WO2022269187A1 - Mid-ir laser additive printing equipment - Google Patents

Mid-ir laser additive printing equipment Download PDF

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Publication number
WO2022269187A1
WO2022269187A1 PCT/FR2022/051206 FR2022051206W WO2022269187A1 WO 2022269187 A1 WO2022269187 A1 WO 2022269187A1 FR 2022051206 W FR2022051206 W FR 2022051206W WO 2022269187 A1 WO2022269187 A1 WO 2022269187A1
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Prior art keywords
laser
equipment according
printed
donor
liquid
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PCT/FR2022/051206
Other languages
French (fr)
Inventor
Bertrand Viellerobe
Antonio IAZZOLINO
Fabien Guillemot
Dan Soto
Original Assignee
Poietis
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Priority to EP22744258.9A priority Critical patent/EP4359194A1/en
Publication of WO2022269187A1 publication Critical patent/WO2022269187A1/en

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Classifications

    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B29—WORKING OF PLASTICS; WORKING OF SUBSTANCES IN A PLASTIC STATE IN GENERAL
    • B29C—SHAPING OR JOINING OF PLASTICS; SHAPING OF MATERIAL IN A PLASTIC STATE, NOT OTHERWISE PROVIDED FOR; AFTER-TREATMENT OF THE SHAPED PRODUCTS, e.g. REPAIRING
    • B29C64/00—Additive manufacturing, i.e. manufacturing of three-dimensional [3D] objects by additive deposition, additive agglomeration or additive layering, e.g. by 3D printing, stereolithography or selective laser sintering
    • B29C64/10—Processes of additive manufacturing
    • B29C64/106—Processes of additive manufacturing using only liquids or viscous materials, e.g. depositing a continuous bead of viscous material
    • B29C64/112—Processes of additive manufacturing using only liquids or viscous materials, e.g. depositing a continuous bead of viscous material using individual droplets, e.g. from jetting heads
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B29—WORKING OF PLASTICS; WORKING OF SUBSTANCES IN A PLASTIC STATE IN GENERAL
    • B29C—SHAPING OR JOINING OF PLASTICS; SHAPING OF MATERIAL IN A PLASTIC STATE, NOT OTHERWISE PROVIDED FOR; AFTER-TREATMENT OF THE SHAPED PRODUCTS, e.g. REPAIRING
    • B29C64/00—Additive manufacturing, i.e. manufacturing of three-dimensional [3D] objects by additive deposition, additive agglomeration or additive layering, e.g. by 3D printing, stereolithography or selective laser sintering
    • B29C64/20—Apparatus for additive manufacturing; Details thereof or accessories therefor
    • B29C64/264—Arrangements for irradiation
    • B29C64/268—Arrangements for irradiation using laser beams; using electron beams [EB]
    • B29C64/273—Arrangements for irradiation using laser beams; using electron beams [EB] pulsed; frequency modulated
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B33—ADDITIVE MANUFACTURING TECHNOLOGY
    • B33Y—ADDITIVE MANUFACTURING, i.e. MANUFACTURING OF THREE-DIMENSIONAL [3-D] OBJECTS BY ADDITIVE DEPOSITION, ADDITIVE AGGLOMERATION OR ADDITIVE LAYERING, e.g. BY 3-D PRINTING, STEREOLITHOGRAPHY OR SELECTIVE LASER SINTERING
    • B33Y10/00—Processes of additive manufacturing
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B33—ADDITIVE MANUFACTURING TECHNOLOGY
    • B33Y—ADDITIVE MANUFACTURING, i.e. MANUFACTURING OF THREE-DIMENSIONAL [3-D] OBJECTS BY ADDITIVE DEPOSITION, ADDITIVE AGGLOMERATION OR ADDITIVE LAYERING, e.g. BY 3-D PRINTING, STEREOLITHOGRAPHY OR SELECTIVE LASER SINTERING
    • B33Y30/00—Apparatus for additive manufacturing; Details thereof or accessories therefor
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B33—ADDITIVE MANUFACTURING TECHNOLOGY
    • B33Y—ADDITIVE MANUFACTURING, i.e. MANUFACTURING OF THREE-DIMENSIONAL [3-D] OBJECTS BY ADDITIVE DEPOSITION, ADDITIVE AGGLOMERATION OR ADDITIVE LAYERING, e.g. BY 3-D PRINTING, STEREOLITHOGRAPHY OR SELECTIVE LASER SINTERING
    • B33Y70/00—Materials specially adapted for additive manufacturing
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B33—ADDITIVE MANUFACTURING TECHNOLOGY
    • B33Y—ADDITIVE MANUFACTURING, i.e. MANUFACTURING OF THREE-DIMENSIONAL [3-D] OBJECTS BY ADDITIVE DEPOSITION, ADDITIVE AGGLOMERATION OR ADDITIVE LAYERING, e.g. BY 3-D PRINTING, STEREOLITHOGRAPHY OR SELECTIVE LASER SINTERING
    • B33Y80/00—Products made by additive manufacturing

Definitions

  • the present invention relates to the field of bio-printing, based on a brick-by-brick reconstruction of a fabric. These methods offer the possibility of arranging each component according to a predefined pattern which guides the subsequent maturation of the tissue construct towards its final functional architecture. Cell distribution can be defined at the micrometric scale. Layer-by-layer control of the distribution of cells and extracellular matrix components within the matrix promotes tissue maturation. Bio-printing benefits from the capabilities of rapid prototyping, facilitated by computer-aided design and/or manufacturing procedures (CAD and CAM respectively), which control the geometry of the internal structure and the external shape of the cellular scaffolds during their impression.
  • CAD and CAM computer-aided design and/or manufacturing procedures
  • Bioprinters based on LIFT (laser-induced forward transfer) technology are composed of three elements: a laser source, generally pulsed, focused on the material to be printed, a donor on which the biological material to be printed is based, and a receiver/target receiver substrate which collects the printed material.
  • LIFT laser-induced forward transfer
  • the donor is composed of a support transparent to the laser (for example glass or quartz) coated with a thin "sacrificial" absorbent metallic layer generally composed of gold or titanium.
  • the organic component molecules or cells
  • a liquid phase for example a culture medium, serum or even a solution of polymer-type biomaterial
  • the laser pulse induces an energy concentration at the metal-liquid interface to be printed, leading to the creation of a cavitation bubble which causes a deformation of the interface of the liquid film located above until the production of a jet. This allows the deposition of droplets on the receiving substrate located opposite the support on which the ink is located.
  • This method makes it possible to deposit cells or biomaterials over a very wide range of sizes and at speeds that can reach several thousand drops/s. Thanks to a resolution that can go down to a few tens of picoliters, laser-assisted bio-printing makes it possible to finely control the cell density and the spatial organization of the 3D printing with great precision. In principle, it makes it possible to individually control the printing of each cell and therefore to exert control of an unprecedented level of the distribution of these in the printed fabric.
  • this emerging technology is suitable for the manufacture of structures which not only mimic the structural organization of native tissues, but are also capable of developing a physiological functionality close to that of their native counterparts.
  • This method also has advantages such as the ability to be automated, the reproducibility and the high throughput of the deposition of the constituent elements of the tissue, which make it possible to envisage the manufacture of 3D constructions of relevant sizes on the physiological and therefore clinical level.
  • the use of a sacrificial layer has several drawbacks, including that of the permanent local ablation of the metal following the laser firing, requiring a frequent change of the substrate supporting the ink and therefore repeated interruptions in the printing sequence of a fabric to renew said substrate.
  • Patent US10940687B2 illustrates an embodiment of the prior art.
  • a laser emitting a beam with a wavelength of 700 nanometers uses a laser emitting a beam with a wavelength of 700 nanometers.
  • a standard LIFT process is described in French patents FR3030360B1 or FR3087702B1 or FR3063930B1.
  • the laser is directed upwards, and the energy provided locally by the laser pulse is intended to create a cavitation bubble from the plasma generated on the sacrificial layer propelling a biological element towards the target, with enough kinetic energy to overcome gravity and surface tension.
  • the substrates must be renewed regularly, which represents both a cost in time and money.
  • particles of the metallic sacrificial layer can be brought to the target by means of the jets and drops generated.
  • the invention here describes a LIFT printing method without recourse to a sacrificial layer ensuring good reproducibility of the jets generated and compatibility with the printing of living cells for the production of 3D tissues.
  • the invention is based on the use of a laser in the field of the IR medium around 2.9 ⁇ m allowing direct linear absorption of the ink liquid for carrying out an efficient and reproducible LIFT process. State of the art
  • the laser-matter interaction is very sensitive to the presence of objects such as cells for example in the absorption zone with the consequence of significant variations in the sizes and height of the jets generated, and hence a quality and random reproducibility of printed patterns.
  • the nonlinear absorption domain is therefore limited in its ability to imprint colloidal media.
  • the focusing of the energy must be perfectly controlled and correctly positioned either in very low ink thicknesses (of the order of a few tens of microns) or in very precise positions under the surface of the liquid. This results in extremely complex systems that are difficult to industrialize or transfer for large-scale applications. [0007] This patent also mentions the fact of working in linear absorption to circumvent the problems of reproducibility mentioned above.
  • the transfer carrier has no absorber layer.
  • the transfer material layer preferably serves as an absorber layer.
  • IR laser beam sources with > 3 pm wavelength are preferably used. ". It therefore describes the use of a wavelength greater than 3 ⁇ m to achieve direct absorption of the laser used by the medium. No details about the process of creating the bubble and the jet are described there. It therefore does not constitute a sufficient description for carrying out the present invention, knowing that the wavelength range proposed is different from that defended by the present invention.
  • shock waves and cavitation bubbles are studied using the technique of ombroscopy and acoustic measurements.
  • the conversion into shock wave energy reaches 67%.
  • Most of the shock wave energy (92%) is dissipated as the shock front travels the first 250 pm, and the remaining 8% is transferred to the acoustic far field.
  • the results of the article can be used for the optimization of the laser parameters to carry out micro-machining or the ablation of biological tissues.
  • the article does not describe a LIFT type process for material transfer but a linear absorption process in a liquid to generate cavitation bubbles.
  • Bio-printing solutions by LIFT can also use a specific interaction layer for the laser but which is not metallic.
  • BA-LIFT blister assisted LIFT
  • LIFT based on the use of a gelatin layer for example. In all cases, this layer plays the role of an actuator which deforms to allow the creation of the jet.
  • the sacrificial nature of the metal layer represents the main drawback of standard laser bio-printing because the donor has a limited use time related to the definitive ablation of the metal following the laser shot.
  • the change of donor is therefore necessarily recurrent and time-consuming, which is potentially prohibitive for use in bio-printing where the manufacturing time of the tissues is critical for their viability.
  • it requires significant user stress depending on the number of cell impressions to be made and therefore the number of donor changes to manufacture a tissue.
  • Another limitation of the sacrificial layers relates to the quality of the metal layer deposited on the glass. Indeed, the latter may suffer from weak adhesion of the gold to the glass, from a variation in the thickness of the metal on the glass surface or even from variation in the method of depositing the metallic film.
  • the direct consequence is a disparity of the jets generated since the initial conditions are not identical at any point of the donor. It may also result in delamination of the metal layer during use. The consequence is a loss of print quality or even the obligation to start a new print.
  • Another known limit lies in the period of validity of the metallization state sacrificial layers, a state which must not change over time in order to ensure high reproducibility of the printing process.
  • the present invention relates, in its most general sense, to additive printing equipment comprising: a pulsed laser source producing a beam and focused on the material to be printed, a donor to from which a biological material is imprinted, and a target recipient substrate which collects the imprinted material.
  • Said donor being constituted by a coated plate, in the interaction zone of the beam laser, by a liquid film intended to contain transferable inhomogeneities, characterized in that said laser emits a beam whose wavelength is between 2 pm and 3.2 pm, said plate being transparent or weakly absorbing at the wavelength of said laser beam.
  • the absorption of the laser in the wavelength range between 2 and 3.2 ⁇ m is proportional to the quantity of liquid present at the level of the illumination plane, one then speaks of linear absorption.
  • the preferred wavelength is around 2.9pm because it corresponds to the highest liquid absorption peak as can be seen in Figure 2.
  • the advantage of linear direct absorption is to open up a new field absorption corresponding to small laser energies making it possible to create small bubbles and therefore small jets, in a regime where the bubble and the jet generated are sufficient to make an impression.
  • printing at 2.9pm opens the way to a wide range of printed drop sizes from picolitre to nanolitre.
  • Direct absorption means that there is no intermediate element or process in the interaction between the laser and the liquid medium. In fact, there are no absorbent elements added to the solution, nor a metallic sacrificial layer which makes it possible to initiate absorption, nor complex optical effects, nor physico-chemical or electromagnetic effects to initiate absorption. 'absorption. This is carried out directly by absorption of the liquid whether it is aqueous (water molecules) or non-aqueous (polymers in solution, etc.).
  • Total absorption means that there is no residual photon of the laser after a certain thickness through which said laser passes in the fluid to be printed. This thickness has been estimated experimentally and is less than 5 ⁇ m.
  • the fluid present in this small thickness plays a role equivalent to that of a sacrificial layer without really being so. It makes it possible in particular to confine the absorption of the laser to the liquid-substrate interface, over an area of a few microns, as is usually done with a metallic sacrificial layer which is generally a few tens of nanometers. This makes it possible to obtain a positioning of the cavitation bubble similar to that obtained conventionally by creating a plasma within a sacrificial layer.
  • the dynamics of the bubble and the jet are very similar to those of a system based on the use of a sacrificial layer with the advantage of being able to reuse the donor as many times as necessary since the quantity of liquid present in this one is important and renewable.
  • the total absorption of the laser on the first microns of the liquid ensures protection of the rest of the liquid with respect to the photons used and therefore the cells which are there. It is a very interesting alternative for cell viability compared to other solutions where UV (ionizing) lasers are used to make laser bio-printing for example.
  • Another advantage is that absorption can be viewed as a 2D process that requires less precision for positioning the laser focal plane.
  • Printing at 2.9 ⁇ m is therefore based on the interaction of the laser with a few ⁇ m of thickness of a homogeneous fluid which guarantees high reproducibility of the laser-material interaction, resulting in high stability of the bubbles and jets generated. Such reproducibility is very important in the field of bio-printing, in particular for industrial and clinical purposes.
  • FR3063931A1 proposes an implementation where the ink to be printed enters and leaves the open printing zone of the donor by fluidic tubes.
  • the laser is fired into said open area which may be circular in shape (similar to what is practiced with conventional laser printing systems) or more special in shape such as a single or multiple fluidic channel.
  • the laser shots can be fired on the same point or on lines of points as many times as necessary without having to change the cartridge.
  • the present invention relates, in its most general sense, to additive printing equipment comprising: a pulsed laser source producing a beam and focused on the material to be printed, a donor without a sacrificial layer, from which a biological material is printed, and a target receiver substrate which collects the printed material.
  • Said donor being constituted by a covered plate, in the interaction zone of the beam laser, by a liquid film intended to contain transferable inhomogeneities, characterized in that the said laser emits a beam whose wavelength is between 2 ⁇ m and 3.2 ⁇ m, the said plate being transparent or weakly absorbing at the length of wave of said laser beam.
  • the emission wavelength of the laser has a peak at 2.9 pm ⁇ 0.3 pm
  • the thickness of the liquid in which the total absorption of the laser energy is achieved is order of a few microns and less than 5 ⁇ m
  • the focusing of the laser at the interface of the donor and the liquid can be modified so as to allow variation of the transverse size of the absorption zone with the effect of modulating the size of the bubbles and the jets generated and therefore drops deposited on the target laser printing at 2.9pm is perfectly suited to the use of a cartridge with continuous fluid refill a cartridge with continuous refill using a pre-wetting film could use technical ink pre-wetting as an absorbent medium for the wavelength at 2.9 pm the fluid is placed in a tank the impression is made from bottom to top in the opposite direction to the force of gravity
  • the performances associated with this equipment make it possible to ensure the following characteristics: the size of the cavitation bubble and the size of the associated jet are linearly dependent on the one hand on the laser energy used which is typically in the range from pJ to a few
  • the invention also relates to a method of additive printing by a pulsed laser source producing a beam in the direction of a donor without a sacrificial layer from which a biological material is printed, towards a receiving substrate which collects the printed material.
  • said donor being constituted by a plate covered, in the zone of interaction of the laser beam, by a film of liquid intended to contain transferable inhomogeneities, characterized in that said laser emits a beam whose wavelength is between 2 ⁇ m and 3.2 ⁇ m and in that said plate is transparent or weakly absorbent at the wavelength of said laser beam.
  • the fluid to be printed comes from a cartridge with continuous reloading.
  • the routing of the ink to the cartridge is automated to avoid any human intervention during the printing phase.
  • FIG. 1 shows a schematic view of a LIFT system according to the invention working at 2.9pm
  • FIG. 2 Figure 2 shows the absorption spectrum of water
  • FIG. 3a represents a curve of the stability of the jets obtained by ombroscopy
  • FIG. 3b represents a curve of the jet height measured on 50 consecutive jets
  • FIG. 4 Figure 4 represents a curve of the dependence of jet heights as a function of time and laser energy
  • Figure 5 represents a graph of the characteristics of the drops printed on the receiver substrate as a function of the distance to the receiver
  • Figure 6 represents the geometry of the drops printed by the direct LIFT technique at 2.9 ⁇ m on a substrate without sacrificial layer. Description of a first example of equipment
  • FIG. 1 shows a schematic view of a first example of equipment.
  • the laser (1) is a pulsed source in the nanosecond regime, the pulse duration of which is 10 ns, emitting between 2.6 ⁇ m and 3.2 ⁇ m in wavelength, in an energy regime capable of reaching 100pJ, with an adjustable rate of fire between 1Hz and 1OKHz.
  • the laser beam is shaped through an optical system (2) composed of two lenses in order to collimate it and give it the desired diameter.
  • the optical components must be made of ZnSe, CaF2, Infrasil or any other material compatible with use in the wavelength range of the IR medium (from 2 to 3.2 pm).
  • the scanner (3) ensures an angular orientation of the beam along two perpendicular axes. It is for example constituted by two mirrors actuated by an electromagnetic actuator, for example a scanner marketed by the company SCANLAB (trade name) under the reference “SCANcube 14”.
  • the mirrors of this scanner must be covered with a layer of gold to ensure total reflection of the laser in the wavelength range of the IR medium (from 2 to 3.2 ⁇ m).
  • the laser (1) is then focused by a dedicated focusing optical system (4) on the donor (5) on which the ink to be printed (6) is deposited.
  • a dedicated focusing optical system (4) on the donor (5) on which the ink to be printed (6) is deposited.
  • the focusing system in standard LIFT systems is based on complex optical systems called F-Theta with a large number of lenses.
  • F-Theta complex optical systems
  • Telecentric lens consisting of 2 custom-made lenses compatible with the 2.9pm wavelength (simpler and less expensive solution)
  • a 3D scanner type solution would be used where the focusing would be done before the scanner and would require the use of an adaptation along the Z direction of the focal length in the field targeted.
  • the film (6) is arranged opposite a target receiver substrate (7) sufficiently transparent to the wavelength of the laser to which the cells or particles are transferred, when a laser beam (1) is triggered. impulsive.
  • Figure 2 illustrates the absorption capacity of water as a function of the illumination wavelength and one can clearly see there the absorption peak at 2.9 pm targeted by the present invention in its implementation. particular work.
  • FIG. 3a represents the stability curve of the jets measured by a ombroscopy technique implemented in the example embodiment described in FIG. 1. It can easily be concluded that the height of the jets remains the same over a large number consecutive throws.
  • FIG. 3b represents the curve of the jet height measured over 50 consecutive jets from shadow data. It can be seen that the repeatability of the jet heights is greater than 90%. This stability is a guarantee of print reproducibility, an absolutely necessary characteristic for using bio-printing in the industrial or clinical field. It was obtained on the system described in figure 1.
  • FIG. 4 represents a curve of the dependence of the jet heights as a function of time and of the laser energy obtained with the system described in FIG. 1.
  • the direct dependence between energy and jet height is clearly observable and provides the proof of the linear character of the absorption at 2.9 pm.
  • the jets have a "lifetime" of a few hundred ps, which is standard in the field of LIFT. We are therefore on short printing times compatible with the printing of biological tissues which must remain viable during this phase.
  • FIG. 5 represents a graph of the characteristics of the drops printed on the receiver substrate as a function of the distance from said receiver in the context of the embodiment described in FIG. 1.
  • the size of the drops follows the same logic as that described in the jets, namely a linear dependence on the laser energy used and an ability to obtain reproducible sizes on a large scale.
  • Figure 6 is the final proof of the ability of the direct LIFT at 2.9 ⁇ m to print drops on a substrate without sacrificial layer with results obtained on the assembly described in Figure 1.
  • the printing relates to a homogeneous medium in this drawing. Similar results were obtained on colloidal media with microbeads or human cells. It illustrates the topography of the target after printing with four different energy levels. Description of a second example of equipment
  • the solution can be achieved without using a 2D or 3D scanner.
  • the optical line is simplified because it only integrates a beam shaping system and focusing optics.
  • the ink deposited at the level of the donor will be sufficiently disturbed by each laser shot resulting in a bubble then a jet to allow a form of renewal of the ink at the level of the zone of the laser shot.
  • This configuration can be imagined both in the case of a printing architecture from bottom to top in the direction opposite to the force of gravity as well as from top to bottom in the direction of gravity.
  • the donor may consist of a fluid film such as a reservoir.
  • the 2.9 ⁇ m wavelength of the laser beam does not allow the use of standard optics.
  • the optical components must therefore be based on other materials which are not absorbent at these wavelengths and which are more expensive. It is therefore preferable to use transparent materials such as CaF2 , ZnSe or Infrasil (trade name) even if it is also possible to use as donor substrate thin layers of silica at the cost of a loss of energy.
  • the lenses used are CaF2 lenses and the mirrors must be in protected gold.
  • the emission at 2.9 ⁇ m is characterized by strong absorption of this radiation by the water. This characteristic could then bring this type of radiation into the category of so-called eye safety lasers such as erbium lasers emitting at 1500 nm. It is therefore the cornea that absorbs this radiation.
  • the water absorption coefficient at 2.9 pm (10000 cm 4 ) is much greater than at 1.5 pm (30 cm -1 ), absorption therefore takes place over distances less than 2 to 3 pm from wavelength versus 700pm to 1.5pm wavelength. The risks of damage to the cornea are therefore greater at 3 pm than at 1.5 pm, but much less concerning the retina.
  • the optical system allowing the propagation of the laser beam to the donor comprises a 2D or 3D scanner making it possible to orient said laser beam in order to create a complete pattern of drops printed by scanning said laser beam on said donor whose surface is at least 0.5 cm2.
  • the optical system does not include a scanner and shoots directly on the donor, each shot making it possible to renew the interaction zone within said donor by the movements induced in the film of ink by the cavitation bubble and the jet.
  • the optical system is integrated into a module allowing humidity regulation.
  • the optical system is integrated in a closed enclosure in which a neutral gas has been introduced in order to avoid any absorption by water vapor
  • the size of the drops deposited is linearly dependent on the laser energy and the thickness of the fluid to be printed

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Abstract

The present invention relates to the field of bioprinting, based on a brick-by-brick reconstruction of a tissue. These methods offer the possibility of arranging each component according to a predefined pattern that guides the subsequent maturation of the tissue construct until its final functional architecture. The distribution of the cells can be defined on a micrometre scale. Layer-by-layer control of the distribution of the cells and the extracellular matrix components in the matrix promotes tissue maturation. Bioprinting benefits from the capabilities of rapid prototyping, facilitated by computer-aided design and/or manufacturing procedures (CAD and CAM, respectively), which control the geometry of the internal structure and the external shape of the cell scaffolds as they are printed.

Description

DESCRIPTION DESCRIPTION
Titre : EQUIPEMENT D'IMPRESSION ADDITIVE PAR LASER MID IR Title: ADDITIVE PRINTING EQUIPMENT BY LASER MID IR
Domaine de l'invention Field of the invention
[0001] La présente invention concerne le domaine de la bio-impression, fondée sur une reconstruction brique par brique d'un tissu. Ces méthodes offrent la possibilité de disposer chaque composant selon un patron prédéfini qui guide la maturation subséquente de la construction tissulaire vers son architecture fonctionnelle finale. La distribution des cellules peut être définie à l'échelle micrométrique. Le contrôle couche par couche de la distribution des cellules et des composants de la matrice extracellulaire dans la matrice favorise la maturation tissulaire. La bio-impression bénéficie des capacités du prototypage rapide, facilitée par des procédures de conception et / ou de fabrication assistée par ordinateur (respectivement CAO et FAO), qui contrôlent la géométrie de la structure interne et la forme externe des échafaudages cellulaires lors de leur impression. The present invention relates to the field of bio-printing, based on a brick-by-brick reconstruction of a fabric. These methods offer the possibility of arranging each component according to a predefined pattern which guides the subsequent maturation of the tissue construct towards its final functional architecture. Cell distribution can be defined at the micrometric scale. Layer-by-layer control of the distribution of cells and extracellular matrix components within the matrix promotes tissue maturation. Bio-printing benefits from the capabilities of rapid prototyping, facilitated by computer-aided design and/or manufacturing procedures (CAD and CAM respectively), which control the geometry of the internal structure and the external shape of the cellular scaffolds during their impression.
[0002] Les bio-imprimantes basées sur la technologie LIFT (laser-induced forward transfer) sont composées de trois éléments : une source laser, généralement pulsée, focalisée sur le matériel à imprimer un donneur sur lequel repose le matériel biologique à imprimer, et un substrat récepteur/ receveur cible qui recueille le matériel imprimé. [0002] Bioprinters based on LIFT (laser-induced forward transfer) technology are composed of three elements: a laser source, generally pulsed, focused on the material to be printed, a donor on which the biological material to be printed is based, and a receiver/target receiver substrate which collects the printed material.
[0003] Dans les premières générations de bio-imprimantes, le donneur est composé d'un support transparent au laser (par exemple du verre ou du quartz) revêtu d'une mince couche métallique absorbante « sacrificielle » composée généralement d'or ou de titane. Le composant organique (molécules ou cellules) est préparé dans une phase liquide (par exemple un milieu de culture, du sérum ou encore une solution de biomatériau de type polymère) puis déposé à la surface dudit film métallique. L'impulsion laser induit une concentration énergétique à l'interface métal - liquide à imprimer, entraînant la création d'une bulle de cavitation qui engendre une déformation de l'interface du film liquide se trouvant au-dessus jusqu'à la production d'un jet. Celui-ci permet le dépôt de gouttelettes sur le substrat receveur situé en regard du support sur lequel est située l'encre. Cette méthode permet de déposer des cellules ou des biomatériaux sur une très large gamme de tailles et selon des vitesses pouvant atteindre plusieurs milliers de gouttes/s. Grâce à une résolution pouvant descendre à quelques dizaines de picolitres, la bio-impression assistée par laser permet de maîtriser finement la densité cellulaire et l'organisation spatiale de l'impression en 3D avec une grande précision. En principe, elle permet de contrôler individuellement l'impression de chaque cellule et donc d'exercer un contrôle d'un niveau sans précédent de la répartition de celles-ci dans le tissu imprimé. [0003] In the first generations of bio-printers, the donor is composed of a support transparent to the laser (for example glass or quartz) coated with a thin "sacrificial" absorbent metallic layer generally composed of gold or titanium. The organic component (molecules or cells) is prepared in a liquid phase (for example a culture medium, serum or even a solution of polymer-type biomaterial) then deposited on the surface of said metal film. The laser pulse induces an energy concentration at the metal-liquid interface to be printed, leading to the creation of a cavitation bubble which causes a deformation of the interface of the liquid film located above until the production of a jet. This allows the deposition of droplets on the receiving substrate located opposite the support on which the ink is located. This method makes it possible to deposit cells or biomaterials over a very wide range of sizes and at speeds that can reach several thousand drops/s. Thanks to a resolution that can go down to a few tens of picoliters, laser-assisted bio-printing makes it possible to finely control the cell density and the spatial organization of the 3D printing with great precision. In principle, it makes it possible to individually control the printing of each cell and therefore to exert control of an unprecedented level of the distribution of these in the printed fabric.
[0004] Ainsi, cette technologie émergente est adaptée à la fabrication de structures qui non seulement miment l'organisation structurelle des tissus natifs, mais sont également susceptibles de développer une fonctionnalité physiologique proche de celle de leurs homologues natifs. Cette méthode présente aussi des avantages comme la capacité à être automatisée, la reproductibilité et le haut débit du dépôt des éléments constituants le tissu, qui permettent d'envisager la fabrication de constructions 3D de tailles pertinentes sur le plan physiologique et donc clinique. L'utilisation d'une couche sacrificielle présente toutefois plusieurs inconvénients, dont celui de l'ablation locale définitive du métal suite au tir laser obligeant un changement fréquent du substrat supportant l'encre et donc des interruptions répétées dans la séquence d'impression d'un tissu pour renouveler ledit substrat. Le brevet US10940687B2 illustre un exemple de réalisation de l'art antérieur. Il utilise un laser émettant un faisceau d'une longueur d'onde de 700 nanomètres. Un autre exemple de procédé LIFT standard est décrit dans les brevets français FR3030360B1 ou FR3087702B1 ou FR3063930B1. Dans ces réalisations, le laser est dirigé vers le haut, et l'énergie apportée localement par l'impulsion laser a pour but de créer une bulle de cavitation à partir du plasma généré sur la couche sacrificielle propulsant un élément biologique vers la cible, avec une énergie cinétique suffisante pour vaincre la gravité et les tensions superficielles. Dans chacun de ces cas de mise en œuvre, les substrats doivent être renouvelés régulièrement ce qui représente à la fois un coût en temps et financier. De plus, des particules de la couche sacrificielle métallique peuvent être amenées jusqu'à la cible par le biais des jets et gouttes générés. L'invention décrit ici un procédé d'impression LIFT sans recours à une couche sacrificielle assurant une bonne reproductibilité des jets générés et une compatibilité avec l'impression de cellules vivantes pour la réalisation de tissus en 3D. L'invention se base sur l'utilisation d'un laser dans le domaine du milieu IR autour de 2.9pm permettant une absorption linéaire directe du liquide de l'encre pour la réalisation d'un procédé LIFT efficace et reproductible. Etat de la technique [0004] Thus, this emerging technology is suitable for the manufacture of structures which not only mimic the structural organization of native tissues, but are also capable of developing a physiological functionality close to that of their native counterparts. This method also has advantages such as the ability to be automated, the reproducibility and the high throughput of the deposition of the constituent elements of the tissue, which make it possible to envisage the manufacture of 3D constructions of relevant sizes on the physiological and therefore clinical level. However, the use of a sacrificial layer has several drawbacks, including that of the permanent local ablation of the metal following the laser firing, requiring a frequent change of the substrate supporting the ink and therefore repeated interruptions in the printing sequence of a fabric to renew said substrate. Patent US10940687B2 illustrates an embodiment of the prior art. It uses a laser emitting a beam with a wavelength of 700 nanometers. Another example of a standard LIFT process is described in French patents FR3030360B1 or FR3087702B1 or FR3063930B1. In these embodiments, the laser is directed upwards, and the energy provided locally by the laser pulse is intended to create a cavitation bubble from the plasma generated on the sacrificial layer propelling a biological element towards the target, with enough kinetic energy to overcome gravity and surface tension. In each of these cases of implementation, the substrates must be renewed regularly, which represents both a cost in time and money. In addition, particles of the metallic sacrificial layer can be brought to the target by means of the jets and drops generated. The invention here describes a LIFT printing method without recourse to a sacrificial layer ensuring good reproducibility of the jets generated and compatibility with the printing of living cells for the production of 3D tissues. The invention is based on the use of a laser in the field of the IR medium around 2.9 μm allowing direct linear absorption of the ink liquid for carrying out an efficient and reproducible LIFT process. State of the art
[0005] L'état de la technique concerne des solutions existantes proposant la réalisation d'un procédé LIFT ou plus généralement d'interaction laser - matière sans recours à une couche sacrificielle. Ces solutions suivent des stratégies assez différentes mais ont toutes des limites que la présente invention propose de résoudre. [0005] The state of the art relates to existing solutions proposing the realization of a LIFT process or more generally of laser-material interaction without recourse to a sacrificial layer. These solutions follow quite different strategies but all have limitations that the present invention proposes to resolve.
[0006] Dans le brevet FR3063931A1 « EQUIPEMENT ET PROCEDE D'IMPRESSION ADDITIVE », l'impression sans couche sacrificielle se base sur l'utilisation d'effets non linéaires d'absorption (multi-photoniques par exemple) permettant la création d'un plasma duquel découle tout le processus d'impression LIFT (bulle de cavitation, jet, goutte). Le principe décrit dans ce brevet est d'atteindre des seuils d'énergie qui dépassent le seuil d'ionisation de la matière afin d'enclencher le processus d'impression. Les lasers utilisés dans ce cadre sont onéreux car leur durée d'impulsion doit être très courte (typiquement en régime picoseconde / femtoseconde). De plus, l'interaction laser - matière est très sensible à la présence d'objets comme des cellules par exemple dans la zone d'absorption avec pour conséquence des variations importantes dans les tailles et la hauteur des jets générés, et de là une qualité et une reproductibilité aléatoires des motifs imprimés. Le domaine d'absorption non linéaire est donc limité dans sa capacité à imprimer des milieux colloïdaux. En sus, la focalisation de l'énergie doit être parfaitement maîtrisée et correctement positionnée soit dans des épaisseurs d'encre très faibles (de l'ordre de quelques dizaines de microns) soit dans des positions très précises sous la surface du liquide. Il en résulte ainsi des systèmes extrêmement complexes et difficiles à industrialiser ou transférer pour des applications à grande échelle. [0007] Ce brevet évoque également le fait de travailler en absorption linéaire pour contourner les problématiques de reproductibilité évoquées ci-avant. Le document ne décrit qu'un cas particulier de mise en oeuvre où des éléments absorbants ont été préalablement intégrés au liquide à imprimer. Ainsi, le milieu liquide joue le rôle d'absorbant et la reproductibilité des jets est bien meilleure. Malheureusement, les éléments absorbants ajoutés comme la mélanine dans ce cas sont généralement d'une nature chimique peu compatible avec l'utilisation d'encres cellulaires notamment pour des applications thérapeutiques qui impliquent de réduire l'introduction d'éléments potentiellement bio-actifs dans les procédés de fabrication. Cette mise en œuvre est donc elle aussi limitée dans ses capacités. [0008] Au final, les conditions décrites et protégées dans ce brevet sont éloignées de celles de la présente invention à la fois par leur nature et par leur performance. [0006] In the patent FR3063931A1 "EQUIPMENT AND METHOD FOR ADDITIVE PRINTING", printing without a sacrificial layer is based on the use of non-linear absorption effects (multi-photonic for example) allowing the creation of a plasma from which the entire LIFT printing process (cavitation bubble, jet, drop) derives. The principle described in this patent is to reach energy thresholds which exceed the ionization threshold of the material in order to initiate the printing process. The lasers used in this context are expensive because their pulse duration must be very short (typically in the picosecond/femtosecond regime). In addition, the laser-matter interaction is very sensitive to the presence of objects such as cells for example in the absorption zone with the consequence of significant variations in the sizes and height of the jets generated, and hence a quality and random reproducibility of printed patterns. The nonlinear absorption domain is therefore limited in its ability to imprint colloidal media. In addition, the focusing of the energy must be perfectly controlled and correctly positioned either in very low ink thicknesses (of the order of a few tens of microns) or in very precise positions under the surface of the liquid. This results in extremely complex systems that are difficult to industrialize or transfer for large-scale applications. [0007] This patent also mentions the fact of working in linear absorption to circumvent the problems of reproducibility mentioned above. The document only describes a particular case of implementation where absorbent elements have been previously integrated into the liquid to be printed. Thus, the liquid medium acts as an absorbent and the reproducibility of the jets is much better. Unfortunately, the absorbent elements added such as melanin in this case are generally of a chemical nature that is not very compatible with the use of cellular inks, in particular for therapeutic applications which involve reducing the introduction of potentially bio-active elements into the manufacturing processes. This implementation is therefore also limited in its capabilities. [0008] In the end, the conditions described and protected in this patent are far removed from those of the present invention both by their nature and by their performance.
[0009] Dans le brevet DE102017207262B4, il est fait mention d'une impression LIFT sans couche sacrificielle au niveau de la description du brevet : « In a preferred embodiment of the présent invention, the transfer carrier has no absorber layer. The transfer material layer preferably serves as an absorber layer. When using the transfer material layer as the absorber layer, IR laser beam sources with > 3 pm wavelength are preferably used. ». Il décrit donc l'utilisation d'une longueur d'onde supérieure à 3pm pour réaliser une absorption directe du laser utilisé par le milieu. Aucun détail sur le processus de création de la bulle et du jet n'y est décrit. Il ne constitue donc pas une description suffisante pour la réalisation de la présente invention en sachant que le domaine de longueur d'onde proposé est différent de celui défendu par la présente invention. [0009] In patent DE102017207262B4, mention is made of a LIFT printing without sacrificial layer in the description of the patent: “In a preferred embodiment of the present invention, the transfer carrier has no absorber layer. The transfer material layer preferably serves as an absorber layer. When using the transfer material layer as the absorber layer, IR laser beam sources with > 3 pm wavelength are preferably used. ". It therefore describes the use of a wavelength greater than 3 μm to achieve direct absorption of the laser used by the medium. No details about the process of creating the bubble and the jet are described there. It therefore does not constitute a sufficient description for carrying out the present invention, knowing that the wavelength range proposed is different from that defended by the present invention.
[0010] Dans l'article, « Cavitation and shock waves émission on the rigid boundary of water under mid-IR nanosecond laser puise excitation » A.V. Pushkin, A. S. Bychkov, A. A. Karabutov and F.V. Potemkin, Laser Phys. Lett. 15 (2018) 065401, les processus de conversion de l'énergie lumineuse en énergie mécanique sous l'excitation d'un laser nanoseconde dans l'IR moyen sur une frontière rigide d'eau sont décrits. La forte absorption par l'eau d'un laser Q.-switched Cr:Yb:Ho:YSGG (2.85 pm, 6 mJ, 45 ns) permet un dépôt rapide d'énergie de ~8 kJ/ cm3 accompagné de fortes transitions mécaniques. L'évolution des ondes de choc et des bulles de cavitation est étudiée à l'aide de la technique d'ombroscopie et de mesures acoustiques. Pour une impulsion laser de 6 mJ avec une fluence de 2.0 J. cm 2, la conversion en énergie d'onde de choc atteint 67%. La majeure partie de l'énergie de l'onde de choc (92 %) est dissipée lorsque le front de choc parcourt les premiers 250 pm, et les 8 % restants sont transférés au champ lointain acoustique. Les résultats de l'article peuvent être utilisés pour l'optimisation des paramètres du laser pour réaliser du micro-usinage ou de l'ablation de tissus biologiques. L'article ne décrit pas un procédé de type LIFT pour le transfert de matière mais un processus d'absorption linéaire dans un liquide pour générer des bulles de cavitation. Les énergies étudiées dans cet article sont de 2 ordres de grandeur supérieures à celles décrites dans la présente invention, de même que la taille des bulles qui est de plusieurs mm dans l'article quand la présente invention décrit des objets de quelques dizaines à centaines de microns. Les conditions illustrées dans cet article sont donc très éloignées de celles recherchées pour la bio impression précise de cellules par transfert LIFT. [0011] Les solutions de bio-impression par LIFT peuvent également utiliser une couche d'interaction spécifique pour le laser mais non métallique. On parle de BA-LIFT (blister assisted LIFT) ou de LIFT basé sur l'utilisation d'une couche de gélatine par exemple. Dans tous les cas, cette couche joue le rôle d'un actuateur qui se déforme pour permettre la création du jet. Son défaut principal réside dans le fait que la couche en question reste mono-utilisable car les déformations liées à l'interaction avec le laser sont plastiques ou laissent des bulles d'air qui empêchent une réutilisation par la suite. Même si elles ne sont pas sacrificielles au même titre qu'une couche métallique, elles restent à usage unique. [0010] In the article, “Cavitation and shock waves emission on the rigid boundary of water under mid-IR nanosecond laser pulse excitation” AV Pushkin, AS Bychkov, AA Karabutov and FV Potemkin, Laser Phys. Lett. 15 (2018) 065401, the processes of converting light energy into mechanical energy under the excitation of a mid-IR nanosecond laser on a rigid water boundary are described. The strong absorption by water of a Q.-switched Cr:Yb:Ho:YSGG laser (2.85 pm, 6 mJ, 45 ns) allows a rapid energy deposition of ~8 kJ/cm 3 accompanied by strong transitions mechanical. The evolution of shock waves and cavitation bubbles is studied using the technique of ombroscopy and acoustic measurements. For a 6 mJ laser pulse with a fluence of 2.0 J.cm 2 , the conversion into shock wave energy reaches 67%. Most of the shock wave energy (92%) is dissipated as the shock front travels the first 250 pm, and the remaining 8% is transferred to the acoustic far field. The results of the article can be used for the optimization of the laser parameters to carry out micro-machining or the ablation of biological tissues. The article does not describe a LIFT type process for material transfer but a linear absorption process in a liquid to generate cavitation bubbles. The energies studied in this article are 2 orders of magnitude higher than those described in the present invention, as well as the size of the bubbles which is several mm in the article when the present invention describes objects of a few tens to hundreds of microns. The conditions illustrated in this article are therefore very far from those sought for the precise bioprinting of cells by LIFT transfer. [0011] Bio-printing solutions by LIFT can also use a specific interaction layer for the laser but which is not metallic. We talk about BA-LIFT (blister assisted LIFT) or LIFT based on the use of a gelatin layer for example. In all cases, this layer plays the role of an actuator which deforms to allow the creation of the jet. Its main defect lies in the fact that the layer in question remains single-use because the deformations linked to the interaction with the laser are plastic or leave air bubbles which prevent reuse thereafter. Even if they are not sacrificial in the same way as a metal layer, they remain disposable.
Inconvénients de l'art antérieur Disadvantages of the prior art
[0012] Le caractère sacrificiel de la couche métallique représente le principal inconvénient de la bio-impression par laser standard car le donneur a un temps d'utilisation limité lié à l'ablation définitive du métal suite au tir laser. Le changement de donneur est donc nécessairement récurrent et consommateur de temps ce qui est potentiellement rédhibitoire pour un usage en bio-impression où le temps de fabrication des tissus est critique pour leur viabilité. De plus, il oblige une sollicitation de l'utilisateur importante en fonction du nombre d'impressions cellulaires à réaliser et donc du nombre de changements du donneur pour fabriquer un tissu. [0012] The sacrificial nature of the metal layer represents the main drawback of standard laser bio-printing because the donor has a limited use time related to the definitive ablation of the metal following the laser shot. The change of donor is therefore necessarily recurrent and time-consuming, which is potentially prohibitive for use in bio-printing where the manufacturing time of the tissues is critical for their viability. In addition, it requires significant user stress depending on the number of cell impressions to be made and therefore the number of donor changes to manufacture a tissue.
[0013] Les systèmes utilisant une couche d'absorption (généralement métallique) dédiée sont limités à un seuil de génération d'une bulle de cavitation en dessous duquel le processus LIFT ne peut pas se déclencher. De fait, la taille minimale des jets et donc des gouttes déposées est déterminée par ce seuil. [0013] Systems using a dedicated (generally metallic) absorption layer are limited to a cavitation bubble generation threshold below which the LIFT process cannot be triggered. In fact, the minimum size of the jets and therefore of the drops deposited is determined by this threshold.
[0014] Une autre limite des couches sacrificielles concerne la qualité de la couche métallique déposée sur le verre. En effet, celle-ci peut souffrir d'une faible adhésion de l'or au verre, d'une variation d'épaisseur du métal sur la surface de verre ou encore de variation dans le procédé de dépôt du film métallique. La conséquence directe est une disparité des jets générés puisque les conditions initiales ne sont pas identiques en tout point du donneur. Il peut aussi en résulter une délamination de la couche métallique pendant l'usage. La conséquence est une perte de qualité d'impression voire l'obligation de devoir recommencer une nouvelle impression. [0014] Another limitation of the sacrificial layers relates to the quality of the metal layer deposited on the glass. Indeed, the latter may suffer from weak adhesion of the gold to the glass, from a variation in the thickness of the metal on the glass surface or even from variation in the method of depositing the metallic film. The direct consequence is a disparity of the jets generated since the initial conditions are not identical at any point of the donor. It may also result in delamination of the metal layer during use. The consequence is a loss of print quality or even the obligation to start a new print.
[0015] Une autre limite connue réside dans la durée de validité de l'état de métallisation des couches sacrificielles, état qui ne doit pas évoluer dans le temps afin d'assurer une reproductibilité élevée du processus d'impression. [0015] Another known limit lies in the period of validity of the metallization state sacrificial layers, a state which must not change over time in order to ensure high reproducibility of the printing process.
[0016] L'art antérieur, pour lequel le procédé LIFT est mis en œuvre sans couche sacrificielle, est relatif à quatre régimes bien spécifiques qui impliquent : The prior art, for which the LIFT process is implemented without a sacrificial layer, relates to four very specific regimes which involve:
- des effets non linéaires dans le domaine proche IR - nonlinear effects in the near IR range
- des absorbants inclus dans l'encre à imprimer - absorbents included in the printing ink
- une absorption par l'encre à une longueur d'onde supérieure à 3pm - absorption by the ink at a wavelength greater than 3 pm
- le recours à une couche de type blister - the use of a blister type layer
[0017] Les inconvénients du régime non linéaire sont liés au mode d'interaction entre le laser et le matériel à imprimer qui est très sensible à la typologie des encres et à leurs propriétés colloïdales. En effet, cette approche est pertinente pour l'impression de liquides homogènes mais se heurte à des problématiques de reproductibilité majeures (grosses disparités de taille de bulle de cavitation et de taille de jets) lors de l'impression d'encres inhomogènes (colloïdales) comme celles utilisées en bio-impression (cellules). Cette approche n'est donc pas la bonne solution pour l'impression cellulaire. En effet, les résultats obtenus amènent à des patterns imprimés incomplets où seulement 50 à 70 % comporte des cellules. Un autre inconvénient du régime non linaire est lié au besoin de positionnement précis en 3D du point focal afin d'obtenir des jets reproductibles. [0017] The drawbacks of the non-linear regime are linked to the mode of interaction between the laser and the material to be printed, which is very sensitive to the type of inks and to their colloidal properties. Indeed, this approach is relevant for printing homogeneous liquids but comes up against major reproducibility issues (large disparities in cavitation bubble size and jet size) when printing inhomogeneous (colloidal) inks. such as those used in bio-printing (cells). So this approach is not the right solution for cellular printing. Indeed, the results obtained lead to incomplete printed patterns where only 50 to 70% comprises cells. Another disadvantage of the nonlinear regime is related to the need for precise 3D positioning of the focal point in order to obtain reproducible jets.
[0018] Concernant le recours à des absorbants, le résultat est beaucoup plus performant d'un point de vue reproductibilité et qualité d'impression. Toutefois, cette approche est contraignante car elle impose l'ajout de molécules généralement non bio-compatibles au sein de l'encre à imprimer. Là encore, cette approche est limitante pour la bio-impression de tissus car susceptible d'entraîner des effets délétères sur les fonctions cellulaires. [0018] Regarding the use of absorbents, the result is much more efficient from a reproducibility and print quality point of view. However, this approach is restrictive because it imposes the addition of generally non-biocompatible molecules within the ink to be printed. Here again, this approach is limiting for the bio-printing of tissues because it is likely to lead to deleterious effects on cellular functions.
Solution apportée par l'invention Solution provided by the invention
[0019] Afin de remédier aux inconvénients de l'art antérieur, la présente invention concerne selon son acception la plus générale un équipement d'impression additive comportant : une source laser pulsée produisant un faisceau et focalisée sur le matériel à imprimer, un donneur à partir duquel un matériel biologique est imprimé, et un substrat receveur cible qui recueille le matériel imprimé. In order to remedy the drawbacks of the prior art, the present invention relates, in its most general sense, to additive printing equipment comprising: a pulsed laser source producing a beam and focused on the material to be printed, a donor to from which a biological material is imprinted, and a target recipient substrate which collects the imprinted material.
Ledit donneur étant constitué par une lame recouverte, dans la zone d'interaction du faisceau laser, par un film liquide destiné à contenir des inhomogénéités transférables, caractérisé en ce que ledit laser émet un faisceau dont la longueur d'onde est comprise entre 2 pm et 3.2 pm ladite lame étant transparente ou faiblement absorbante à la longueur d'onde dudit faisceau laser. Said donor being constituted by a coated plate, in the interaction zone of the beam laser, by a liquid film intended to contain transferable inhomogeneities, characterized in that said laser emits a beam whose wavelength is between 2 pm and 3.2 pm, said plate being transparent or weakly absorbing at the wavelength of said laser beam.
[0020] Cette solution a pour conséquence : [0020] This solution has the following consequences:
Une interaction entre ledit laser et ledit fluide se basant sur une absorption linéaire, directe et totale de l'énergie laser par le liquide, An interaction between said laser and said fluid based on a linear, direct and total absorption of the laser energy by the liquid,
La création d'une bulle de cavitation par des effets d'élévation de température et de pression localisée au niveau du point d'absorption The creation of a cavitation bubble by the effects of temperature rise and localized pressure at the level of the absorption point
L'entraînement par ladite bulle de cavitation d'un flux de matière qui par expansion ou rétraction modifie la surface libre dudit film fluide jusqu'à la création d'un jet, le transfert de matière par ledit jet depuis le film fluide jusqu'à la surface d'une cible. Une reproductibilité élevée de la taille de bulle de cavitation générée, du jet et de la goutte déposée étant liés linéairement à l'énergie absorbée par ledit fluide la possibilité de créer des impressions à haute résolution par le biais de gouttes dont le volume est dans le domaine de quelques dizaines de picolitres à plusieurs nanolitres en régime stable. The entrainment by said cavitation bubble of a flow of material which, by expansion or retraction, modifies the free surface of said fluid film until the creation of a jet, the transfer of material by said jet from the fluid film to the surface of a target. A high reproducibility of the size of the cavitation bubble generated, of the jet and of the deposited drop being linearly related to the energy absorbed by said fluid the possibility of creating high resolution impressions through drops whose volume is in the range from a few tens of picoliters to several nanoliters in steady state.
[0021] L'absorption linéaire suit la loi de Beer Lambert qui établit que l'absorbance A d'une solution est proportionnelle, d'une part, à sa concentration c et, d'autre part, à la longueur b du trajet parcouru par la lumière dans la solution : A = £ c où : Linear absorption follows Beer Lambert's law which establishes that the absorbance A of a solution is proportional, on the one hand, to its concentration c and, on the other hand, to the length b of the path traveled. by light in the solution: A = £ c where:
• e est le coefficient d'extinction molaire ou d'absorptivité propre à l'entité chimique (constante) ; • e is the molar extinction or absorptivity coefficient specific to the chemical entity (constant);
• b est la longueur du trajet parcouru par la lumière dans le milieu considéré ; • b is the length of the path traveled by the light in the medium considered;
• C désigne la concentration de l'entité chimique. • C designates the concentration of the chemical entity.
[0022] L'absorption du laser dans le domaine de longueur d'onde entre 2 et 3,2pm est proportionnelle à la quantité de liquide présente au niveau du plan d'illumination, on parle alors d'absorption linéaire. La longueur d'onde privilégiée est autour de 2.9pm car elle correspond au pic d'absorption du liquide le plus élevé comme on peut le voir sur la figure 2. L'intérêt de l'absorption directe linéaire est d'ouvrir un champ nouveau d'absorption correspondant à des petites énergies laser permettant de créer des petites bulles et donc des petits jets, dans un régime où la bulle et le jet générés sont suffisants pour réaliser une impression. De fait, l'impression à 2.9pm ouvre la voie à une large palette de tailles de gouttes imprimées du picolitre au nanolitre. [0022] The absorption of the laser in the wavelength range between 2 and 3.2 μm is proportional to the quantity of liquid present at the level of the illumination plane, one then speaks of linear absorption. The preferred wavelength is around 2.9pm because it corresponds to the highest liquid absorption peak as can be seen in Figure 2. The advantage of linear direct absorption is to open up a new field absorption corresponding to small laser energies making it possible to create small bubbles and therefore small jets, in a regime where the bubble and the jet generated are sufficient to make an impression. In fact, printing at 2.9pm opens the way to a wide range of printed drop sizes from picolitre to nanolitre.
[0023] L'absorption directe signifie qu'il n'y a pas d'élément ou de processus intermédiaire dans l'interaction entre le laser et le milieu liquide. De fait, il n'y pas d'éléments absorbants ajoutés dans la solution, ni de couche sacrificielle métallique qui permette d'initier l'absorption, ni d'effets optiques complexes, ni d'effets physico-chimiques ou électromagnétiques pour initier l'absorption. Celle-ci est directement réalisée par absorption du liquide qu'il soit aqueux (molécules d'eau) ou non aqueux (polymères en solution, etc.). [0024] L'absorption totale signifie qu'il n'y a pas de photon résiduel du laser après une certaine épaisseur traversée par ledit laser dans le fluide à imprimer. Cette épaisseur a été estimée expérimentalement et est inférieure à 5 pm. On peut donc considérer que le fluide présent dans cette faible épaisseur joue un rôle équivalent à celui d'une couche sacrificielle sans l'être véritablement. Elle permet notamment de confiner l'absorption du laser à l'interface liquide - substrat, sur une zone de quelques microns, comme on le fait habituellement avec une couche sacrificielle métallique qui elle fait généralement quelques dizaines de nanomètres. Cela permet d'obtenir un positionnement de la bulle de cavitation similaire à celui qui est obtenu classiquement par création d'un plasma au sein d'une couche sacrificielle. Ainsi, la dynamique de la bulle et du jet est très similaire à celle d'un système basé sur l'utilisation d'une couche sacrificielle avec l'avantage de pouvoir réutiliser le donneur autant de fois que nécessaire puisque la quantité de liquide présente dans celui-ci est importante et renouvelable. L'absorption totale du laser sur les premiers microns du liquide assure une protection du reste du liquide vis à vis des photons utilisés et donc des cellules qui s'y trouvent. C'est une alternative très intéressante pour la viabilité cellulaire par comparaison à d'autres solutions où des lasers UV (ionisants) sont utilisés pour faire de la bio-impression par laser par exemple. Un autre avantage réside dans le fait que l'absorption peut être vue comme un processus en 2D qui nécessite une précision moindre pour le positionnement du plan focal laser. De fait, ces conditions d'interaction favorisent une très haute viabilité des cellules dans le cas spécifique de la bio-impression puisque celles-ci n'auront pas d'interaction directe avec les photons du laser. L'absorption totale a également un impact sur la sécurité laser pour l'utilisateur d'un tel système. En effet, l'absorption par le liquide ou par les interfaces en verre ou autre substrat garantie qu'aucun photon du laser ne sera en mesure d'atteindre l'utilisateur. Les moyens de sécurité des personnes utilisant le système peuvent donc être aménagés dans ce contexte. [0023] Direct absorption means that there is no intermediate element or process in the interaction between the laser and the liquid medium. In fact, there are no absorbent elements added to the solution, nor a metallic sacrificial layer which makes it possible to initiate absorption, nor complex optical effects, nor physico-chemical or electromagnetic effects to initiate absorption. 'absorption. This is carried out directly by absorption of the liquid whether it is aqueous (water molecules) or non-aqueous (polymers in solution, etc.). [0024] Total absorption means that there is no residual photon of the laser after a certain thickness through which said laser passes in the fluid to be printed. This thickness has been estimated experimentally and is less than 5 μm. It can therefore be considered that the fluid present in this small thickness plays a role equivalent to that of a sacrificial layer without really being so. It makes it possible in particular to confine the absorption of the laser to the liquid-substrate interface, over an area of a few microns, as is usually done with a metallic sacrificial layer which is generally a few tens of nanometers. This makes it possible to obtain a positioning of the cavitation bubble similar to that obtained conventionally by creating a plasma within a sacrificial layer. Thus, the dynamics of the bubble and the jet are very similar to those of a system based on the use of a sacrificial layer with the advantage of being able to reuse the donor as many times as necessary since the quantity of liquid present in this one is important and renewable. The total absorption of the laser on the first microns of the liquid ensures protection of the rest of the liquid with respect to the photons used and therefore the cells which are there. It is a very interesting alternative for cell viability compared to other solutions where UV (ionizing) lasers are used to make laser bio-printing for example. Another advantage is that absorption can be viewed as a 2D process that requires less precision for positioning the laser focal plane. In fact, these interaction conditions favor a very high viability of the cells in the specific case of bio-printing since these will not have a direct interaction with the photons of the laser. Total absorption also has an impact on laser safety for the user of such a system. Indeed, the absorption by the liquid or by the interfaces in glass or other substrate guarantee that no photon of the laser will be able to reach the user. The means of safety of the people using the system can therefore be arranged in this context.
[0025] L'impression à 2.9pm se base donc sur l'interaction du laser avec quelques pm d'épaisseur d'un fluide homogène qui garantit une grande reproductibilité de l'interaction laser - matière avec pour conséquence une grande stabilité des bulles et des jets générés. Une telle reproductibilité est très importante dans le domaine de la bio-impression notamment à visée industrielle et clinique. [0025] Printing at 2.9 μm is therefore based on the interaction of the laser with a few μm of thickness of a homogeneous fluid which guarantees high reproducibility of the laser-material interaction, resulting in high stability of the bubbles and jets generated. Such reproducibility is very important in the field of bio-printing, in particular for industrial and clinical purposes.
[0026] L'utilisation d'une cartouche à rechargement continu faisant office de substrat donneur et d'un bras robotisé pour pipeter automatiquement l'encre et la disposer sur la surface ouverte de ladite cartouche a été décrite dans l'art antérieur (FR3093944). Elle permet d'automatiser le processus d'impression afin de réduire l'intervention humaine mais se heurte à l'ablation successive de la couche d'or à chaque étape d'impression, obligeant un changement régulier de ladite cartouche. L'utilisation conjointe de cette cartouche avec un système travaillant à 2.9pm tel que décrit dans cette invention permettrait d'imprimer des grandes surfaces / épaisseur d'objets (tissus) sans intervention humaine puisque la cartouche serait utilisable sur un très grand nombre de processus d'impression sans avoir à la changer. Le liquide technique de pré-mouillage servirait alors de milieu absorbant dédié, capacité qui n'est pas suggérée dans le brevet en question. [0026] The use of a cartridge with continuous reloading acting as a donor substrate and of a robotic arm for automatically pipetting the ink and placing it on the open surface of said cartridge has been described in the prior art (FR3093944 ). It makes it possible to automate the printing process in order to reduce human intervention, but comes up against the successive removal of the gold layer at each printing step, requiring a regular change of said cartridge. The joint use of this cartridge with a system working at 2.9pm as described in this invention would make it possible to print large surfaces / thickness of objects (tissues) without human intervention since the cartridge would be usable on a very large number of processes. printing without having to change it. The technical pre-wetting liquid would then serve as a dedicated absorbent medium, a capability which is not suggested in the subject patent.
[0027] On pourrait aussi utiliser la cartouche décrite dans la demande (FR3063931A1) qui propose une mise en œuvre où l'encre à imprimer entre et sort de la zone d'impression ouverte du donneur par des tubes fluidiques. Le laser est tiré dans ladite zone ouverte qui peut être de forme circulaire (similaire à ce qui se pratique avec les systèmes d'impression par laser conventionnels) ou de forme plus spéciale comme un canal fluidique simple ou multiple. Ainsi, quelle que soit la forme de la zone ouverte, les tirs lasers peuvent réalisés sur un même point ou sur des lignes de points autant de fois que nécessaire sans avoir à changer la cartouche. [0028] La présente invention concerne selon son acception la plus générale un équipement d'impression additive comportant : une source laser pulsée produisant un faisceau et focalisé sur le matériel à imprimer, un donneur dépourvu de couche sacrificielle, à partir duquel un matériel biologique est imprimé, et un substrat receveur cible qui recueille le matériel imprimé Ledit donneur étant constitué par une lame recouverte, dans la zone d'interaction du faisceau laser, par un film liquide destiné à contenir des inhomogénéités transférables, caractérisé en ce que ledit laser émet un faisceau dont la longueur d'onde est comprise entre 2 pm et 3,2 pm ladite lame étant transparente ou faiblement absorbante à la longueur d'onde dudit faisceau laser. One could also use the cartridge described in the application (FR3063931A1) which proposes an implementation where the ink to be printed enters and leaves the open printing zone of the donor by fluidic tubes. The laser is fired into said open area which may be circular in shape (similar to what is practiced with conventional laser printing systems) or more special in shape such as a single or multiple fluidic channel. Thus, whatever the shape of the open area, the laser shots can be fired on the same point or on lines of points as many times as necessary without having to change the cartridge. The present invention relates, in its most general sense, to additive printing equipment comprising: a pulsed laser source producing a beam and focused on the material to be printed, a donor without a sacrificial layer, from which a biological material is printed, and a target receiver substrate which collects the printed material. Said donor being constituted by a covered plate, in the interaction zone of the beam laser, by a liquid film intended to contain transferable inhomogeneities, characterized in that the said laser emits a beam whose wavelength is between 2 μm and 3.2 μm, the said plate being transparent or weakly absorbing at the length of wave of said laser beam.
Selon des variantes avantageuses : la longueur d'onde d'émission du laser présente un pic à 2,9 pm ± 0,3 pm l'épaisseur du liquide dans laquelle l'absorption totale de l'énergie laser est réalisée est de l'ordre de quelques microns et inférieure à 5pm la focalisation du laser à l'interface du donneur et du liquide peut être modifiée de façon à permettre variation de la taille transversale de la zone d'absorption avec pour effet de moduler la taille des bulles et des jets générés et donc des gouttes déposées sur la cible l'impression par laser à 2.9pm est parfaitement adaptée à l'utilisation d'une cartouche à rechargement continu en fluide une cartouche à rechargement continu utilisant un film de prémouillage pourrait utiliser l'encre technique de prémouillage comme milieu absorbant pour la longueur d'onde à 2.9pm le fluide est disposé dans un réservoir l'impression se fait de bas vers le haut en sens opposé à la force de gravité Les performances associées à cet équipement permettent d'assurer les caractéristiques suivantes : la taille de la bulle de cavitation et la taille du jet associé sont linéairement dépendantes d'une part de l'énergie laser utilisée qui est typiquement dans le domaine du pJ à quelques dizaines de pJ et d'autre part de l'épaisseur du liquide qui peut varier de quelques microns à quelques centaines de microns les bulles de cavitation ont un grand spectre de tailles allant de quelques microns à plusieurs centaines de microns les gouttes déposées ont un grand spectre de tailles allant de très petits volumes de quelques picolitres à de très grands volumes d'un nanolitre à un microlitre l'absorption directe du liquide assure une condition d'impression constante pour un jeu de paramètres d'impression donnés avec pour conséquence une très grande stabilité des jets générés ouvrant la voie à une très importante reproductibilité du transfert du fluide vers une cible According to advantageous variants: the emission wavelength of the laser has a peak at 2.9 pm ± 0.3 pm the thickness of the liquid in which the total absorption of the laser energy is achieved is order of a few microns and less than 5 μm the focusing of the laser at the interface of the donor and the liquid can be modified so as to allow variation of the transverse size of the absorption zone with the effect of modulating the size of the bubbles and the jets generated and therefore drops deposited on the target laser printing at 2.9pm is perfectly suited to the use of a cartridge with continuous fluid refill a cartridge with continuous refill using a pre-wetting film could use technical ink pre-wetting as an absorbent medium for the wavelength at 2.9 pm the fluid is placed in a tank the impression is made from bottom to top in the opposite direction to the force of gravity The performances associated with this equipment make it possible to ensure the following characteristics: the size of the cavitation bubble and the size of the associated jet are linearly dependent on the one hand on the laser energy used which is typically in the range from pJ to a few tens of pJ and on the other hand on the thickness of the liquid, which can vary from a few microns to a few hundred microns the cavitation bubbles have a large spectrum of sizes ranging from a few microns to several hundred microns the deposited drops have a large spectrum of sizes ranging from very small volumes of a few picoliters to very large volumes from a nanoliter to a microliter the direct absorption of the liquid ensures a constant printing condition for a given set of printing parameters with the consequence of a very large stability of the jets generated opening the way to a very high reproducibility of fluid transfer to a target
[0029] L'invention concerne aussi une méthode d'impression additive par une source laser pulsée produisant un faisceau en direction d'un donneur dépourvu de couche sacrificielle à partir duquel un matériel biologique est imprimé, vers un substrat receveur qui recueille le matériel imprimé, ledit donneur étant constitué par une lame recouverte, dans la zone d'interaction du faisceau laser, par un film de liquide destiné à contenir des inhomogénéités transférables, caractérisé en ce que ledit laser émet un faisceau dont la longueur d'onde est comprise entre 2 pm et 3,2 pm et en ce que ladite lame est transparente ou faiblement absorbante à la longueur d'onde dudit faisceau laser. The invention also relates to a method of additive printing by a pulsed laser source producing a beam in the direction of a donor without a sacrificial layer from which a biological material is printed, towards a receiving substrate which collects the printed material. , said donor being constituted by a plate covered, in the zone of interaction of the laser beam, by a film of liquid intended to contain transferable inhomogeneities, characterized in that said laser emits a beam whose wavelength is between 2 μm and 3.2 μm and in that said plate is transparent or weakly absorbent at the wavelength of said laser beam.
[0030] Avantageusement, le fluide à imprimer provient d'une cartouche à rechargement continu. [0030] Advantageously, the fluid to be printed comes from a cartridge with continuous reloading.
[0031] De préférence l'acheminement de l'encre vers la cartouche est automatisé pour éviter toute intervention humaine pendant la phase d'impression. [0031] Preferably, the routing of the ink to the cartridge is automated to avoid any human intervention during the printing phase.
Description détaillée d'un exemple non limitatif de réalisation Detailed description of a non-limiting embodiment
[0032] D'autres caractéristiques et avantages ressortiront de la description qui va suivre de l'invention, description donnée à titre d'exemple uniquement, se référant aux dessins annexés sur lesquels : Other characteristics and advantages will emerge from the following description of the invention, description given by way of example only, with reference to the accompanying drawings in which:
[FIG. 1] La figure 1 représente une vue schématique d'un système LIFT selon l'invention travaillant à 2.9pm, [FIG. 1] Figure 1 shows a schematic view of a LIFT system according to the invention working at 2.9pm,
[FIG. 2] La figure 2 représente le spectre d'absorption de l'eau, [FIG. 2] Figure 2 shows the absorption spectrum of water,
[FIG. 3a] La figure 3a représente une courbe de la stabilité des jets obtenus par ombroscopie, [FIG. 3b] La figure 3b représente une courbe de la hauteur de jet mesurée sur 50 jets consécutifs, [FIG. 3a] FIG. 3a represents a curve of the stability of the jets obtained by ombroscopy, [FIG. 3b] Figure 3b represents a curve of the jet height measured on 50 consecutive jets,
[FIG. 4] La figure 4 représente une courbe de la dépendance des hauteurs de jets en fonction du temps et de l'énergie laser, [FIG. 4] Figure 4 represents a curve of the dependence of jet heights as a function of time and laser energy,
[FIG. 5] La figure 5 représente un graphe des caractéristiques des gouttes imprimées sur substrat receveur en fonction de la distance au receveur, [FIG. 5] Figure 5 represents a graph of the characteristics of the drops printed on the receiver substrate as a function of the distance to the receiver,
[FIG. 6] La figure 6 représente la géométrie des gouttes imprimées par la technique LIFT direct à 2.9 pm sur un substrat sans couche sacrificielle. Description d'un premier exemple d'équipement [FIG. 6] Figure 6 represents the geometry of the drops printed by the direct LIFT technique at 2.9 μm on a substrate without sacrificial layer. Description of a first example of equipment
[0033] La figure 1 représente une vue schématique d'un premier exemple d'équipement. [0034] Le laser (1) est une source pulsée en régime nanoseconde, dont la durée d'impulsion est de 10 ns, émettant entre 2,6 pm et 3,2 pm de longueur d'onde, dans un régime d'énergie pouvant atteindre lOOpJ, avec une cadence de tirs réglable entre 1 Hz et lOKHz. Figure 1 shows a schematic view of a first example of equipment. The laser (1) is a pulsed source in the nanosecond regime, the pulse duration of which is 10 ns, emitting between 2.6 μm and 3.2 μm in wavelength, in an energy regime capable of reaching 100pJ, with an adjustable rate of fire between 1Hz and 1OKHz.
[0035] Le faisceau laser est mis en forme à travers un système optique (2) composé de deux lentilles afin de le collimater et lui donner le diamètre souhaité. Les composants optiques doivent être en ZnSe, CaF2, Infrasil ou tout autre matériau compatible avec un usage dans le domaine de longueur d'onde du milieu IR (de 2 à 3,2 pm). The laser beam is shaped through an optical system (2) composed of two lenses in order to collimate it and give it the desired diameter. The optical components must be made of ZnSe, CaF2, Infrasil or any other material compatible with use in the wavelength range of the IR medium (from 2 to 3.2 pm).
[0036] Le scanner (3) assure une orientation angulaire du faisceau selon deux axes perpendiculaires. Il est par exemple constitué par deux miroirs actionnés par un actionneur électromagnétique, par exemple un scanner commercialisé par la société SCANLAB (nom commercial) sous la référence « SCANcube 14 ». Les miroirs de ce scanner doivent être recouverts d'une couche d'or pour assurer une réflexion totale du laser dans le domaine de longueur d'onde du milieu IR (de 2 à 3,2 pm). The scanner (3) ensures an angular orientation of the beam along two perpendicular axes. It is for example constituted by two mirrors actuated by an electromagnetic actuator, for example a scanner marketed by the company SCANLAB (trade name) under the reference “SCANcube 14”. The mirrors of this scanner must be covered with a layer of gold to ensure total reflection of the laser in the wavelength range of the IR medium (from 2 to 3.2 μm).
[0037] Le laser (1) est ensuite focalisé par un système optique de focalisation dédié (4) sur le donneur (5) sur lequel est déposé l'encre à imprimer (6). Habituellement, le système de focalisation dans les systèmes LIFT standards est basé sur des systèmes optiques complexes appelés F-Theta comportant un grand nombre de lentilles. Malheureusement, il n'existe pas d'équivalent dans le domaine mid-IR visé par cette invention. Des solutions sur mesure peuvent alors répondre à cette problématique : The laser (1) is then focused by a dedicated focusing optical system (4) on the donor (5) on which the ink to be printed (6) is deposited. Usually, the focusing system in standard LIFT systems is based on complex optical systems called F-Theta with a large number of lenses. Unfortunately, there is no equivalent in the mid-IR domain covered by this invention. Tailor-made solutions can then respond to this problem:
- F-théta sur mesure compatible avec la longueur d'onde à 2.9pm (solution complexe et onéreuse) - Customized F-theta compatible with the wavelength at 2.9pm (complex and expensive solution)
- Objectif télécentrique constitué de 2 lentilles sur mesure compatible avec la longueur d'onde à 2.9pm (solution plus simple et moins onéreuse) - Telecentric lens consisting of 2 custom-made lenses compatible with the 2.9pm wavelength (simpler and less expensive solution)
- Lentille ou objectif simple disposé avant le scanner: dans ce cas, on utiliserait une solution de type scanner 3D où la focalisation se ferait en amont du scanner et nécessiterait l'utilisation d'une adaptation selon la direction Z de la focale dans le champ visé. - Simple lens or objective placed before the scanner: in this case, a 3D scanner type solution would be used where the focusing would be done before the scanner and would require the use of an adaptation along the Z direction of the focal length in the field targeted.
[0038] Le film (6) est disposé en face d'un substrat receveur cible (7) suffisamment transparent à la longueur d'onde du laservers lequel sont transférées les cellules ou particules, lors du déclenchement d'un faisceau laser (1) impulsionnel. [0039] La figure 2 illustre la capacité d'absorption de l'eau en fonction de la longueur d'onde d'illumination et on peut y voir clairement le pic d'absorption à 2.9pm visé par la présente invention dans sa mise en œuvre particulière. The film (6) is arranged opposite a target receiver substrate (7) sufficiently transparent to the wavelength of the laser to which the cells or particles are transferred, when a laser beam (1) is triggered. impulsive. [0039] Figure 2 illustrates the absorption capacity of water as a function of the illumination wavelength and one can clearly see there the absorption peak at 2.9 pm targeted by the present invention in its implementation. particular work.
[0040] La figure 3a représente la courbe de stabilité des jets mesurée par une technique d'ombroscopie mise en œuvre dans l'exemple de réalisation décrit en figure 1. On peut aisément conclure que la hauteur des jets reste la même sur un nombre important de jets consécutifs. FIG. 3a represents the stability curve of the jets measured by a ombroscopy technique implemented in the example embodiment described in FIG. 1. It can easily be concluded that the height of the jets remains the same over a large number consecutive throws.
[0041] La figure 3b représente la courbe de la hauteur de jet mesurée sur 50 jets consécutifs à partir des données d'ombroscopie. On constate que la répétabilité des hauteurs de jet est supérieure à 90 %. Cette stabilité est un gage de reproductibilité des impressions, caractéristique absolument nécessaire pour utiliser la bio-impression dans le domaine industriel ou clinique. Elle a été obtenue sur le système décrit en figure 1. [0041] FIG. 3b represents the curve of the jet height measured over 50 consecutive jets from shadow data. It can be seen that the repeatability of the jet heights is greater than 90%. This stability is a guarantee of print reproducibility, an absolutely necessary characteristic for using bio-printing in the industrial or clinical field. It was obtained on the system described in figure 1.
[0042] La figure 4 représente une courbe de la dépendance des hauteurs de jets en fonction du temps et de l'énergie laser obtenue avec le système décrit sur la figure 1. La dépendance directe entre énergie et hauteur de jet est clairement observable et apporte la preuve du caractère linéaire de l'absorption à 2.9pm. Temporellement, on constate que les jets ont une « durée de vie » de quelques centaines de ps ce qui est standard dans le domaine du LIFT. On est donc sur des temps d'impression courts compatibles avec l'impression de tissus biologiques qui doivent rester viables pendant cette phase. [0042] FIG. 4 represents a curve of the dependence of the jet heights as a function of time and of the laser energy obtained with the system described in FIG. 1. The direct dependence between energy and jet height is clearly observable and provides the proof of the linear character of the absorption at 2.9 pm. Temporally, we see that the jets have a "lifetime" of a few hundred ps, which is standard in the field of LIFT. We are therefore on short printing times compatible with the printing of biological tissues which must remain viable during this phase.
[0043] La figure 5 représente un graphe des caractéristiques des gouttes imprimées sur substrat receveur en fonction de la distance audit receveur dans le cadre de l'exemple de réalisation décrit en figure 1. La taille des gouttes suit la même logique que celle décrite sur les jets à savoir une dépendance linéaire à l'énergie laser utilisée et une capacité à obtenir des tailles reproductibles sur une échelle importante. Ces résultats corroborent les performances décrites de façon générale sur cette invention. [0043] FIG. 5 represents a graph of the characteristics of the drops printed on the receiver substrate as a function of the distance from said receiver in the context of the embodiment described in FIG. 1. The size of the drops follows the same logic as that described in the jets, namely a linear dependence on the laser energy used and an ability to obtain reproducible sizes on a large scale. These results corroborate the performances generally described for this invention.
[0044] La figure 6 est la preuve finale de la capacité du LIFT direct à 2.9 pm à imprimer des gouttes sur un substrat sans couche sacrificielle avec des résultats obtenus sur le montage décrit en figure 1. L'impression concerne un milieu homogène dans cette illustration. Des résultats similaires ont été obtenus sur des milieux colloïdaux avec microbilles ou cellules humaines. Elle illustre la topographie de la cible après impression avec quatre niveaux d'énergie différents. Description d'un second exemple d'équipement [0044] Figure 6 is the final proof of the ability of the direct LIFT at 2.9 μm to print drops on a substrate without sacrificial layer with results obtained on the assembly described in Figure 1. The printing relates to a homogeneous medium in this drawing. Similar results were obtained on colloidal media with microbeads or human cells. It illustrates the topography of the target after printing with four different energy levels. Description of a second example of equipment
[0045] Dans une autre configuration, la solution peut être réalisée sans recours à un scanner 2D ou 3D. Dans ce cas, la ligne optique est simplifiée car elle intègre uniquement un système de mise en forme de faisceau et une optique de focalisation. Dans ce cas, on considère que l'encre déposée au niveau du donneur sera suffisamment perturbée par chaque tir laser entraînant une bulle puis un jet pour permettre une forme de renouvellement de l'encre au niveau de la zone du tir laser. Cette configuration peut être imaginée à la fois dans le cas d'une architecture d'impression de bas vers le haut en sens opposé à la force de gravité comme de haut vers le bas dans le sens de la gravité. Le donneur peut être constitué d'un film fluide comme d'un réservoir. In another configuration, the solution can be achieved without using a 2D or 3D scanner. In this case, the optical line is simplified because it only integrates a beam shaping system and focusing optics. In this case, it is considered that the ink deposited at the level of the donor will be sufficiently disturbed by each laser shot resulting in a bubble then a jet to allow a form of renewal of the ink at the level of the zone of the laser shot. This configuration can be imagined both in the case of a printing architecture from bottom to top in the direction opposite to the force of gravity as well as from top to bottom in the direction of gravity. The donor may consist of a fluid film such as a reservoir.
Spécificités du domaine de longueur d'ondes Milieu IR Specificities of the Medium IR wavelength range
[0046] La longueur d'onde de 2,9 pm du faisceau laser ne permet pas l'utilisation d'optiques standards. Les composants optiques doivent donc se baser sur d'autres matériaux non absorbants à ces longueurs d'onde et qui sont plus dispendieux. Il faut donc utiliser de manière préférentielle des matériaux transparents comme le CaF2 , le ZnSe ou l'Infrasil (nom commercial) même si on peut aussi utiliser comme substrat donneur de fines lames de silice au prix d'une perte d'énergie. Les lentilles utilisées par contre sont des lentilles de CaF2 et les miroirs doivent être en or protégé. The 2.9 μm wavelength of the laser beam does not allow the use of standard optics. The optical components must therefore be based on other materials which are not absorbent at these wavelengths and which are more expensive. It is therefore preferable to use transparent materials such as CaF2 , ZnSe or Infrasil (trade name) even if it is also possible to use as donor substrate thin layers of silica at the cost of a loss of energy. The lenses used, on the other hand, are CaF2 lenses and the mirrors must be in protected gold.
[0047] L'émission à 2.9 pm est caractérisée par une forte absorption de ce rayonnement par l'eau. Cette caractéristique pourrait alors faire entrer ce type de rayonnement dans la catégorie des lasers dit de sécurité oculaire comme les lasers à erbium émettant à 1500 nm. C'est donc la cornée qui absorbe ce rayonnement. Cependant, le coefficient d'absorption de l'eau à 2.9 pm (10000 cm4) est beaucoup plus important qu'à 1.5pm (30 cm-1), l'absorption se fait donc sur des distances inférieures à 2 à 3pm de longueur d'onde contre 700 pm à 1.5pm de longueur d'onde. Les risques d'endommagement de la cornée sont donc plus importants à 3pm qu'à 1.5 pm mais beaucoup moins concernant la rétine. The emission at 2.9 μm is characterized by strong absorption of this radiation by the water. This characteristic could then bring this type of radiation into the category of so-called eye safety lasers such as erbium lasers emitting at 1500 nm. It is therefore the cornea that absorbs this radiation. However, the water absorption coefficient at 2.9 pm (10000 cm 4 ) is much greater than at 1.5 pm (30 cm -1 ), absorption therefore takes place over distances less than 2 to 3 pm from wavelength versus 700pm to 1.5pm wavelength. The risks of damage to the cornea are therefore greater at 3 pm than at 1.5 pm, but much less concerning the retina.
[0048] La mise en œuvre d'un système travaillant à 2.9pm va être sensible à l'environnement, notamment la quantité de vapeur d'eau présente dans l'air. Pour y remédier, une variante de l'équipement peut être imaginée comme un système fermé vis à vis de l'extérieur afin d'éviter les changements d'humidité de l'air. Selon une autre variante, on peut aussi prévoir un gaz neutre présent au cœur du système optique. Le module laser peut donc être constitué d'une enveloppe permettant de protéger le faisceau de changement d'hygrométrie voir d'y placer le gaz neutre évoqué auparavant. L'intérêt d'une telle configuration est d'assurer une stabilité de la quantité d'énergie laser arrivant sur l'échantillon. [0048] The implementation of a system working at 2.9 pm will be sensitive to the environment, in particular the quantity of water vapor present in the air. To remedy this, a variant of the equipment can be imagined as a closed system with respect to the outside in order to avoid changes in air humidity. According to another variant, it is also possible to provide an inert gas present at the heart of the optical system. The laser module can therefore consist of an envelope making it possible to protect the hygrometry change beam or even to place the inert gas mentioned above therein. The interest of such a configuration is to ensure stability of the quantity of laser energy arriving on the sample.
Selon des variantes : According to variants:
- la modification de la focalisation du laser à l'interface verre liquide par changement de plan focal va permettre de faire varier la taille transversale de la zone d'absorption avec pour effet de moduler la taille des bulles et des jets générés et donc des gouttes déposées sur la cible.- the modification of the focusing of the laser at the liquid glass interface by changing the focal plane will make it possible to vary the transverse size of the absorption zone with the effect of modulating the size of the bubbles and the jets generated and therefore of the drops deposited on the target.
- le système optique permettant la propagation du faisceau laser jusqu'au donneur comporte un scanner 2D ou 3D permettant d'orienter ledit faisceau laser afin de créer un pattern complet de gouttes imprimées par balayage dudit faisceau laser sur ledit donneur dont la surface est d'au moins 0,5 cm2. - the optical system allowing the propagation of the laser beam to the donor comprises a 2D or 3D scanner making it possible to orient said laser beam in order to create a complete pattern of drops printed by scanning said laser beam on said donor whose surface is at least 0.5 cm2.
- le système optique ne comporte pas de scanner et tire directement sur le donneur, chaque tir permettant de renouveler la zone d'interaction au sein dudit donneur par les mouvements induits dans le film d'encre par la bulle de cavitation et le jet. - the optical system does not include a scanner and shoots directly on the donor, each shot making it possible to renew the interaction zone within said donor by the movements induced in the film of ink by the cavitation bubble and the jet.
- le système optique est intégré à un module permettant une régulation de l'hygrométrie.- the optical system is integrated into a module allowing humidity regulation.
- le système optique est intégré dans une enceinte fermée dans laquelle un gaz neutre a été introduit afin d'éviter toute d'absorption par vapeur d'eau - the optical system is integrated in a closed enclosure in which a neutral gas has been introduced in order to avoid any absorption by water vapor
- l'acheminement de l'encre vers la cartouche est robotisé. - the routing of the ink towards the cartridge is robotized.
- la taille des gouttes déposées est linéairement dépendante de l'énergie laser et de l'épaisseur du fluide à imprimer - the size of the drops deposited is linearly dependent on the laser energy and the thickness of the fluid to be printed

Claims

Revendications Claims
1 - Équipement d'impression additive comportant : une source laser pulsée produisant un faisceau et focalisé sur le matériel à imprimer, un donneur dépourvu de couche sacrificielle, à partir duquel un matériel biologique est imprimé, et un substrat receveur cible qui recueille le matériel imprimé Ledit donneur étant constitué par une lame recouverte, dans la zone d'interaction du faisceau laser, par un film liquide destiné à contenir des inhomogénéités transférables, caractérisé en ce que ledit laser émet un faisceau dont la longueur d'onde est comprise entre 2 pm et 3,2 pm ladite lame étant transparente ou faiblement absorbante à la longueur d'onde dudit faisceau laser ledit liquide présente une absorption directe totale sur une épaisseur inférieure de 5pm. 1 - Additive printing equipment comprising: a pulsed laser source producing a beam and focused on the material to be printed, a donor without a sacrificial layer, from which a biological material is printed, and a target receiver substrate which collects the printed material Said donor being constituted by a plate covered, in the zone of interaction of the laser beam, by a liquid film intended to contain transferable inhomogeneities, characterized in that said laser emits a beam whose wavelength is between 2 μm and 3.2 μm, said plate being transparent or weakly absorbing at the wavelength of said laser beam, said liquid has total direct absorption over a thickness less than 5 μm.
2 - Équipement d'impression additive selon la revendication 1 caractérisé en ce que ledit liquide est aqueux. 2 - additive printing equipment according to claim 1 characterized in that said liquid is aqueous.
3 - Équipement d'impression additive selon la revendication 1 caractérisé en ce que ledit liquide est de l'eau. 3 - additive printing equipment according to claim 1 characterized in that said liquid is water.
4 - Équipement d'impression additive selon la revendication 1 caractérisé en ce que ledit liquide est constitué de polymères en solution. 4 - additive printing equipment according to claim 1 characterized in that said liquid consists of polymers in solution.
5 - Équipement d'impression additive selon la revendication 2 ou 3 caractérisé en ce que la longueur d'onde d'émission du laser présente un pic à 2,9 pm ± 0,3 pm. 5 - Additive printing equipment according to claim 2 or 3 characterized in that the laser emission wavelength has a peak at 2.9 pm ± 0.3 pm.
6 - Équipement selon la revendication 1, caractérisé en ce que la focalisation du laser à l'interface substrat / liquide est modifiable par changement de plan focal pour faire varier la taille transversale de la zone d'absorption avec pour effet de moduler la taille des bulles et des jets générés et donc des gouttes déposées sur la cible. 7 - Équipement selon la revendication 7, caractérisé en ce qu'une cartouche à rechargement continu mettant en oeuvre un film de prémouillage exploite l'encre technique de prémouillage comme milieu absorbant pour la longueur d'onde à 2.9pm. 6 - Equipment according to claim 1, characterized in that the focusing of the laser at the substrate / liquid interface can be modified by changing the focal plane to vary the transverse size of the absorption zone with the effect of modulating the size of the bubbles and jets generated and therefore drops deposited on the target. 7 - Equipment according to claim 7, characterized in that a cartridge with continuous reloading implementing a pre-wetting film uses the technical pre-wetting ink as an absorbent medium for the wavelength at 2.9 μm.
8 - Équipement selon la revendication 1, caractérisé en ce que le fluide à imprimer est disposé dans un réservoir. 8 - Equipment according to claim 1, characterized in that the fluid to be printed is disposed in a reservoir.
9 - Équipement selon la revendication 1, caractérisé en ce que l'impression se fasse de bas vers le haut en sens opposé à la force de gravité. 9 - Equipment according to claim 1, characterized in that the impression is made from bottom to top in the opposite direction to the force of gravity.
10 - Équipement selon la revendication 1, caractérisé en ce que l'impression se fasse de haut vers le bas selon la direction de la force de gravité. 10 - Equipment according to claim 1, characterized in that the printing is done from top to bottom in the direction of the force of gravity.
11 - Équipement selon la revendication 1, caractérisé en ce que le système optique permettant la propagation du faisceau laser jusqu'au donneur comporte un scanner 2D ou 3D permettant d'orienter ledit faisceau laser afin de créer un pattern complet de gouttes imprimées par balayage dudit faisceau laser sur ledit donneur dont la surface est d'au moins 0,5 cm2. 11 - Equipment according to claim 1, characterized in that the optical system allowing the propagation of the laser beam to the donor comprises a 2D or 3D scanner making it possible to direct said laser beam in order to create a complete pattern of drops printed by scanning said laser beam on said donor whose surface is at least 0.5 cm2.
12 - Équipement selon la revendication 1, caractérisé en ce que le système optique ne comporte pas de scanner et tire directement sur le donneur, chaque tir permettant de renouveler la zone d'interaction au sein dudit donneur par les mouvements induits dans le film d'encre par la bulle de cavitation et le jet. 12 - Equipment according to claim 1, characterized in that the optical system does not include a scanner and shoots directly on the donor, each shot making it possible to renew the interaction zone within said donor by the movements induced in the film of ink by cavitation bubble and jet.
13 - Équipement selon la revendication 1, caractérisé en ce que le système optique est intégré à un module permettant une régulation de l'hygrométrie. 13 - Equipment according to claim 1, characterized in that the optical system is integrated into a module allowing regulation of the hygrometry.
14 - Équipement selon la revendication 1, caractérisé en ce que le système optique est intégré dans une enceinte fermée dans laquelle un gaz neutre a été introduit afin d'éviter toute d'absorption par vapeur d'eau. 14 - Equipment according to claim 1, characterized in that the optical system is integrated in a closed enclosure into which an inert gas has been introduced in order to avoid any absorption by water vapour.
15 - Méthode d'impression additive par une source laser pulsée produisant un faisceau en direction d'un donneur dépourvu de couche sacrificielle à partir duquel un matériel biologique est imprimé, vers un substrat receveur cible qui recueille le matériel imprimé, ledit donneur étant constitué par une lame recouverte, dans la zone d'interaction du faisceau laser, par un film de liquide destiné à contenir des inhomogénéités transférables, caractérisé en ce que ledit laser émet un faisceau dont la longueur d'onde est comprise entre 2 pm et 3,2 pm et en ce que ladite lame est transparente à la longueur d'onde dudit faisceau laser. 15 - Method of additive printing by a pulsed laser source producing a beam in direction of a donor without a sacrificial layer from which a biological material is printed, towards a target receiver substrate which collects the printed material, said donor being constituted by a plate covered, in the interaction zone of the laser beam, by a film of liquid intended to contain transferable inhomogeneities, characterized in that the said laser emits a beam whose wavelength is between 2 pm and 3.2 pm and in that the said plate is transparent to the wavelength of the said laser beam.
16 - Méthode selon la revendication 13, caractérisée en ce que le fluide à imprimer est supporté par une cartouche à rechargement continu. 16 - Method according to claim 13, characterized in that the fluid to be printed is supported by a cartridge with continuous reloading.
17 - Méthode selon la revendication 14, caractérisée en ce que l'acheminement de l'encre vers la cartouche est robotisé. 17 - Method according to claim 14, characterized in that the delivery of the ink to the cartridge is robotized.
18 - Méthode selon la revendication 16, caractérisée en ce que l'absorption directe du liquide assure une condition d'impression constante pour un jeu de paramètres d'impression donnés avec pour conséquence une très grande stabilité des jets générés ouvrant la voie à une très importante reproductibilité du transfert du fluide vers une cible. 18 - Method according to claim 16, characterized in that the direct absorption of the liquid ensures a constant printing condition for a set of given printing parameters with the consequence of a very high stability of the jets generated opening the way to a very high reproducibility of fluid transfer to a target.
PCT/FR2022/051206 2021-06-25 2022-06-21 Mid-ir laser additive printing equipment WO2022269187A1 (en)

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FR2106803A FR3124523A1 (en) 2021-06-25 2021-06-25 ADDITIVE PRINTING EQUIPMENT BY MID IR LASER

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