanol.
For characterisation, powder X-ray diffraction (XRD) patterns were obtained on a PANalytical X'Pert Pro MPD X-ray diffractometer using Cu Kul radiation (40 kV, 40 mA, λ=0.15418 nm). Transmission electron microscope (TEM) measurements were performed using a Philips CM10 TEM with Olympus Sis Megaview G2 Digital Camera. The samples for TEM analysis were prepared by placing a drop of a dilute suspension of nanocrystals onto formvar-coated copper grids (300 mesh). The XRD patterns are shown in
A single-mode 980 nm diode laser beam was launched into a suspended-core fibre (see
To quantify the analysis above in Example 2 a matrix of power-dependent (1.6×104 up to 2.5×106 W/cm2) luminescence spectra from six samples of upconversion nanocrystals with Tm3+ concentrations ranging from 0.2′mol % to 8 mol % were collected. With, reference to the simplified excited-state levels in
The selected evolution of spectra for 0.5 mol %, 4 mol %, and 8 mol % Tm3+ as a function of excitation reveals the weight for multiple emission peaks (see
To further explore the factors that contribute to upconversion enhancement,
In order to establish the potential of Tm3+ upconversion nanocrystals as fluorescent probes for trace-molecular detection, NaYF4:Yb/Tm (20/4 mol %) nanocrystals in cyclohexane at various dilutions were introduced into microstructured fibres, as described. above. The Tm3+ emission was clearly detectable at a level of 5 ng/mL, corresponding to 39 fM nanocrystals in a 20 nL suspension (which is equivalent to approximately 635 nanocrystals distributed along about a 12 cm long fibre sensor) as shown in
To further investigate the detection limit, 8 mol % Tm3+ nanocrystals were diluted to 3.9 fM. Interestingly, a digitized signal of ˜30 counts was observed (as background noise), ˜250 counts (220 net counts) and ˜470 counts (440 net counts) for the 802 nm emission, as shown in
Confocal microscopes, though widely used in cell biology labs, only give optical resolution approaching the theoretical Abbe diffraction limit of ˜200 nm, larger than DNA, RNA, proteins, and cytoskeletons (5-50 nm). Super-resolution microscopy, wherein the diffraction limit of light is overcome, has been the subject of several major developments during the past decade. STimulated Emission Depletion (STED) can be used as an approach to achieving super-resolution in fluorescence microscopy. In one example, STED uses an intense doughnut-shaped laser beam to trim the primary excitation focus by “switching off” the surrounding excited fluorophore(s) through a stimulated emission pathway (“de-excitation”). The spatial resolution achieved in STED microscopy is strongly dependent on the intensity of the depletion-laser beam: for standard biolabels (e.g. Alexa Fluor, and Atto dyes) lateral resolution of 62 nm has been reported for depletion-laser intensity of 400 MW/cm2, while resolution of 8 nm has been reported for depletion-laser intensity 3.7 GW/cm2. However, such large laser intensities commonly cause photobleaching of the biolabels and photo-thermal damage to the fragile sub-cellular structures of biological samples. Other associated issues, such as the laser complexity, stability and cost, are also becoming major impediments to advanced applications of STED in cell biology. Thus, a critical advance needed to extend the capabilities of STED microscopy in biomedical research is a new way to achieve high stimulated emission depletion factors (switch-off) at low laser pump intensities.
The fundamental problem of very high depletion pump intensities arises from the short (nanosecond) lifetimes of the biolabels used in STED. The depletion intensity is inversely proportional to the fluorescence lifetime of the target fluorophore, thus intensities of 108˜109 W/cm2 are needed in the depletion pump beam. This requires precisely synchronizing a pulsed laser within a very short time window or a CW synchronization-free laser of hundreds of milliwatts; both approaches are challenging, and in the case of “soft materials” impractical. Consistent with theory, it has previously been suggested that a solution to this problem is to employ target fluorophores with much longer lifetimes to reduce the depletion-intensity requirements commensurately. However, implementation of this simple idea has been precluded by the lack of practical fluorescent or luminescent materials or particles which have the requisite long lifetimes, are sufficiently bright and have sufficient depletion cross-section.
This offers another example application for the previously discussed upconversion particles/materials. The inventors use a lanthanide-based luminescent nanomaterial, being bright with both long excited-state lifetime and large depletion cross-section, suitable for low power stimulated emission depletion. The inventors found that the critical factors of both brightness and large depletion cross-section are only accessible by significantly increasing the doping concentration of activators in the upconversion nanocrystals. This condition has only become accessible after the inventors surprisingly realised the optimum concentration was power-dependent, as previously discussed. Sufficient excitation power (i.e. irradiance) under a laser scanning confocal microscope has been used to overcome the fundamental barrier of so-called concentration quenching (e.g. 0.5 mol % Tm3+), allowing tens of thousands of photostable emission centres (e.g. up to 8 mol % Tm3+) to be densely packed into a single dot.
Moreover, the ladder-like arranged energy levels in these crystals provide multiple intermediate excited states for the step-wise upconversion process, so that by. indirectly depleting the lower intermediate states it is possible to effectively switch “off” the higher level emissions. In comparison to current STED techniques, which use fluorescence biolabels, the advantages of this technique include high contrast in on-to-off ratio and high depletion efficiency.
An upconversion approach enables separation of the depletion wavelength from excitation wavelength. Clear separation of the de-excitation wavelength from the absorption wavelength is important, otherwise the depleted molecule may be re-excited by the strong depletion beam when the excitation spectra and emission spectra overlap. This overlap occurs for most fluorochromes used in STED, so that re-excitation caused by the depletion beam has been one of the major limitations for most dyes (including quantum dots), where depletion was chosen at the red-shifted tail of the emission band in STED.
To test the depletion efficiency, a single-mode 976 nm laser was employed as the primary excitation source in a confocal microscopy setup (x-y-z stage scan), and an 808 nm single-mode laser was coupled to the primary beam. Precision nanophotonics engineering was applied to ensure the two confocal beams precisely overlap through a high-performance objective. This setup allowed testing of the depletion efficiency of Tm3+-doped upconversion nanocrystals. While an upconversion nanocrystal with a conventional doping concentration of 0.5 mol % was difficult to switch off (they are even less efficient than the best-performing dye, Dylight 650, depleted at 783 nm in our previous CW STED system), a high doping concentration of 6 mol % Tm3+ was surprisingly easy to deplete. Indeed the upconversion nanocrystals were fully depleted at sub-milliwatt levels, three orders of magnitude lower power than the 0.5% crystals (see
To evaluate the optical resolution of upconversion particle based powered high-contrast STED nanoscopy, a phase plate was employed to generate an 808 nm “doughnut” PSF surrounding the excitation PSF to form the STED nanoscopy architecture. The efficacy of the new generation of luminescent upconversion particles and intermediate optical pumping scheme was evaluated for single nanocrystal STED imaging (refer to
Application of the upconversion particles in this manner provides luminescent biolabels that feature multiple, long-lived intermediate excited states, and produce bright and sharp luminescence emissions. Thus, this example application solves the main limitation of current STED-based super-resolution microscopy, namely that the high laser powers required to deplete the fluorescent dyes, and so achieve sub-100 nm resolution, also cause photobleaching and sample damage, thereby limiting the utility of the technique. Use of upconversion particles can provide important opportunities for practical improvements in super-resolution microscopy.
Excitation-dependent upconversion particles also enable a new approach to security inks, because highly doped (typically>4 mol %) Tm3+ nanocrystals remain dark unless high infrared excitation irradiance is used, in contrast to low level doped Tm3+ nanocrystals. Additionally, nanocrystal suspensions can be dispersed in traditional inkjet printer inks to print highly secure images, such as trademarks or logos, on papers and plastics.
This demonstration shows an application for security inks using power dependent Tm3+ concentration. In another example, low concentration (for example, 0.2 mol % Tm3+) nanocrystals can be used to stain a masking pattern which is visible under both low power illumination (about 104 W/cm2) and high power illumination (about 106 W/cm2 or greater). High concentration (for example, 4 mol % Tm3+) nanocrystals can be used to stain a hidden pattern (e.g. the Macquarie University logo or the fireworks in
Efficient upconversion emission can be realized at a high activator doping, but only when sufficient irradiance is provided. Sufficient excitation irradiance can unlock otherwise dark activators, thereby enhancing the upconversion brightness. This effect is independent of particle or crystal size (for example from tens to several hundreds of nanometres, to ‘bulk material’), surface conditions and synthesis conditions.
This effect in bulk crystals is demonstrated in
Various other applications using the upconversion particles are possible. For example, in detection, sensing, imaging, such as of biological material, flow cytometry, solar cell or display applications. A sensing application may be, for example, a fibre sensing method, such as a fibre dip sensing method. Display applications can include televisions and monitors.
The exceptional nanocrystal brightness provides compelling advantages to a wide range of fields including immunofluorescence imaging, rare event cell detection and quantification, document security and security printing. The ultrabright upconversion nanocrystals can be used to provide high-contrast biolabels. As a further illustrative example, Giardia lamblia cells can be labelled by nanocrystals conjugated to suitable monoclonal antibodies (G203). The labelled Giardia cells can be imaged by a scanning system at only about 0.1 s exposure time by a standard charge-coupled device (CCD) camera. The absence of autofluorescence background at 980 nm excitation enables the quantification of the absolute signal intensities of each single microorganism, as well as quantification of the level of surface antigens. Single labelled cells on a glass slide have been detected within 3 min without background interference. This shows that these bioprobes are capable of rare event detection.
Those skilled in the art will appreciate that the invention described herein is susceptible to variations and modifications other than those specifically described. It is to be understood that the invention includes all such variations and modifications.
The reference in this specification to any prior publication (or information derived from the prior publication), or to any matter which is known, is not, and should not be taken as an acknowledgment or admission or any form of suggestion that the prior publication (or information derived from the prior publication) or known matter forms part of the common general knowledge in the field of endeavour to which this specification relates.
| Number | Date | Country | Kind |
|---|---|---|---|
| 2012904043 | Sep 2012 | AU | national |
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/AU2013/001055 | 9/17/2013 | WO | 00 |
