Composition for hematology analyses

Abstract

The invention relates to a composition for hematology analyses and to the method for producing same. The composition comprises an anticoagulant, in an amount of between 0.1% and 8% by weight, preferably between 2% and 6%; a salt in an amount of between 30% and 90% by weight, preferably between 50% and 80%; a pH buffer in an amount of between 0.1% and 50% by weight, preferably greater than or equal to 10%; and a surfactant in an amount of between 0.01% and 2% by weight, preferably between 0.05% and 1.5%, in solid form. The invention also relates to a solution obtained from the solid composition.

Description

Description Title of the invention: Composition for hematology analyses.

[0001] The invention relates to a composition for hematology analyses and its manufacturing process. The invention also relates to a solution prepared from the composition of the invention and intended to be used in hematology analysis devices.

[0002] Hematology devices use specific reagents to analyze biological samples. The biological sample is preferably a blood sample, but it can also be, for example, cerebrospinal fluid, bone marrow fluid, or urine. The sample can contain particles of any nature (cells, proteins, biomarkers, etc.) that will need to be identified and counted. Prior to analysis, the biological sample can be diluted and treated with one or more reagents. The sample is then transported, for example, to an automated flow measurement system that uses electrical and optical measurements to count and differentiate the cells present in the sample to be analyzed.

[0003] The sample is then sent to an analyzer, or, in the case of specific blood analyses, to a hematology analyzer. The analyzer begins by distributing volumes of sample into containers that already contain, or receive shortly after or simultaneously, a reagent. This reagent is used, for example, to detect one or more elements of interest present in the sample, or to characterize these elements (determination of volumes, hemoglobin dosage or other). It may in particular be a diluting reagent, a lysis reagent, a staining reagent, a sheathing reagent or a cleaning reagent.

[0004] The reagent is therefore an essential element in the operation of biological analysis devices, and in particular hematology analyzers. It must be present at all times and, in particular, the diluent in considerable quantities.

[0005] One of the challenges is the need for frequent replenishment of this reagent. For example, when the reagent is diluent manufactured and / or sold at working concentration (ready-to-use), a laboratory or hospital may need to schedule a high-volume replenishment in a single day. This requires dedicated storage space, and sometimes even dedicated replenishment staff. Staff also face handling challenges due to the heavy loads that need to be handled and moved. However, most laboratories are limited in space, and it is usually difficult to keep large quantities of diluent on hand. Furthermore, there is the problem of having to store used containers until they can be removed from the laboratory.

[0006] An attempt has been made to develop preparations of concentrated diluents. The document WO 19222620 discloses such a product.

[0007] However, the preparation of these concentrated diluents has encountered maximum concentrations reaching only about 25 times the working concentration. This is mainly due to problems of non-homogeneous mixing.

[0008] Another problem with known concentrated diluents is the presence of harmful components, such as formaldehyde, imidazole, glutaraldehyde, or sodium azide, used as a preservative. This poses a health risk to personnel as well as environmental problems. State-of-the-art products are therefore classified as seriously harmful to health and are typically marked with hazard pictograms. This leads to transport and / or storage problems, particularly due to the need for specific packaging.

[0009] The present invention improves the situation.

[0010] Thus, the invention relates to a composition for hematology analyses comprising: - at least one anticoagulant, in an amount of between 0.1% and 8% by weight, preferably between 2% and 6%; - at least one salt in an amount of between 30% and 90% by weight, preferably between 50% and 80%; - at least one pH buffer in an amount of between 0.1% and 50% by weight, preferably greater than or equal to 10%; and - at least one surfactant in an amount of between 0.01% and 2% by weight, preferably between 0.05% and 1.5%,

[0011] which is in solid form.

[0012] The invention thus provides a reagent in solid form for the field of hematology (in particular in the form of a pellet, powder or granule). In particular, in a preferred embodiment, the invention provides a solid diluent type reagent which is a pioneer in the field of hematology. It can be used directly by simple dissolution in a solvent, such as osmosis water for example, and thus avoids the problems of the state of the art.

[0013] In one embodiment, the anticoagulant is a chelating agent, preferably EDTA, or one selected from K2-EDTA, K3-EDTA, Na2EDTA, T acid-citrate-dextrose (ACD), lithium heparin (LH), sodium heparin (NH), and sodium citrate.

[0014] In a preferred embodiment, the amount by weight of the anticoagulant (which is preferably a chelating agent) is between about 3% and about 4%; the amount by weight of the salt is between about 50% and about 70%; the amount by weight of the pH buffer is between about 30% and about 45%; and the amount by weight of the surfactant is between about 0.05% and about 0.80%. In this embodiment, the amount of the salts can be adapted depending on the buffer pH, and vice versa. The less salt you have, the more buffer you will need to add and vice versa.

[0015] In another preferred embodiment, the salt is composed of a first salt, preferably NaCl, in an amount of between 30% and 90% by weight, preferably between 50% and 80%, and a second salt, preferably KCl, in an amount of between 0.1% and 90% by weight of said composition, preferably between 1% and 5%. The use of two salts has an effect on the pH. This allows in particular to use less pH buffer, which optimizes costs.

[0016] In another preferred embodiment, the pH buffer is chosen so as to obtain a pH between 6.5 and 8.5, preferably between 7.5 and 8.5. The pH buffer may be chosen from the group consisting of K2HPO4,3H2O, NaH2PO4,2H2O, Na2HPO4,12H2O and KH2PO4 OR a combination thereof. Preferably, the Tris buffer NH2C(CH2OH)3, NH2C(CH2OH)3HC1 is used.

[0017] Thus, the pH buffer may be selected from the group consisting of K2HPO4,3H2O, NaH2PO4,2H2O, Na2HPO4,12H2O, KH2PO4 and Tris buffer NH2C(CH2OH)3, NH2C(CH2OH)3HC1 OR a combination thereof

[0018] In another preferred embodiment, the surfactant is selected from the group consisting of Tween 20, Tween 80, Tergitol 15-S-9, Tergitol 15-S-15, Brij L23, Brij 35, SB3-12 and SB3-14. Combinations of surfactants are particularly envisaged to improve stability or to increase the penetration of an active ingredient or a coloring agent into cells. The choice of surfactant depends on the final application. Each surfactant has a specific interaction with the cell membrane, for example to stiffen the membrane or to make the cell more spherical.

[0019] In another preferred embodiment, the composition has the following formulation:

[0020] [Tables 1]

[0021] In another preferred embodiment, the composition has the following formulation:

[0022] [Tables 2]

[0023] Preferably, the composition is in a form selected from a tablet, a powder or a granule.

[0024] In one embodiment, the composition of the invention consists of: - an anticoagulant (which is preferably a chelating agent) in an amount of between 0.1% and 8% by weight, preferably between 2% and 6%; - a salt in an amount of between 30% and 90% by weight, preferably between 50% and 80%; - a pH buffer in an amount of between 0.1% and 50% by weight, preferably greater than or equal to 10%; - a surfactant in an amount of between 0.01% and 2% by weight, preferably between 0.05% and 1.5%; - and optionally traces of solvent.

[0025] The invention also relates to a solution for hematology analyses obtained by dissolving the composition described above comprising: - an anticoagulant (which is preferably a chelating agent, preferably EDTA), in an amount of between 0.01% and 0.2% by weight, preferably between 0.05% and 0.15%; - a first salt, preferably NaCl, in an amount of between 1.15% and 3.5% by weight, preferably between 0.8% and 0.9%; - a second salt, preferably KC1, in an amount of between 0.01% and 0.9% by weight, preferably between 0.02% and 0.04%; - a pH buffer in an amount of between 0.01% and 0.9% by weight, preferably 0.5%; - a surfactant in an amount of between 0.001% and 0.02% by weight, preferably between 0.01% and 0.015%; and - a solvent, preferably osmosis water, in a quantity remaining to obtain 100% by weight of said solution.

[0026] The solvent may contain lysing agents, staining agents, enzymes or other elements commonly used in hematology analyses.

[0027] Furthermore, the present invention relates to a method for manufacturing a composition for hematology analyses, comprising the following steps:

[0028] a. the provision of the following components: - an anticoagulant (which is preferably a chelating agent), in an amount of between 0.1% and 8% by weight of said composition, the anticoagulant being in the form of a fine powder having a grain size of between approximately 0.8 mm and approximately 2.2 mm; - a salt, in an amount of between 30% and 90% by weight of said composition, the salt being in the form of a fine powder having a grain size of between approximately 0.8 mm and approximately 2.2 mm; - a pH buffer, in an amount of between 0.1% and 50% by weight of said composition, the pH buffer being in the form of a fine powder having a grain size of between approximately 0.8 mm and approximately 2.2 mm; - a liquid surfactant in an amount of between 0.01% and 2% by weight of said composition, mixed with a solvent in a surfactant:solvent proportion of between 1:10 and 1:40;

[0029] b. mixing the anticoagulant with the pH buffer to obtain a fine powder mixture;

[0030] c. adding the liquid surfactant mixed with the solvent to said fine powder mixture so as to obtain a pasty premix;

[0031] d. adding salt to said pasty premix followed by homogenization so as to obtain a paste; and

[0032] e. resting said paste to obtain said composition in a solid form.

[0033] In a preferred embodiment, the salt is composed of: - a first salt, preferably NaCl, in an amount of between 30% and 90% by weight, preferably between 50% and 80%, and - a second salt, preferably KC1, in an amount of between 0.1% and 90% by weight of said composition, preferably between 1% and 5%.

[0034] In another preferred embodiment, the solvent is chosen from osmosis water, distilled water and ethanol.

[0035] In another preferred embodiment, the method further comprises a step of homogenizing the grain size of said salt to obtain said grain size of between 0.8 mm and 2.2 mm.

[0036] In a preferred embodiment of the invention, the homogenization of step d. comprises the coating of the salt grains with at least part of the surfactant.

[0037] Other features, details and benefits will become apparent upon reading the des- detailed description below, and to the analysis of the attached drawings on which:

[0038] [Fig- 1] shows a microscopic photograph of a grain of the composition of the invention in granular form;

[0039] [Fig.2] shows a microscopic photograph of several grains of the composition of the invention in granular form;

[0040] [Fig.3] shows histograms of red blood cells (RBCs) obtained using a state-of-the-art reagent and a solution prepared from the composition of the invention; and

[0041] [Fig.4] shows comparative blood counts between an analysis with a state-of-the-art reagent and with a solution prepared from the composition of the invention.

[0042] The figures, tables and description below contain, for the most part, elements of a certain nature. The figures and tables are an integral part of the description, and may therefore not only serve to better understand the present invention, but also contribute to its definition, where appropriate.

[0043] Optimal sample flow management has become an important element in laboratory organization. Two criteria are particularly important to consider: sample processing time (Turn Around Time) and the workload that each action imposes on staff (Full Time Equivalent).

[0044] Reagent supply influences both of these parameters, as well as carbon impact, storage costs, and purchasing procedures. Reagents are often contained in large containers called cubitainers, which are directly connected to the analyzers.

[0045] Containers are heavy and bulky and must be changed regularly by staff when empty. Since diluted reagent containers are composed of approximately 80% water, reagent preparation units have been developed to allow for less frequent changing of these containers by filling them with diluted reagent produced from concentrated reagent and water available on site in the laboratory.

[0046] The reagent must have precise characteristics so as not to distort the measurements of the devices in which it is used. For this reason, it is crucial that the preparation devices produce a reagent that reliably presents the desired concentration. However, the water or diluent used and the concentrated reagent often have different temperatures, temperatures that can also vary greatly depending on the laboratory and working methods. These differences are sources of errors linked to the expansion of the volumes of the solutions as a function of temperature in the dosing systems. Indeed, for the same quantity of the same liquid, the volume of the liquid can be different depending on the temperature. To overcome this problem and other sources of error in the dosage, existing devices are based on a panel of physical measurements in order to validate the dilution carried out in the preparation device such as conductivity and pH.

[0047] In very general terms, the present invention relates to a composition in solid form, containing all the ingredients and / or chemical products necessary to constitute a reagent intended to be used in a hematology analyzer. The solid composition may be in particular in the form of a pellet, powder or granule. Mixing this solid composition with osmosis water makes it possible to reconstitute the reagent in liquid form. The reagent is thus ready for use in a hematology analyzer. In other words, the reagent is reconstituted by adding osmosis water to the solid composition of the invention. Mixing the composition with osmosis water may preferably be followed by filtration. Then, the reagent can be directly used in a hematology analyzer, typically as a diluent, lysis, sheathing or rinsing reagent.

[0048] The invention thus provides a solid composition for hematological analysis, comprising only the substances necessary for the preparation and analysis of a biological sample such as a blood sample, plasma, platelet-rich plasma (PRP) preparation, cerebrospinal fluid, pleural fluid, liquid taken during a bone marrow puncture or even a synovial fluid. The solid composition of the invention comprises a chelator, a surfactant (also called a detergent) and a pH buffer.

[0049] More specifically, the composition of the invention comprises: - an anticoagulant, which is preferably a chelating agent (preferably EDTA), in an amount of between 0.1% and 8% by weight, preferably between 2% and 6%; - a salt in an amount of between 30% and 90% by weight, preferably between 50% and 80%; - a pH buffer in an amount of between 0.1% and 50% by weight, preferably greater than or equal to 10%; and - a surfactant in an amount of between 0.01% and 2% by weight, preferably between 0.05% and 1.5%.

[0050] The composition of the invention is dissolvable in an inert liquid, such as reverse osmosis water, distilled water or ethanol. In particular, this dissolution is facilitated by the surfactant, also called detergent, which prevents crystallization. Non-ionic surfactants are mainly used. But other families of surfactants are possible such as non-ionic, cationic, anionic and zwitterionic surfactants. The surfactant used in the invention allows almost 100% of the composition to be dissolved. Indeed, when the composition in solid form of the invention is brought into contact with an inert liquid, it dissolves in the liquid and passes into the mass of this liquid to give a homogeneous whole.

[0051] Thus, the present invention also relates to a solution for hematology analyses obtained by dissolving the solid composition described above. The solution of the invention comprises: - an anticoagulant, which is preferably a chelating agent, preferably EDTA, in an amount of between 0.01% and 0.2% by weight, preferably between 0.05% and 0.15%; - a first salt, preferably NaCl, in an amount of between 1.15% and 3.5% by weight, preferably between 0.8% and 0.9%; - a second salt, preferably KC1, in an amount of between 0.01% and 0.9% by weight, preferably between 0.02% and 0.04%; - a pH buffer in an amount of between 0.01% and 0.9% by weight, preferably 0.5%; - a surfactant in an amount of between 0.001% and 0.02% by weight, preferably between 0.01% and 0.015%; and - a solvent, preferably osmosis water, in a quantity remaining to obtain 100% by weight of said solution.

[0052] The Applicant tested various surfactants which showed satisfactory results. These surfactants are reported in Table 3 below.

[0053] [Tables 3]

[0054] Once the solid composition is dissolved in osmosis water, it can be used in the form of a solution for hematological analyses directly in a hematological analysis device. Indeed, the composition has all the physicochemical characteristics so as not to distort the hematological analyses and / or measurements. Thus, the solution of the invention (obtained from the solid composition of the invention) has the required functions, in particular in terms of osmolarity, pH, surfactant and conductivity. Other functions such as lysis or staining are possible. In particular, the solution meets the requirements relating to hypertonicity and hypotonicity concentration limits. For example, the salt concentration of the composition is not higher than 1.2% (12 g / L, 200 mM) - Hypertonicity 400 mOsm / L, and not lower than 0.6% (6 g / L, 100 mM) - Low hypotonicity 100 mOsm / L. Furthermore, the composition allows cell analysis, counting and differentiation by resistivity and / or optical measurements.

[0055] The Applicant tested the salt concentration limits in the composition, defined by the resistivity target 60.4 Qm ± 2 (acceptable tolerance identified)

[0056] [Tables 4]

[0057] In practice, we aim broadly for an operating range between: maximum 200 mM, 400 mOsm / L (hypertonicity) and minimum 100 mM, 100 mOsm / L (low hypotonicity).

[0058] More generally, the salt of the present invention encompasses so-called molten salts, i.e. dissociated salts consisting solely of ions. This particularly encompasses electrolytes. The term "salt" of the invention can thus be read as "salt" or "at least one electrolyte".

[0059] A major advantage of the composition of the invention is that it can be produced without harmful compounds. The problems of prior art reagents related to safety, the environment, transport and / or storage therefore do not arise.

[0060] Another advantage of the composition of the invention is the possibility of a preservative-free embodiment. The absence of preservative in the product helps avoid chemical risks. This makes it possible to provide a composition free from any mention of hazardous classification and / or labeling by hazard pictogram.

[0061] Another advantage is related to the solid form of the composition. Indeed, the solid form of the invention offers increased storage capacity. As such, the composition offers a double skill: on the one hand it is stable over time, and, on the other hand, it allows a concentration going up to well beyond 60 times the working concentration. In a particular embodiment of the invention, nearly 90 times (approximately 87.7 for the example described) the working concentration is reached, and 126 times in the low hypotonicity limit to preserve the integrity of the sample (NaCl 0.6%, 100 mM, 100 mOsm / L). This results in easier storage in a reduced space. It also results in ease of use.

[0062] Overall, the composition and / or solution of the invention can be used in particle analysis devices. The analysis devices are of the biological analyzer type, for example for the preparation, counting and / or identification of particles. A particle is understood to mean any product that can be analyzed and requires, in the context of this analysis, the use of at least one reagent. Particles include, in particular: blood cells, crystals or other molecules.

[0063] The invention also relates to the method of manufacturing the solid composition.

[0064] There are various methods in the state of the art for manufacturing a composition in solid form. In particular, techniques commonly used in the field of biotechnology and the medical field are known. For example, techniques such as extrusion in a melted medium, electrospinning, atomization (spray drying), lyophilization, freezing-thawing, and co-precipitation can be mentioned. However, all these manufacturing techniques require the suspension of the various chemical compounds in solution and generally the addition of an adjuvant and / or a binder.

[0065] The invention provides another type of manufacturing process. Thus, the composition of the invention can be produced as follows:

[0066] a. the provision of the following components: - an anticoagulant, which is preferably a chelating agent, in an amount of between 0.1% and 8% by weight of said composition, the anticoagulant being in the form of a fine powder having a grain size of between approximately 0.8 mm and approximately 2.2 mm; - a salt, in an amount of between 30% and 90% by weight of said composition, the salt being in the form of a fine powder having a grain size of between approximately 0.8 mm and approximately 2.2 mm; - a pH buffer, in an amount of between 0.1% and 50% by weight of said composition, the pH buffer being in the form of a fine powder having a grain size of between approximately 0.8 mm and approximately 2.2 mm; - a liquid surfactant in an amount of between 0.01% and 2% by weight of said composition, mixed with a solvent in a surfactant:solvent proportion of between 1:10 and 1:40;

[0067] b. mixing the anticoagulant with the pH buffer to obtain a fine powder mixture;

[0068] c. adding the liquid surfactant mixed with the solvent to said fine powder mixture so as to obtain a pasty premix;

[0069] d. adding salt to said pasty premix followed by homogenization so as to obtain a paste; and

[0070] e. resting said paste to obtain said composition in a solid form.

[0071] Unlike the prior art, the method of the invention does not require any addition of binders or other additives. Furthermore, the method of the invention avoids freeze-drying. It also avoids a drying step. And, the components used do not contain any preservatives or other toxic elements.

[0072] The surfactant can advantageously coat the powder grains, particularly the salt and / or electrolyte used.

[0073] The hygroscopic properties of the salt contribute significantly to the shaping of the composition of the invention. In particular during the manufacturing process, at step e. of resting the dough, the ionic charges of the salt attract the molecules of the solvent. In this way, the composition of the invention is formulated in a solid form.

[0074] It is also possible to provide a step of grinding said salt until a powder is obtained whose grain size is between approximately 0.8 mm and 2.2 mm.

[0075] A dough shaping step can be provided, for example for shaping pellets or granules.

[0076] Salt can be advantageously composed: - a first salt, preferably NaCl, in an amount of between 30% and 90% by weight, preferably between 50% and 80%, and - a second salt, preferably KC1, in an amount of between 0.1% and 90% by weight of said composition, preferably between 1% and 5%.

[0077] Indeed, the combined use of NaCl with KC1 has shown particularly good performance. Indeed, this guarantees the integrity of the sample while respecting the osmotic limit.

[0078] The solvent is preferably chosen from osmosis water, distilled water and an organic solvent such as ethanol, methanol or ethylene glycol. The choice is gener- generally linked to planned hematology analyses.

[0079] Advantageously, the method comprises a step of homogenizing the grain size of said salt. Optimally, a grain size of between 0.8 mm and 2.2 mm is targeted.

[0080] In this description, when reference is made to a grain size, this is a size measured by microscopy. However, the shape of the grains is polymorphic. Thus, since the shape can vary greatly (in particular spherical, ellipsoidal, cubic, regular or irregular prismoidal, etc.), the sizes are expressed here by referring to a measurement of a distance made on the top of a chosen face of a grain. In practice, the length from one edge to another is measured. Reference is generally not made to the diameter or to a diagonal. This is exemplified in Figures 2 and 3. It is estimated that each face measured on a grain is representative, to within approximately 10%, of the other faces of this grain.

[0081] EXAMPLES OF ACHIEVEMENTS

[0082] EXAMPLE 1: Manufacturing process

[0083] A composition according to the invention was prepared according to the preparatory protocol below.

[0084] I. Mixing of liquid components:

[0085] LI. Tergitol (concentration between 0 and 3%, preferably 1.5%) + Distilled water (or organic solvent (Ethanol, methanol, ethylene glycol) pure or in water (concentration between 1 and 2%).)

[0086] II. Crushing of powdered components (separately):

[0087] II.1. NaCl (concentration between 30% and 90%, preferably between 60% and 90%) and KC1 (concentration between 0% and 90%, preferably between 2.5% and 25%): homogenization and reduction of the size of the seeds (from more than 2.3 mm to a size between 0.8 and 2.2 mm);

[0088] III. Mixing fine powders (0.8-2.2 mm):

[0089] EDTA (concentration between 0% and 8%, preferably between 1% and 6%), pH buffer (e.g. TRIZMA base, Tris HCl): seed size already quite small (concentration between 0% and 20%, preferably between 5% and 10%);

[0090] IV. Mixing of liquid components with fine powders;

[0091] V. Addition of powders (large grains, > 2.3 mm) and homogenization of the mixture;

[0092] VI. Shaping the product (granules or pellets)

[0093] This process makes it possible to obtain in particular a granule composed of solid grains.

[0094] EXAMPLE 2: Granules

[0095] The process described in the present example was implemented with different choices among the components cited in the general description above. Various granules were thus obtained which meet the definition of the composition of the invention.

[0096] [Fig. 1] shows a microscopic photograph of a grain of the composition of the invention in granular form. The microscope is the Olympus Microscope BX-51 model with a Nikon xl0 / 0.25 air objective. The camera is an Olympus DP-23. The figure shows a grain with a diametrical dimension between 0.8 mm and 2.2 mm, more precisely about 1.8 mm x 1.8 mm.

[0097] [Fig. 2] shows a microscopic photograph of several grains of the composition of the invention in granular form. The microscope is the one also used for [Fig. 1]. Grains with dimensions between 0.8 mm and 2.2 mm can be seen.

[0098] The solid composition in granular form shown in Figures 1 and 2 offers increased stability and shelf life. This form also reduces transport and storage costs. In particular, this form allows for a reduction in water volumes and their transport. It is easy to use: simply dilute with an inert liquid. All of this has a positive impact on the environment.

[0099] EXAMPLE 3: Preferred Diluent Formulations

[0100] A first solid composition according to the invention was produced in the proportions indicated in the table below.

[0101] [Tables 5]

[0102] A solution for hematology analyses according to the invention was produced from this first composition. For this, the solid composition is dissolved in osmosis water. This results in a solution in the proportions indicated in the table below.

[0103] [Tableauxô]

[0104] A second solid composition according to the invention was produced in the proportions indicated in the table below.

[0105] [Paintings?]

[0106] A solution for hematology analyses according to the invention was produced from this second composition. For this purpose, the solid composition is dissolved in osmosis water. This results in a solution in the proportions indicated in the table below.

[0107] [Tables 8]

[0108] The performance of the above compositions or solutions has shown particularly satisfactory performance.

[0109] EXAMPLE 3: Formulation of lysis reagent with dye

[0110] [Tables 9] [OR I] EXAMPLE 4: Exploration test

[0112] An exploration test on ten blood samples was conducted to test the performance of the solid composition of the invention (in its dissolved form) in comparison with a conventional liquid diluent for hematology analyzers called "ABX Diluent".

[0113] The ABX Diluent was compared with the solid composition formulated according to the procedure described above, then reconstituted in liquid form in hematological solution by adding osmosis water. In summary, a hematological reagent of the state of the art is compared here with the reconstituted hematological reagent of the invention.

[0114] An example of a red blood cell (RBC) histogram is shown for both reagents. The histograms were produced using a Yumizen H500-OT device available from HORIBA MEDICAL.

[0115] [Fig. 3] shows the GR histograms of the state-of-the-art reagent and the solution prepared from the composition of the invention. Both compositions have almost the same performance, but the solid composition of the invention has all the advantages discussed above.

[0116] This is confirmed by blood counts performed with the Yumizen H500-OT device available from HORIBA MEDICAL.

[0117] [Eig.4] shows comparative blood counts between the state-of-the-art reagent ABX diluent and a solution (or reagent) prepared from the composition of the invention referred to in the figure as "reconstituted solid diluent". Indeed, the composition for hematology analyses of the invention, which is in solid form, is capable of being dissolved in osmosis water to give a hematological solution of the diluent type. The composition can therefore be referred to as a "solid diluent".

[0118] More specifically, [Eig.4] shows the correlations on the main CBC parameters (from the English Complete Blood Count for Blood Count (NES) or "hemogram") for the ten samples tested.

[0119] The table below shows the coefficients of determination for each measured parameter of [Eig.4]

[0120] [Tables 10]

[0121] A very good correlation between the ABX Diluent product and the reconstituted solid Diluent of the invention is observed for all parameters.

[0122] A quality control of the composition of the invention can be carried out from the reconstituted reagent (i.e. from the solution of the invention). Conductivity represents in particular an important parameter which can serve as a basis for quality control. Conductivity measurement is also a relatively simple method to set up and allows a continuous measurement process. In general, existing hematology devices are regulated systems arranged to maintain a conductivity setpoint in the reagent produced. The conductivity is measured within a reservoir and is compared to a reference value. If necessary, the devices are arranged to add osmosis water and / or reagent until the target conductivity is obtained. Existing devices therefore use a control loop allowing the volumes of water and / or reagent to be adjusted until the solution reaches the target conductivity.

[0123] Other control options are possible: detergent concentration, surface tension test and / or verification of certain hematological parameters such as MCV or hemoglobin using control blood.

Claims

Claims

1. Composition for hematology analyses, characterized in that it comprises: - an anticoagulant in an amount of between 0.1% and 8% by weight, preferably between 2% and 6%; - a salt in an amount of between 30% and 90% by weight, preferably between 50% and 80%; - a pH buffer in an amount of between 0.1% and 50% by weight, preferably greater than or equal to 10%; and - a surfactant in an amount of between 0.01% and 2% by weight, preferably between 0.05% and 1.5%, and in that it is in solid form.

2. A composition according to claim 1, wherein - the amount by weight of the anticoagulant is between approximately 3% and approximately 4%; - the amount by weight of the salt is between about 50% and about 70%; - the amount by weight of the pH buffer is between about 30% and about 45%; and - the amount by weight of the surfactant is between about 0.05% and about 0.80%.

3. Composition according to one of the preceding claims, in which the salt is composed of: - a first salt, preferably NaCl, in an amount of between 30% and 90% by weight, preferably between 50% and 80%, and - a second salt, preferably KC1, in an amount of between 0.1% and 90% by weight of said composition, preferably between 1% and 5%.

4. Composition according to one of the preceding claims, in which the anticoagulant is chosen from the group consisting of EDTA, K2-EDTA, K3-EDTA, Na2EDTA, F acid-citrate-dextrose (ACD), lithium heparin (LH), sodium heparin (NH), and sodium citrate.

5. Composition according to one of the preceding claims, in which the anticoagulant is a chelating agent.

6. Composition according to one of the preceding claims, in which the pH buffer is chosen from the group consisting of K 2 HPO 4 ,3H 2 O, NaH 2 PO 4 ,2H 2 O, Na 2 HPO 4 ,12H 2 O, KH 2 PO 4 and the Tris NH buffer 2 C(CH 2 OH) 3 , NH 2 C(CH 2 OH)3HC1 OR a combination thereof.

7. A composition according to any preceding claim, wherein the surfactant is selected from the group consisting of Tween 20, Tween 80, Tergitol 15-S-9, Brij L23, Brij 35, and SB3-12.

8. A composition according to claim 1, having the following formulation:

9. A composition according to claim 1, having the following formulation:

10. Composition according to one of the preceding claims in a form chosen from, a tablet, a powder or a granule.

11. Composition according to one of the preceding claims consisting of: an anticoagulant in an amount of between 0.1% and 8% by weight, preferably between 2% and 6%; a salt in an amount of between 30% and 90% by weight, preferably between 50% and 80%; a buffer pH in an amount between 0.1% and 50% by weight, preferably greater than or equal to 10%; a surfactant in an amount between 0.01% and 2% by weight, preferably between 0.05% and 1.5%; and optionally traces of solvent.

12. Composition according to one of the preceding claims, free from preservative.

13. Solution for hematology analyses obtained by dissolving the composition according to one of claims 1 to 12 comprising: - an anticoagulant, in an amount of between 0.01% and 0.2% by weight, preferably between 0.05% and 0.15%; - a first salt, preferably NaCl, in an amount of between 1.15% and 3.5% by weight, preferably between 0.8% and 0.9%; - a second salt, preferably KC1, in an amount of between 0.01% and 0.9% by weight, preferably between 0.02% and 0.04%; - a pH buffer in an amount of between 0.01% and 0.9% by weight, preferably 0.5%; - a surfactant in an amount of between 0.001% and 0.02% by weight, preferably between 0.01% and 0.015%; and - a solvent, preferably osmosis water, in a quantity remaining to obtain 100% by weight of said solution.

14. Method for manufacturing a composition for hematology analyses, characterized in that it comprises the following steps: a. providing the following components: - an anticoagulant, in an amount of between 0.1% and 8% by weight of said composition, the anticoagulant being in the form of a fine powder having a grain size of between approximately 0.8 mm and approximately 2.2 mm; - a salt, in an amount of between 30% and 90% by weight of said composition, the salt being in the form of a fine powder having a grain size of between approximately 0.8 mm and approximately 2.2 mm; - a pH buffer, in an amount of between 0.1% and 50% by weight of said composition, the pH buffer being in the form of a fine powder having a grain size of between approximately 0.8 mm and about 2.2 mm; - a liquid surfactant in an amount of between 0.01% and 2% by weight of said composition, mixed with a solvent in a surfactant:solvent ratio of between 1:10 and 1:40 b. mixing the anticoagulant with the pH buffer so as to obtain a fine powder mixture; c. adding the liquid surfactant mixed with the solvent to said fine powder mixture so as to obtain a pasty premix; d. adding the salt to said pasty premix followed by homogenization so as to obtain a paste; and e. resting said paste to obtain said composition in a solid form.

15. A method according to claim 14, wherein the salt is composed of: - a first salt, preferably NaCl, in an amount of between 30% and 90% by weight, preferably between 50% and 80%, and - a second salt, preferably KC1, in an amount of between 0.1% and 90% by weight of said composition, preferably between 1% and 5%.

16. Method according to one of claims 15 and 16, in which the solvent is chosen from osmosis water, distilled water and an organic solvent.

17. Method according to one of claims 14 to 16, further comprising a step of homogenizing the grain size of said salt to obtain said grain size between 0.8 mm and 2.2 mm.

18. A method according to one of claims 14 to 17, wherein the homogenization of step d. comprises coating the salt grains with at least a portion of said surfactant.

Images