METHOD FOR PRODUCING A MONOLITHIC SUPPORT WITH IMMOBILIZED URANYLKATIONS IMMOBILIZED ON IT, AND ASSOCIATED METHOD FOR COLLECTION AND RECOVERY

Description

technical field

[0001] The present invention relates to a method for preparing a monolithic support on which uranyl cations are immobilized, this monolithic support being more particularly synthesized and anchored in situ in the channel(s) of a miniaturized analytical system.

[0002] The present invention also relates to a method for capturing proteins that selectively bind uranium immobilized on such a monolithic support.

[0003] The invention also relates to a method for recovering uranium-selectively binding proteins, this recovery method implementing the previous capture method. Prior art

[0004] Neurotoxic effects induced by low concentrations of uranium are suspected in humans. While the biodistribution of uranium in the human body is well described, the biochemical mechanisms occurring at the cellular and molecular levels that are thought to be responsible for these neurotoxic effects remain to be elucidated. The identification of target molecules, particularly proteins that selectively bind uranium, in a human neuronal cell model should allow us to predict the uranium protein species likely to form and to precisely characterize the biochemical mechanisms associated with uranium neurotoxicity.

[0005] To identify such target molecules, the publication by C. Basset et al. ("Specific capture of uranyl protein targets by metal affinity chromatography", Journal of Chromatography A, 2008, 1185, 233-240), referenced [1]at the end of this description, described the exploitation of the separation mode by immobilized metal ion affinity chromatography (in English, "Immobilized Metal Affinity Chromatography" and abbreviated "IMAC") in order to selectively capture uranium-binding proteins in its uranyl form UO 2 2+< .

[0006] In the remainder of this description, the expression "proteins selectively binding uranium" may be used instead of the expression "proteins selectively binding uranium in its uranyl form UO 2 2+<".

[0007] In the publication [1],Selective capture experiments, based on the IMAC method, were conducted using a support consisting of styrene-divinylbenzene copolymer microbeads functionalized with aminophosphonate groups (Duolite®< C467), through which uranyl ions were immobilized. The presence of the aminophosphonate groups proved satisfactory for immobilizing uranyl ions by complexation and preserving free uranyl bonds that bind with proteins, particularly uranium target proteins, contained in human serum complex samples.

[0008] As reported in the publication by A. Dedieu et al. ("Identification of uranyl binding proteins from human kidney-2 cell extracts by immobilized uranyl affinity chromatography and mass spectrometry", Journal of Chromatography A, 2009, 1216, 5365-5376), referenced [2], the support described in the publication [1]was used for the capture and identification of uranium-selectively binding proteins contained in extracts of human HK-2 kidney cells. These proteins were captured in batch mode and then identified by proteomics.

[0009] publications [1] And [2] They therefore report the first work that has been carried out to immobilize UO 2 2+< ions for the capture of uranium-selectively binding proteins and their identification.

[0010] However, capturing and identifying uranium target proteins in cell extracts remains a challenge, as these extracts are only available in very limited quantities. Consequently, the target proteins are present in these cell extracts only in low abundance. Furthermore, the batch capture method requires a minimum volume of 50 µL of microbeads in a suspension system, necessitating the use of 20 µg to 50 µg of protein samples. One of the major direct consequences is the difficulty in performing experimental replicates, which are essential to validate the repeatability of the capture method and the identification of these uranium target proteins in the cell extracts.

[0011] Therefore, there is an imperative need to reduce the scale of the uranium target protein capture process given the very low availability of biological samples, the low abundance of these target proteins and the limitations of the batch mode.

[0012] The miniaturization of a microbead support presents several technical challenges: not only is filling channels in miniaturized analytical systems with microbeads laborious and difficult to reproduce, but it also requires the use of sintered media to hold the microbeads in place. However, sintered media can lead to the formation of air bubbles and / or solute adsorption during analyses. All of these factors generate significant reproducibility problems for selective capture experiments.

[0013] We also know of document WO 2019 / 008278 A1 which relates to a process for manufacturing chromatography columns comprising monolithic stationary phases and document US 2014 / 0178252 A1 which relates to microfluidic devices containing porous monoliths.

[0014] However, neither of these two documents describes whether these chromatography columns and microfluidic devices are equipped with monolithic stationary phases. on which are immobilized of the cations UO 2 2< .

[0015] The aim of the present invention is, therefore, to overcome the drawbacks of the prior art and to propose a method for preparing a support on which UO 2 2+< cations are immobilized, this support allowing the continuous capture of uranium-selectively binding proteins contained in a biological sample regardless of its volume.

[0016] Another objective of the present invention is to propose a method for capturing uranium-selectively binding proteins contained in a biological sample, this method exhibiting improved repeatability and robustness compared to those of the capture methods described in the publications [1] And [2]. Description of the invention

[0017] The goals stated above, as well as others, are achieved, firstly, by a process of preparing a support on which UO2 2+< cations are immobilized, more particularly by a process of preparing, in the internal volume of at least one channel of a miniaturized analytical system, a monolithic support on which UO2 2+< cations are immobilized.

[0018] According to the invention, this method comprises the following successive steps (a) to (e): (a) activation of the internal surface of the channel(s); (b) introduction of a polymerization solution for the synthesis of a monolithic support into the internal volume of the channel(s), the polymerization solution comprising: a monomer including a phosphate group, at least one crosslinking agent, several solvents, and a radical polymerization initiator; (c) polymerization of the polymerization solution obtained in step (b), thereby obtaining a monolithic support anchored to the walls of the channel(s); (d) rinsing of the monolithic support obtained in step (c); and (e) contacting of the rinsed monolithic support obtained in step (d) with a solution comprising UO2 2+< cations, thereby obtaining the monolithic support on which UO2 2+< cations are immobilized.

[0019] The choice of this monomer containing a phosphate group, in combination with a crosslinking agent, solvents and a radical polymerization initiator, allows for the localized and in situ, in the internal volume of the channel(s) of the miniaturized analytical system, a monolithic support having a mechanically and chemically stable three-dimensional porous polymeric structure on which UO 2 2+< cations are immobilized.

[0020] As previously stated, the process for preparing a monolithic support on which UO2 2+< cations are immobilized according to the invention comprises successive steps (a) to (e).

[0021] The activation step (a) has the effect of functionalizing the internal surface of the channel(s) in such a way as to allow the subsequent anchoring of the monolithic support on its internal wall(s).

[0022] In cases where the channel(s) are made of glass, step (a) of activation can be performed by silanization. Indeed, the silanization reaction allows the silanol groups on the internal surface of the glass channels to be converted into vinyl groups.

[0023] One could, for example, consider using gamma-methacryloxy-propyltrimethoxysilane (γ-MAPS) as a silanizing agent.

[0024] The synthetic polymerization solution of the monolithic support which is introduced into the internal volume of the channel(s) during step (b) of the preparation process according to the invention can be made by a mixture of the monomer comprising a phosphate group, the crosslinking agent(s), solvents and the radical polymerization initiator.

[0025] The polymerization solution generally comprises a single monomer including a phosphate group.

[0026] This monomer can in particular be chosen from among the methacrylate monomers comprising a phosphate group or any other monomer allowing to covalently bind a phosphate group to the surface of a monolithic solid support.

[0027] In a variant of the preparation process according to the invention, the mass proportion of the monomer comprising a phosphate group, relative to the total mass of the polymerization solution, is between 4% by mass and 10% by mass.

[0028] It is specified that the expressions "between ... and ..." and "includes from ... to ..." which are used in this application should be understood as defining not only the values ​​of the interval, but also the values ​​of the bounds of this interval.

[0029] In a particularly advantageous way, the mass proportion of the monomer comprising a phosphate group, relative to the total mass of the polymerization solution, is between 8.7% by mass and 9.7% by mass.

[0030] In a preferred embodiment of the preparation process according to the invention, the polymerization solution implemented in step (b) comprises a methacrylate monomer comprising a phosphate group, this methacrylate monomer being, more preferably still, polyethylene glycol phosphate methacrylate (EGMP).

[0031] The polymerization solution implemented in step (b) of the preparation process according to the invention also includes at least one crosslinking agent.

[0032] In other words, the polymerization solution may consist of only one crosslinking agent but may just as easily consist of a mixture of two, three, or even more crosslinking agents.

[0033] This or these crosslinking agents may in particular be chosen from a mixture of acrylamide (AA) and bisacrylamide (BAA), bisacrylamide (BAA), ethylene dimethacrylate and a derivative of ethylene dimethacrylate.

[0034] In an advantageous variant, the crosslinking agent is composed of an acrylamide / bisacrylamide (AA / BAA) mixture, preferably in a mass ratio of 19 / 1.

[0035] In a variant of the preparation process according to the invention, the mass proportion of the AA / BAA mixture, preferably in a mass ratio of 19 / 1 (AA / BAA), relative to the total mass of the polymerization solution, is between 3% by mass and 8% by mass.

[0036] More particularly advantageously, the mass proportion of the AA / BAA mixture, preferably in a mass ratio of 19 / 1 (AA / BAA), relative to the total mass of the polymerization solution, is between 6.6% by mass and 7.3% by mass.

[0037] The polymerization solution implemented in step (b) of the preparation process according to the invention also includes several solvents.

[0038] These solvents can notably be chosen from dodecanol (DOC), dimethyl sulfoxide (DMSO) and dimethylformamide (DMF).

[0039] In an advantageous variant of the preparation process according to the invention, the polymerization solution implemented in step (b) comprises three solvents, which are preferably dodecanol, dimethylformamide and dimethyl sulfoxide.

[0040] In this advantageous variant, the mass proportion of dodecanol, relative to the total mass of the polymerization solution, is between 28% by mass and 59% by mass and, preferably, between 52.7% by mass and 58.3% by mass; the mass proportion of dimethyl sulfoxide, relative to the total mass of the polymerization solution, is between 22% by mass and 53% by mass and, preferably, between 22.8% by mass and 25.2% by mass; and the mass proportion of dimethylformamide, relative to the total mass of the polymerization solution, is between 7% by mass and 9% by mass.

[0041] The polymerization solution implemented in step (b) of the preparation process according to the invention further comprises a radical polymerization initiator.

[0042] This radical polymerization initiator can be chosen from among the radical polymerization initiators that can be used for the preparation of monolithic supports, namely azobisisobutyronitrile (AIBN), 2,2-diethoxyacetophenone (DEA), the α -dialkoxyacetophenones, 2,2-dimethyl-2-hydroxyacetophenone (DARO), the α -hydroxyacetophenones, benzoin methyl ether (BME), 2-methyl-4'-(methylthio)-2-morpholino-propiophenone (IRG), the α- alkylaminoacetophenones or 2,2-dimethoxy-2-phenylacetophenone (DMPA).

[0043] In a variant of the preparation process according to the invention, the mass proportion of radical polymerization initiator, relative to the total mass of the polymerization solution, is between 0.05% by mass and 0.2% by mass.

[0044] In a particularly advantageous manner, the mass proportion of radical polymerization initiator, relative to the total mass of the polymerization solution, is between 0.14% by mass and 0.16% by mass.

[0045] In a preferred variant of the preparation process according to the invention, the polymerization solution uses azobisisobutyronitrile (AIBN) as a radical polymerization initiator.

[0046] It is worth noting that a synthetic polymerization solution for a monolithic support comprising polyethylene glycol phosphate methacrylate, acrylamide, bisacrylamide, dodecanol, dimethyl sulfoxide, dimethylformamide, and azobisisobutyronitrile was very recently described in the publication by M. Araya-Farias et al. ("A lab-on-chip for monolith-based preconcentration and electrophoresis separation of phosphopeptides", Analyst, 2017, 142, 485-494), referenced [3],to pre-concentrate phosphopeptides, which are potential biomarkers of Alzheimer's disease. To this end, Zr 4+ cations were immobilized on the surface of a synthesized monolithic phosphate support in situ and anchored in the microchannel of a commercial cross-shaped chip. The miniaturized analytical system incorporating such a monolithic support has demonstrated excellent performance in terms of selectivity and phosphopeptide enrichment factor.

[0047] However, unexpectedly and surprisingly, the Inventors observed that the support described in the publication [3] is perfectly suited for an application other than that described in this publication, no longer for the immobilization of Zr 4+< cations and the capture of certain phosphopeptides, but for the immobilization of UO 2 2+< cations, the capture of proteins selectively binding uranium and their recovery.

[0048] In a particularly preferred version, the polymerization solution that is introduced into the internal volume of the channel(s) during step (b) of the preparation process according to the invention comprises, relative to the total mass of the polymerization solution: from 4% to 10% by mass and, advantageously, from 8.7% to 9.7% by mass of polyethylene glycol phosphate methacrylate, from 3% to 8% by mass and, advantageously, from 6.6% to 7.3% by mass of an acrylamide / bisacrylamide mixture, preferably in a mass ratio of 19:1, from 28% to 59% by mass and, advantageously, from 52.7% to 58.3% by mass of dodecanol, from 22% to 53% by mass and, advantageously, from 22.8% to 25.2% by mass of dimethyl sulfoxide, from 7% to 9% by mass of dimethylformamide, and from 0.05% to 0.2% by mass and, advantageously, from 0.14% by mass to 0.16% by mass of azobisisobutyronitrile.

[0049] After step (b) of introducing the polymerization solution into the internal volume of the channel(s) of the miniaturized analytical system, step (c) of polymerization of this polymerization solution is carried out to obtain a monolithic support.

[0050] This step (c) of polymerization of the preparation process according to the invention is a radical chain polymerization, initiated by the radical polymerization initiator.

[0051] This polymerization step (c) is advantageously a photopolymerization, that is, a polymerization carried out by irradiation of the polymerization solution with ultraviolet (UV) rays. This irradiation can, in particular, be carried out on a localized area of ​​the canal.

[0052] In an advantageous variant of the preparation process according to the invention, the wavelength of the ultraviolet rays is between 320 nm and 380 nm, preferably between 330 nm and 370 nm and, more preferably still, is 347 nm, which is the maximum absorption of AIBN.

[0053] In an advantageous variant of the preparation process according to the invention, the irradiation time using ultraviolet rays is between 5 min and 60 min, advantageously between 15 min and 45 min and, preferably, is 25 min.

[0054] At the end of step (c), a monolithic support is obtained which has been synthesized in situ and which is anchored to the walls of the channel(s) of the miniaturized analytical system.

[0055] The preparation process according to the invention comprises, after step (c) of polymerization, a step (d) of rinsing the monolithic support. This rinsing step (d) removes excess reagents from the polymerization solution that have not reacted.

[0056] This rinsing step (d) can in particular be carried out successively with alcohol and then with water, the alcohol being advantageously methanol.

[0057] The preparation process then includes a step (e) of bringing the rinsed monolithic support, as obtained at the end of step (d), into contact with a solution comprising the UO2 2+< cations. By doing so, a monolithic support is obtained within the internal volume of the channel(s) on which the UO2 2+< cations are immobilized.

[0058] In an advantageous variant of the preparation process according to the invention, the solution comprising the UO2 2+< ions is prepared in an aqueous solution of ammonium acetate.

[0059] Monitoring the fixation of UO 2 2+< ions on the monolithic support can be advantageously carried out by coupling with an inductively coupled plasma mass spectrometer (ICP-MS).

[0060] In an advantageous variant of the preparation process according to the invention, the step (e) of bringing the monolithic support into contact with the solution comprising the UO2 2+< cations is carried out by circulating this solution comprising the UO2 2+< cations through the monolithic support.

[0061] The preparation process according to the invention therefore makes it possible to synthesize in situ a monolithic support on which the uranyl cations UO 2 2+< are immobilized, in miniaturized analytical systems whose size is far smaller, at a minimumby a factor of 1000, to those of analytical systems formed by microbeads such as those described in the publications [1] And [2].

[0062] In particular, the process according to the invention makes it possible to prepare supports on which UO 2 2+< cations are immobilized in channels whose internal diameter can advantageously be less than or equal to 300 µm and, preferably, can be between 50 µm and 90 µm.

[0063] The scale reduction achievable with the preparation process according to the invention also reduces the consumption of solvents and reagents, as well as the quantity of reaction by-products and their treatment, which offers a clear industrial advantage. This scale reduction is particularly beneficial for applications in the nuclear field, as it reduces the constraints associated with handling radioactive samples and also limits the volume of waste and the costs associated with its specific management.

[0064] The present invention relates, secondly, to a method for capturing uranium-selectively binding proteins, these proteins being contained in a biological sample.

[0065] According to the invention, this capture method comprises the following steps (i) and (ii): (i) the preparation, in the internal volume of at least one channel of a miniaturized analytical system, of a monolithic support on which UO2 2+< cations are immobilized, by implementing the preparation process as defined above, and (ii) at least the circulation of a solution containing the biological sample through the monolithic support on which UO2 2+< cations are immobilized, obtained at the end of step (i), thereby achieving the capture of uranium-selectively binding proteins on the monolithic support.

[0066] In the capture process according to the invention, during step (i), preparation is prepared in situa monolithic support by the preparation process as defined above, it being specified that the advantageous characteristics of this preparation process, in particular those relating to the implementation methods of steps (a) to (e) and those relating to the channel(s) of the miniaturized analytical system, may be taken alone or in combination.

[0067] As previously described, this monolithic support is anchored to the internal walls of the channel(s) and comprises immobilized uranyl ions.

[0068] During the circulation step (ii), there is selective capture of the protein(s) which, among those contained in the biological sample, have an affinity for the UO2 2+< ions which are immobilized on the monolithic support.

[0069] The selective capture of these uranium-selectively binding proteins is therefore carried out according to the IMAC method, using a biological sample whose quantity can be drastically reduced compared to that required for capture using the publication support. [1] And [2].

[0070] Thirdly, the present invention relates to a method for recovering uranium-selectively binding proteins, these proteins being contained in a biological sample.

[0071] According to the invention, this method comprises the following steps (1) to (3): (1) the capture of uranium-selectively binding proteins by implementing the capture process as defined above, (2) the removal of unbound proteins by rinsing the monolithic support by circulating a protein-free solution, and (3) at least one elution step, by circulating an eluent solution through the monolithic support obtained at the end of step (2), thereby recovering the uranium-selectively binding proteins in the eluent solution.

[0072] In the recovery process according to the invention, during step (1), the selectively uranium-binding proteins are captured by implementing the capture process as defined above, it being specified that the advantageous characteristics of this capture process can be taken alone or in combination.

[0073] These uranium-selectively binding proteins, which were captured in step (1), are thus recovered from the monolithic support by implementing step (3), which includes at least one elution step using an eluent solution.

[0074] The recovery process according to the invention is particularly simple to implement and allows, by elution, the selective recovery of uranium-binding proteins that have been previously captured on the monolithic support on which UO2 2+ cations are immobilized.

[0075] It is specified that the biological sample involved in the process of capturing uranium-selectively binding proteins and, consequently, in the process of recovering uranium-selectively binding proteins, may in particular consist of protein solutions, cell extracts or biological fluids, and may in particular be derived from a human neuronal cell line.

[0076] Other features and advantages of the invention will become apparent from the following supplementary description, which relates to an example of the preparation of an 8 mm monolithic support on which uranyl cations are immobilized, and to an example illustrating the performance of such a monolithic support for capturing and then recovering uranium-selectively binding proteins contained in a biological sample.

[0077] Of course, these examples are given only as an illustration of the object of the invention and do not constitute, in any way, a limitation of this object. Detailed description of specific implementation methods

[0078] A synthetic polymerization solution for a monolithic support was prepared from the following compounds, in the mass proportions specified below: 2.22 mg (0.15% by mass) of azobisisobutyronitrile (AIBN), 100.05 mg (6.7% by mass) of an acrylamide (AA) and bisacrylamide (BAA) mixture (ratio 19 / 1), 130.60 mg (8.7% by mass) of polyethylene glycol phosphate methacrylate (EGMP), 346.03 mg (23.1% by mass) of dimethyl sulfoxide (DMSO), 799.93 mg (53.4% ​​by mass) of dodecanol (DOC), and 118.80 mg (7.9% by mass) of dimethylformamide (DMF).

[0079] In a bottle, AIBN, AA-BAA, EGMP, DMSO, DOC, then DMF were successively introduced, and then all of these compounds were mixed to obtain the polymerization solution for the synthesis of the monolithic support.

[0080] After degassing, the polymerization solution obtained was introduced into one or more of the four straight channels, parallel to each other and having a width of 50 µm, a depth of 50 µm and a length of 58.5 mm, of a glass analytical microsystem marketed by the company ChipShop.

[0081] It is specified that, before the introduction of this polymerization solution, the internal surface of the channel(s) was functionalized by a mixture comprising gamma-methacryloxy-propyltrimethoxysilane (γ-MAPS) solubilized in acetone (50 / 50, v / v).

[0082] After introducing the polymerization solution into one or more of the four channels, the microsystem was placed in an oven under UV radiation with a maximum wavelength of 365 nm, at 14.5 cm from the source for 25 min so as to have a power of 3.1 mW.cm -2< in order to obtain the polymerization of the polymerization solution.

[0083] The monolithic support thus formed in the channel(s) was rinsed using a mixture comprising methanol and water.

[0084] A solution comprising 500 parts per billion (ppb) of uranyl ions in ammonium acetate buffer at a molar concentration of ammonium acetate between 25 mmol / L and 50 mmol / L and a pH between 4 and 5, is prepared.

[0085] This solution is then circulated through the synthesized monolithic support. in situand anchored to the walls of the channel(s) of the microsystem, for 60 min, at a flow rate of 0.24 mL / h (4 µL / min) so as to immobilize the uranyl ions on the surface of the monolithic support.

[0086] The efficiency of uranyl ion immobilization is determined using an inductively coupled plasma mass spectrometer (ICP-MS).

[0087] This step can be performed offline by fraction retrieval. However, it can advantageously be performed online, i.e., continuously, by coupling the microsystem to the mass spectrometer via a micro-nebulizer. In the present example, 50 ng of uranium were fixed onto the 8 mm monolithic support.

[0088] In the case where the mass spectrometer is a triple quadrupole ICP, the signal of phosphorus and / or sulfur contained in proteins can also be followed in addition to that of uranium, when monitoring protein capture.

[0089] The ability to track both uranyl ions and their target molecules is a significant advantage, allowing us to determine both the immobilization efficiency of these ions on the monolithic support and its capacity to capture target molecules through interaction with the immobilized ions. For example, the amount of apotransferrin protein captured by this monolithic support, on which the uranyl ions are immobilized, and recovered, is 0.75 µg.

[0090] Table 1 below shows the amount of uranium immobilized on the monolithic support prepared by the process according to the invention, as well as the amount of apotransferrin recovered following its capture by this same support, compared to the amounts obtained with the reference support formed by the microbeads as described in publication [1]. Table 1 Microbead support (reference) Monolithic support (according to the invention) Media volume 50 µL of a solution containing 1.1 g of microbeads / mL 16 nL (8 mm x 50 µm x 50 µm) Quantity of immobilized uranium 5.95 mg 50 ng Quantity of apotransferrin eluted 40 µg 0,75 µg References

[0091] [1] C. Basset et al., Journal of Chromatography A, 2008, 1185, pages 233-240 [2] A. Dedieu et al., Journal of Chromatography A, 2009, 1216, pages 5365-5376 [3] M. Araya-Farias et al., Analyst, 2017, 142, pages 485-494

Claims

1. A method for preparing, in the internal volume of at least one channel of a miniaturised analytical system, a monolithic support on which UO22+ cations are immobilised, which method comprises the following successive steps (a) to (e) of: (a) activating the inner surface of the channel(s); (b) introducing a polymerisation solution for synthesising a monolithic support into the internal volume of the channel(s), the polymerisation solution comprising: - a monomer comprising a phosphate group, - at least one crosslinking agent - several solvents, and - a radical polymerisation initiator; (c) polymerising the polymerisation solution obtained in step (b), whereby a monolithic support anchored onto the walls of the channel(s) is obtained; (d) rinsing the monolithic support obtained in step (c); and (e) contacting the rinsed monolithic support obtained in step (d) with a solution comprising UO22+ cations, whereby a monolithic support on which the UO22+ cations are immobilized is obtained.

2. The method according to claim 1, wherein the monomer comprising a phosphate group is a methacrylate monomer, preferentially polyethylene glycol methacrylate phosphate.

3. The method according to claim 1 or 2, wherein the crosslinking agent is composed of a mixture of acrylamide / bisacrylamide.

4. The method according to any of claims 1 to 3, wherein the solvents are selected from dodecanol, dimethylformamide and dimethylsulphoxide.

5. The method according to any of claims 1 to 4, wherein the radical polymerisation initiator is azobisisobutyronitrile (AIBN).

6. The method according to claim 5, wherein the polymerisation solution comprises, based on the total mass of the polymerisation solution: - from 0.05 mass % to 0.2 mass % and, advantageously, from 0.14 mass % to 0.16 mass % of azobisisobutyronitrile, - from 3 mass % to 8 mass % and, advantageously, from 6.6 mass % to 7.3 mass % of acrylamide and bisacrylamide, - from 4 mass % to 10 mass % and, advantageously, from 8.7 mass % to 9.7 mass % of polyethylene glycol methacrylate phosphate, - from 22 mass % to 53 mass % and, advantageously, from 22.8 mass % to 25.2 mass % of dimethylsulphoxide, - from 28 mass % to 59 mass % and, advantageously, from 52.7 mass % to 58.3 mass % of dodecanol, and - from 7 mass % to 9 mass % dimethylformamide.

7. The method according to any of claims 1 to 6, wherein the polymerisation step (c) is carried out by irradiation by means of ultraviolet rays, the wavelength of these ultraviolet rays being advantageously between 320 nm and 380 nm, preferentially between 330 nm and 370 nm and, even more preferentially, being 347 nm.

8. The method according to claim 7, wherein, in step (c), the duration of irradiation by means of the ultraviolet rays is between 5 min and 60 min, advantageously between 15 min and 45 min and, more preferentially, is 25 min.

9. The method according to any of claims 1 to 8, wherein the rinsing step (d) is carried out successively with an alcohol and then with water, the alcohol being advantageously methanol.

10. The method according to any of claims 1 to 9, wherein, the channel(s) being made of glass, the activation step (a) is carried out by silanisation, advantageously by means of gamma-methacryloxypropyltrimethoxysilane (γ-MAPS) as silanising agent.

11. The method according to any of claims 1 to 10, wherein the internal diameter of the channel(s) is advantageously less than or equal to 300 µm and, preferentially, is between 50 µm and 90 µm.

12. The method according to any of claims 1 to 11, wherein the solution comprising the UO22+ cations is prepared in an aqueous solution of ammonium acetate.

13. The method according to any of claims 1 to 12, wherein step (e) of contacting the monolithic support with the solution comprising the UO22+ cations is carried out by circulating this solution comprising the UO22+ cations through the monolithic support.

14. A method for capturing proteins that selectively bind uranium, these proteins being contained in a biological sample, this method comprising the following steps (i) and (ii) of: (i) preparing, in the internal volume of at least one channel of a miniaturised analytical system, a monolithic support on which UO22+ cations are immobilised, by implementing the preparation method according to any of claims 1 to 13, and (ii) at least circulating a solution containing the biological sample through the monolithic support on which UO22+ cations are immobilised, obtained at the end of step (i), whereby the capture of the proteins that selectively bind uranium on the monolithic support is achieved.

15. A method for recovering proteins that selectively bind uranium, these proteins being contained in a biological sample, this method comprising the following steps (1) to (3) of: (1) capturing the proteins that selectively bind uranium by implementing the capture method of claim 14, (2) removing the unbound proteins by rinsing the monolithic support by circulating a solution containing no proteins, and (3) at least one elution step, by circulating an eluting solution through the monolithic support obtained at the end of step (2), whereby the proteins that selectively bind uranium are recovered in the eluting solution.