Apparatus for diffusing or colouring and determining settling rate

Abstract

A device for diffusing or colouring and determining sedimentation rate comprises a first group (4) arranged to sample blood samples in a cuvette and to perform smear testing on the samples, and a second group (6) arranged to sample blood samples in a cuvette and to perform sedimentation rate determination. The device comprises at least one sampling member (8) controllable to operate by said first group (4) and by said second group (6) to sample blood samples such that samples collected for said first group (4) are not used by said second group (6) and samples collected for said second group (6) are not used by said first group (4), the second group (6) is equipped with a sensor (20) comprising an infrared light source (12) and an optical sensor (14), which are arranged substantially oppositely around a cuvette (16) connected to the output of the at least one sampling element (8), the light emitted by the infrared light source (12) reaches the optical sensor (14) after passing through the test tube (16). The apparatus further comprises a converter (10) arranged to receive the one or more light transmission measurements from the optical sensor (14) and to determine the settling rate.

Description

[0001] The present invention relates to the field of hematology, and more specifically to the determination of sedimentation rate (hereinafter referred to as ESR, also known in English as "erythrocyte sedimentation rate" or "ESR").

[0002] The sedimentation rate is part of the routine examination of the complete blood count (hereinafter referred to as CBC, standing for "complete blood count") performed during the blood evaluation and can detect the presence of inflammatory or infectious pathologies. For example, rheumatic diseases, cancer and other diseases that cause changes in the concentration of proteins in the blood. A high ESR indicates an inflammatory state, but does not prejudge its nature. However, when an inflammatory syndrome is present, the ESR remains low. Therefore, a high ESR usually indicates the presence of pathology. Combining this nonspecific test with complementary tests allows a more accurate diagnosis.

[0003] The reference method for measuring ESR is the Westergren method. A blood sample collected with an anticoagulant (e.g. EDTA) is placed in a standard-sized capillary tube, diluted with sodium citrate and placed vertically. The ESR measurement is the height of the plasma column in millimeters after one hour of sedimentation. For example, a height of less than 10 millimeters after one hour of sedimentation is considered normal for an adult male. Normal limits are not absolute values ​​but depend on the patient's age and sex.

[0004] ESR is the result of three steps: aggregation, sedimentation and settling of red blood cells or blood cells.

[0005] The phenomenon of erythrocyte aggregation is caused in particular by blood proteins, whose aggregation force increases with increasing concentration. Aggregates are formed by stacking of erythrocytes into "rolls" and then into three-dimensional structures. The sedimentation rate depends on the size of the aggregates and the viscosity of the plasma.

[0006] In the final step, the aggregates sediment, i.e. they gradually fall to the bottom of the tube and compact, thus separating the sample into a clear portion at the top (plasma) and a much darker portion at the bottom (red blood cells). The ESR is measured by the height of the clear portion.

[0007] The most relevant inflammatory parameter is the aggregation phenomenon, which directly depends on the concentration of plasma proteins. It is this phenomenon that is measured in the present invention to return the ESR value. It is also a non-specific parameter, but it suppresses interference caused by the last two steps of the ESR. There is no standardized parameter that can express this aggregation rate or dynamics / kinetics. Therefore, it is relevant to convert the aggregation measurement into ESR.

[0008] Due to the amount of blood (1.6 ml) and the time required for completion (1 hour), the Westergren method is not compatible with hematology machines used for CBC analysis. To overcome these problems, studies have been conducted, such as in patent US 6,632,679, to determine the sedimentation rate using optical measurements of extinction (absorption and scattering).

[0009] Some patents, such as EP2921862, have proposed integrating improved ESR measurements into blood testing devices that collect a volume of blood and separate it into two parts for separate counts (CBC) and sedimentation rate measurements.

[0010] Therefore, the device links the count measurement and the sedimentation rate measurement, which is disadvantageous because it requires a complex device to distribute the two blood sample parts through one sample to the count part on one side and the sedimentation rate measurement part on the other side, especially since the CBC and ESR measurement operations do not have the same requirements in terms of sample preparation.

[0011] More generally, in automated laboratories of the TLA type ("Total Laboratory Automation" in English), the tube flows are managed in a fully automated way, from the most frequent to the least frequent tests. The instrumentation required to supply the sample to the measuring cell is by far the most expensive and complex part, and includes:

[0012] -interconnection with the chain,

[0013] -Management of test tube racks,

[0014] - stirring of the test tube,

[0015] - Management of test tubes,

[0016] - sampling and transfer in the measuring unit,

[0017] - Flushing of sampling equipment, and

[0018] - Equipment for control and transmission of results.

[0019] Furthermore, this results in a very high ratio between the cost of the measurement system and the cost of the infrastructure required to manage the samples (greater than 100). In contrast, for CBC / DIF / RET measurement systems or automated diffusion or staining systems commonly used in hematology laboratories, this ratio is much lower (2 to 5).

[0020] On the other hand, laboratories that process a large number of test tubes every day require a limited processing time for newly arrived test tubes (in English, "turnaround time" or TAT), which directly affects the size of the CBC / DIF / RET measurement system and thus its hardware cost.

[0021] On the other hand, since a small fraction of processed tubes require diffusion or staining (only 5% to 20%), all manufacturers typically oversize the system.

[0022] Document WO2021 / 097610A1 describes a sample analyzer, including a module for measuring erythrocyte sedimentation rate, a module for measuring blood image, and a sample distribution module. The sample distribution module is used to collect a blood sample, distribute a first portion of the blood sample to the erythrocyte sedimentation rate measurement module, and distribute a second portion of the blood sample to the complete blood cell count measurement module. The erythrocyte sedimentation rate measurement module includes a measuring tube and an optical measuring device.

[0023] There is no known equipment that can effectively determine sedimentation rate measurements in a fast and comprehensive manner under the requirements of modern laboratories.

[0024] The present invention improves this situation. To this end, it provides a device for diffusing or staining and determining sedimentation rate, the device comprising a first group arranged to sample a blood sample in a test tube and perform a smear test on the sample, and a second group arranged to sample a blood sample in a test tube and perform a sedimentation rate determination. The device comprises at least one sampling member, which can be controlled by the first group to operate and controlled by the second group to operate to sample the blood sample, so that the second group does not use the sample collected for the first group, and the first group does not use the sample collected for the second group, and the second group is equipped with a sensor, which includes an infrared light source and an optical sensor, which are arranged approximately oppositely around a test tube connected to the output end of at least one sampling member, so that the light emitted by the infrared light source reaches the optical sensor after passing through the test tube. The second group is also arranged to rinse the sampling member and the tube between two sedimentation rate measurement determinations, and the optical sensor is arranged to perform a blank measurement after the rinsing operation. The apparatus further comprises a transducer arranged to receive a blank measurement and one or more light transmission measurements from the optical sensor and to determine a sedimentation rate based on a ratio between the blank measurement and the light transmission measurements.

[0025] The device is particularly advantageous because it allows sedimentation rate measurement to be performed independently of the complete blood count. Therefore, the sedimentation rate measurement remains a less systematic test and does not interfere with the architecture of the device in a way that could affect other functions.

[0026] According to various embodiments, the present invention may have one or more of the following features:

[0027] - The infrared light source and optical sensor are arranged at a distance of less than 10 cm from the sampling end of the sampling piece,

[0028] The second group is arranged at the inlet of the device, upstream of the first group, or at the outlet of the device for carrying out complementary measurements of the diffusion or coloration type downstream of the first group.

[0029] - a converter is arranged to determine a minimum light transmission measurement point in time and a final light transmission measurement point in time,

[0030] - the optical sensor is arranged to achieve a maximum gain between the lowest light transmission measurement time point and the final light transmission measurement time point, and a minimum gain at the rest of the time,

[0031] - the converter is arranged to calculate the sedimentation rate from the ratio between the ratio of the blank measurement value and the measurement value of the final light transmission measurement time point on the one hand and the ratio of the blank measurement value and the measurement value of the lowest light transmission measurement time point on the other hand.

[0032] - the optical sensor is controlled at a low gain before the blood passes through the substantially transparent portion and at a high gain thereafter, and

[0033] The sampling member is a needle which can be controlled to collect a blood sample, the needle being connected to a test tube in which a substantially transparent portion is formed.

[0034] The invention also relates to a method for diffusion or staining and determining sedimentation rate, characterized in that the method comprises the use of a device according to the invention and that the diffusion or staining on the one hand and the determination of the sedimentation rate on the other hand comprise the acquisition of two different samples.

[0035] Other characteristics and advantages of the invention will better emerge on reading the following description, with reference to examples given for illustrative and non-limiting purposes, with reference to the accompanying drawings, in which:

[0036] - Figure 1 shows a general schematic diagram of an apparatus according to the invention,

[0037] - Figure 2 Shows Figure 1 The manufacturing details of the components,

[0038] - Figure 3 Shows Figure 1 Measurement diagram of the equipment, and

[0039] - Figure 4 Shows Figure 1 A schematic diagram of an embodiment of a device.

[0040] The accompanying drawings and the following description essentially contain components of a certain nature. Therefore, they not only contribute to a better understanding of the invention, but also contribute to the definition of the invention where appropriate.

[0041] Figure 1 The overall schematic diagram of the device 2 for performing a complete blood cell count and determining a sedimentation rate measurement according to the present invention is shown. The device 2 comprises a first set 4, a second set 6, a sampling member 8 and a converter 10.

[0042] The first group 4 is arranged to perform blood smear tests. Devices that automatically perform blood smear tests are very common in laboratories that have to process a large number of test tubes every day. Their processing rate is usually too large relative to the laboratory's needs.

[0043] This oversizing is directly related to the technical complexity of the different steps required to prepare the slides for each device (sampling, depositing a drop of aliquot on the slide, diffusion, slide identification, vital staining, etc.): high rates are the only means by which manufacturers can optimize marginal costs.

[0044] Since the blood smear test is a less systematic test than the CBC, the Applicant found it interesting to pool the resources of the first group 4 in order to add the determination of the ESR measurement value through the second group 6 .

[0045] The second group 6 is arranged to determine the ESR measurement. This second group is the main object of the present invention. In fact, as mentioned in the background introduction section, this test, whether carried out according to the conventional method or by measuring the change in light absorbance, is either cumbersome or combined with a CBC measurement, which slows down the whole device.

[0046] According to the invention, the implementation of the second set 6 allows to infer sedimentation rate measurements by changing the light absorbance in a manner separate from any CBC measurement or other blood measurement. To this end, the second set 6 is arranged to control the sampling member 8, here a needle, independently of the first set 4, i.e. the sample collected by the first set 4 is only used to perform a blood smear test, while the sample collected by the second set 6 is only used for ESR measurement.

[0047] Therefore, if Figure 1 As shown, the first group 4 or the second group 6 controls the needle 8 to collect a blood sample in the blood vessel 11. Afterwards, the sample is taken to the first group 4 for a blood smear test, or to the second group 6 for measurement of light absorbance changes. In the latter case, the measurement result is transmitted to the converter 10, which returns an ESR signal and / or an ESR measurement value.

[0048] Figure 2 An embodiment of a second measurement group 6 is shown. According to this example, the second measurement group 6 is constituted by a light source 12 and an optical sensor 14 assembled relative to each other around a tube 16 for conveying a blood sample collected by a needle 8 from a blood vessel 11 to the interior of the device 2 .

[0049] In the example described herein, the light source 12 is of LED type, and its infrared wavelength is preferably in the range of 700-980 nm, in particular 800 nm, which is the isosbestic point between oxygenated hemoglobin and deoxygenated hemoglobin, which makes the measurement insensitive to blood oxygen saturation. In the example described herein, the optical sensor 14 is of photodiode type, which can be selected from sensors such as PMT, PDA, CMOS, etc. In the example described herein, the cuvette 16 is made of Teflon and is connected to the needle 8. Alternatively, the cuvette 16 can be made of glass or plastic and should be selected to provide good transparency to the wavelength of the light source 12.

[0050] The light source assembly 12 and the optical sensor 14 can be considered as a single sensor 20 of the second group 6. As in the example described herein, it can be made in two parts, assembled together to buckle the needle 8 (for example metal) and the test tube 16. Alternatively, the light source 12 and the optical sensor 14 can be made in one piece. Preferably, the sensor 20 is arranged very close to the end of the needle 8, less than 10 cm from the latter, to optimize the shearing of the red blood cells. Preferably, this distance is about 5 cm to obtain the best results. In a variant, the sensor 20 can be placed directly at the outlet of the needle 8.

[0051] Whatever the configuration, the second set 6 allows advantageous use of a diameter of the needle 8 which is generally in the range of 1 mm. In fact, during the sampling process with the needle 8, the red blood cells undergo a continuous deformation, which will be described below and as Figure 3 shown.

[0052] Thus, once the needle 8 is immersed in the blood vessel 11, the second group 6 controls the aspiration of the needle 8, which has been primed with the diluent, in order to collect the blood. This is done thanks to automated control methods, preparation trays and solenoid valves and syringes, which are also known and, for the sake of simplicity, are not shown in the figures. These methods cause the diluent occupying the test tube 16 to move. During this step, the sensor 20 measures a constant light signal due to the translucent nature of the test tube 16 and the diluent contained therein.

[0053] Afterwards, the blood aliquot is moved upward to the sensor 20 by suction. The red blood cells are sheared, thereby breaking up the aggregation. During this step, the optical signal remains at a maximum because the blood is not yet opposite to the sensor 20. Figure 3 This is indicated by reference numeral 30 .

[0054] When the blood reaches the sensor, the optical signal indicates the shear state of the blood. In this blood shear stage, the red blood cells have an elongated shape, as shown by reference numeral 31.

[0055] When aspiration is stopped to stop shearing, the optical transmission signal decreases as the red blood cells resume their biconcave disc-shaped relief form. Figure 3In the embodiment, this is indicated by reference numeral 32. Afterwards, the optical signal increases according to a pseudo-logarithmic progression. The gradual increase in the optical signal measured by the optical sensor 14 is associated with the gradual aggregation of free red blood cells. Aggregation is accomplished by stacking red blood cells to form a roll and subsequent three-dimensional structure, which is achieved in Figure 3 In the drawings, reference numerals 34 and 36 are used to indicate the same.

[0056] After about 40 seconds, depending on the blood, aggregation slows down considerably and sedimentation begins. The onset of sedimentation interferes with the measurement of interest, namely aggregation and its correlation with sedimentation rate. 4 Afterwards, the blood aliquot is discharged, for example by moving the needle 8 onto the tray. Thus, the blood is discharged and the test tube 16 is flushed with the diluent, and at the end of this step, the test tube 16 is completely filled with the diluent, just as at the beginning of the above-described procedure.

[0057] Figure 3 Four time points are shown:

[0058] - At time t 1 Previously, test tube 16 was filled with diluent,

[0059] - Time point t 1 Mark the point in time when the blood reaches the level of the sensor 20 by aspiration,

[0060] - Time point t 2 Mark the time when the suction stops.

[0061] - Time point t 3 Marks the point in time when red blood cells regain their relaxed shape and begin to aggregate,

[0062] - Time point t 4 Mark the end of the procedure, when the blood is drained and replaced again with diluent, as at time t 1 Same as before.

[0063] Figure 3 Observations show that at time t 4 It may seem paradoxical that the optical signal is less important when the tube 16 is filled with diluent than when the tube 16 is filled with blood. This is explained by the fact that in the example described herein, the optical sensor 14 is Figure 3 Time point t 1 Before and at time t 4 The subsequent adjustments are different.

[0064] Therefore, at time t 1 Before and at time t 4Afterwards, it is determined that the measured value corresponds to a “blank” measured value. During this time, the optical sensor 14 is regulated by the converter 10 with the minimum measurement gain.

[0065] Time point t 3 Corresponding to the period t 1 -t 4 The minimum point beyond which the optical signal begins to increase, which, as mentioned above, marks the beginning of the phenomenon that the present invention aims to measure. For this reason, and because the variation of the optical signal between time point t0 and time point t 1 In the example described herein, the optical sensor 14 is adjusted with minimum gain until time t 1 , then at time t 1 and t 4 The maximum gain is adjusted between 4 Then adjust again to minimum gain for the next measurement.

[0066] This is even more advantageous because, in the embodiment described herein, the converter 10 is arranged to determine the measurement value based on the optical density of the signal measured by the optical sensor 14. It should be recalled that the optical density is defined by the formula DO(t)=log(KI0 / I(t)), where I(t) is the measurement value of the optical sensor at the time point t and K is the maximum gain / minimum gain ratio. More specifically, the converter 10 is arranged to determine the measurement value based on DO(t 3 ) / DO(t 4 ) ratio returns a sedimentation rate measurement. Applicants have made a number of sedimentation rate measurements using the Westergren reference method, which enables the converter 10 to correlate the measurements calculated therefrom with ESR values.

[0067] The applicant has found that the use of optical density is particularly advantageous, as it makes it independent of any variations in the transmittance measurement. The applicant has also found that the converter 10 can also be based on the ratio I(t 4 ) / I(t 3 ) without having to consider the optical density in the Beer-Lambert meaning and using I0 differently in the calculations.

[0068] The converter 10 can be made in various ways, such as in the form of a suitable computer code executed on one or more processors. The processor should be understood as any processor suitable for the calculations described below. Such a processor can be made in any known way, such as a microprocessor of a personal computer, laptop, tablet or smart phone, a dedicated chip of the type FPGA or SoC, a computing resource in a grid or cloud, a graphics processor (GPU) cluster, a microcontroller, or any other form that can provide the computing power required to complete the process described below. One or more of these elements can also be made in the form of a dedicated electronic circuit (such as an ASIC). A combination of a processor and an electronic circuit can also be considered. In the case of a machine learning unit based on gradient boosting, a processor dedicated to machine learning can also be considered. Alternatively, the converter 10 can be an analog computer without any programming or computer code.

[0069] Alternatively, the converter 10 may use a machine learning algorithm, which may or may not involve a neural network (deep or non-deep). This involves correlating the intensity measurement of the optical sensor 14 with the sedimentation rate value. This variant may be particularly suitable for the case where there is no optical density. Alternatively, the gain of the optical sensor 14 may be the same for all time points of the measurement.

[0070] Figure 4 Shows Figure 1 Schematic diagram of an embodiment of a device. As shown, the device 2 is a conventional hematology device, including a syringe 40 with a diameter of 1.5 mm and a syringe 42 with a diameter of 16 mm, both of which are connected to a needle 8. The sensor 20 has been integrated on the needle 8 so that it covers the end of the test tube 16 connected to the needle 8.

[0071] Figure 4 The device 2 is suitable for performing smear tests, ESR measurements, or both.

[0072] To perform a smear test on the first group 4, the entire circuit is primed with a diluent, then the needle 8 is inserted into the blood vessel 11 and about 75 microliters are sampled with a syringe. During its lifting, the outside of the needle 8 is gradually washed by the liquid flow pushed to one side and is continuously sucked and discharged into the waste liquid on the other side.

[0073] After that, the syringe performs a column removal in order to transfer the aliquot at the Y level of the pinch valve 44 opened on the needle 8 side. The pinch valve is activated to close the channel originating from the needle 8 and open the channel towards the drop needle 46. After that, the syringe performs a push, allowing the sample to be transferred to the drop needle. A series of peripheral flushing and then drying of the drop needle ensures the quality of the initial front end of the aliquot. Finally, a drop of a few microliters is deposited on the slide 48 by pushing the syringe.

[0074] For ESR measurements, it is performed as described above and refers to Figures 1 to 3 . The entire circuit is pretreated with a diluent drawn from the reservoir 44, and then a few microliters of bubbles are generated at the tip of the needle 8. The needle 8 is placed in the blood vessel 11, and an aliquot of 50 to 100 microliters is collected by the syringe 42. The needle 8 is lifted and the aliquot is transferred to the sensor 20 and an optical measurement of the aggregate is performed. Finally, the needle 8 is internally and externally flushed in the tray and the discharge is discharged into the waste liquid. A blank measurement (I0) can be performed before sampling or at the end of flushing. The blank measurements before and after can optionally be compared to enable abnormal handling, such as flushing control.

[0075] The smear test and the ESR cycle are performed independently of each other and in particular involve separate sampling of needle 8: an aliquot of the smear test cannot be used for the ESR measurement and vice versa.

[0076] This allows the two operations to be separated without any problematic blood sampling when two measurements need to be made. This independence makes measuring the ESR much less cumbersome for the device 2's operating rate and can be integrated at a lower cost, both in terms of instrumentation and manpower.

[0077] The device of the present invention is much more efficient than all known systems which, in addition to providing diffusion or staining modules, also provide specific modules for measuring sedimentation rate or save CBC / DIF / RET processing time on a universal module.

[0078] Alternatively, the second set 6 may be integrated on a sampling needle of a high-end device comprising a specific sampling module for subsequent diffusion or colorimetric measurements.

[0079] The present invention thus allows sedimentation rate measurements to be efficiently integrated into existing equipment without affecting its operating rate or structure.

Claims

1. An apparatus for diffusion or staining and determining sedimentation rate, comprising a first group (4) arranged to take a sample of blood from a test tube and to perform a smear test on the sample, characterized in that The device comprises a second group (6) arranged to sample a blood sample in a test tube and to determine the sedimentation rate, the device comprising at least one sampling member (8) which can be controlled by the first group (4) and controlled by the second group (6) to sample the blood sample, so that the sample collected for the first group (4) is not used by the second group (6), and the sample collected for the second group (6) is not used by the first group (4), the second group (6) is equipped with a sensor (20), the sensor (20) comprising an infrared light source (12) and an optical sensor (14), the infrared light source (12) and the optical sensor (14) surrounding a device connected to the at least one sampling member (8). The test tube (16) at the output end of the sample piece (8) is arranged approximately opposite to each other so that the light emitted by the infrared light source (12) reaches the optical sensor (14) after passing through the test tube (16). The second group (6) is also arranged to rinse the sampling piece (8) and the test tube (16) between two sedimentation rate measurements, and the optical sensor (14) is arranged to perform a blank measurement after the rinsing operation. The device also includes a converter (10), which is arranged to receive a blank measurement value and one or more light transmission measurement values ​​from the optical sensor (14) and determine the sedimentation rate based on the ratio between the blank measurement value and the light transmission measurement value.

2. The device according to claim 1, wherein: The infrared light source (12) and the optical sensor (14) are arranged at a position less than 10 centimeters away from the sampling end of the sampling member (8).

3. The device according to any one of the preceding claims, wherein: The second group (6) is arranged at the inlet of the device (2), upstream of the first group (4), or at the outlet of the device (2) for performing complementary measurements of the diffusion or coloration type downstream of the first group (4).

4. The device according to any one of the preceding claims, wherein: The converter (10) is arranged to determine a minimum light transmission measurement time point (t3) and a final light transmission measurement time point (t4).

5. The device according to claim 4, wherein: The optical sensor (14) is arranged to achieve a maximum gain between a minimum light transmission measurement time point (t3) and a final light transmission measurement time point (t4), and to achieve a minimum gain at the remaining time.

6. The device according to claim 4 or 5, wherein: The converter (10) is arranged to calculate the sedimentation rate based on the ratio between the ratio of the blank measurement value and the measurement value of the final light transmission measurement time point (t4) on the one hand and the ratio of the blank measurement value and the measurement value of the minimum light transmission measurement time point (t3) on the other hand.

7. The device according to any one of the preceding claims, wherein: The optical sensor (14) is controlled at a low gain before blood passes through the substantially transparent portion and at a high gain thereafter.

8. The device according to any one of the preceding claims, wherein The sampling member is a needle (8) which can be controlled to collect a blood sample. The needle is connected to a test tube (16), and a substantially transparent portion (16) is formed in the test tube (16).

9. A method for diffusing or coloring and determining sedimentation rate, characterized in that The method comprises the use of a device according to one of claims 1 to 8 and is characterized in that carrying out the diffusion or staining on the one hand and determining the sedimentation rate on the other hand comprises taking two different samples.

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