Bone marrow smear preparation method based on air source purging, system equipment and medium

Abstract

The invention discloses a bone marrow smear preparation method based on air source purging, system equipment and a medium, and relates to the technical field of medical examination and cell preparation, and the method comprises the following steps: obtaining and processing a bone marrow cell sample to form a cell suspension; applying the cell suspension to a glass slide, and performing directional treatment on the cell suspension in an air source purging manner; fixing and dyeing the bone marrow smear which is formed by blowing through an air source, so as to complete the smear preparation operation of the bone marrow smear. According to the method, smear quality fluctuation caused by sample state difference is remarkably reduced, a uniform basic suspension condition is provided for stable development of subsequent airflow purging, smear preparation consistency and reproducibility are optimized from the source, smear region distribution quality improvement and stable acquisition of morphological analysis results are achieved, and the method is suitable for large-scale popularization and application. The dyeing uniformity and the color development consistency are improved, and better effects are achieved in the aspects of cell resuspension uniformity, cell spreading consistency and film production process standardization.

Description

Technical Field

[0001] The present invention relates to the technical field of medical testing and cell preparation, in particular to a bone marrow smear preparation method based on air source purge, system equipment and medium. Background Art

[0002] In clinical hematology and bone marrow disease research, bone marrow morphology is an important tool for directly observing the developmental state and proportions of hematopoietic cells and is widely used in the detection and differential diagnosis of various hematologic diseases. Since the 20th century, bone marrow smear preparation, a key pre-processing step for morphological analysis, has been primarily based on the traditional manual push-slide method. This method involves the operator stretching a cell suspension along the surface of a glass slide, spreading the cells into a thin layer, thereby obtaining diagnostically valuable cellular images for subsequent staining and microscopic observation. With the rapid growth in clinical testing demand and the increasing demand for image standardization and consistent results, the traditional push-slide method, which relies heavily on operator skill and experience, has gradually exposed problems such as poor reproducibility, uneven cell distribution, and time-consuming and labor-intensive procedures. To meet the demands of modern medicine for high-throughput, standardized, and intelligent testing processes, research has begun exploring automated smear systems, electronically controlled push-slide devices, and even microfluidic coating technologies, aiming to achieve consistent smear quality and automation through mechanical control. However, most automated smear devices are currently limited to peripheral blood cell smear preparation, and most methods still rely on mechanical push-pull or slide spreading, resulting in complex structures and poor adaptability. These methods also have limited control over the uniform distribution of diverse cell types and morphologies, such as granulocytes and primitive cells, within bone marrow samples. Furthermore, the inherent high viscosity and large fluctuations in cell concentration of bone marrow samples make traditional push-slide or mechanical spreading methods even more challenging in terms of sample compatibility. The resulting smears often exhibit problems such as cell accumulation, blank areas, or discontinuous staining.

[0003] In response to the above problems, some studies in recent years have introduced gas source or airflow-assisted distribution strategies, attempting to use gas force instead of traditional mechanical action to achieve cell spreading on the slide. For example, some studies have used directional gas flow to promote the diffusion of blood samples, thereby improving the problem of cell accumulation. However, in bone marrow smear applications, there is still a lack of detailed parameter control and a mechanism for implementing standardized paths. In the existing literature, there are few technical solutions for systematic modeling and purge path optimization for bone marrow sample characteristics. The following deficiencies are common: there is a lack of real-time identification and feedback adjustment mechanism for sample distribution status. Airflow control parameters (such as pressure, flow, and angle) are often statically set, making it difficult to dynamically respond to changes in sample status, which can easily lead to cell rupture or over-sparse distribution; the purge operation is not coordinated with the slide motion path, making it impossible to achieve controllable and uniform coating across the entire slide area; the fixation and staining process is disconnected from the purge step, making it impossible to implement a closed operation process in a unified system. Manual step-by-step processing is still required, which restricts the development of system integration. Summary of the Invention

[0004] In view of the above-mentioned problems, the present invention is proposed.

[0005] Therefore, the technical problem solved by the present invention is: the existing bone marrow smear preparation method has the problems of manual operation, uneven cell distribution, low standardization, poor adaptability, and how to achieve uniform spreading of cell suspension and high-quality preparation based on air source control.

[0006] To solve the above technical problems, the present invention provides the following technical solutions: a bone marrow smear preparation method based on air source purge, comprising obtaining and processing a bone marrow cell sample to form a cell suspension; applying the cell suspension to a glass slide and directionally treating the cell suspension by air source purge; fixing and staining the bone marrow smear formed by the air source purge to complete the bone marrow smear preparation operation; the air source purge comprises starting an air source purge device after dropping the cell suspension on the glass slide, and forming a directional airflow path by adjusting the gas output parameters. , so that the cell suspension is spread on the surface of the slide, and the gas parameters are dynamically adjusted during the purge process according to the spreading state of the cell suspension, so that the cell suspension is evenly spread to a preset area; the bone marrow smear fixation and bone marrow smear staining include placing the bone marrow smear in a fixed environment after the gas source is purged to form a bone marrow smear, applying a fixative to the surface of the bone marrow smear and maintaining it for a preset fixation time, applying a staining solution to the surface of the fixed bone marrow smear and maintaining it for a primary staining time, applying a buffer solution and maintaining it for a secondary staining time, and then rinsing and drying the stained bone marrow smear.

[0007] As a preferred embodiment of the bone marrow smear preparation method based on air source purge described in the present invention, the obtaining and processing of the bone marrow cell sample includes centrifuging the collected bone marrow suspension to form a bottom sediment of the bone marrow cells in the centrifuge tube, removing the supernatant, adding physiological saline to the bottom sediment for resuspending to form a cell suspension, and pipetting the cell suspension with a pipette.

[0008] As a preferred embodiment of the bone marrow smear preparation method based on air source purge described in the present invention, the method comprises: inserting a pipette tip equipped with a pipette tip below the liquid surface of the cell suspension, performing suction and discharge operations in sequence, and fully mixing the cells and the liquid by a preset number of blows. The blows are performed until the cell suspension reaches a preset uniformity requirement, and the pipette tip is prevented from contacting the container wall during the blows.

[0009] As a preferred embodiment of the bone marrow smear preparation method based on gas source purge described in the present invention, the gas source purge includes: after the cell suspension is dropped onto the glass slide, the gas source purge device is started, and the gas output parameters are adjusted according to the sample characteristics. By adjusting the gas output parameters, the airflow stability and coverage range are controlled to form a directional purge path. During the purge process, the gas output parameters are adjusted according to the spreading state of the cell suspension, and the cell suspension is evenly spread along the surface to a predetermined area in combination with the movement rhythm of the glass slide.

[0010] As a preferred embodiment of the bone marrow smear preparation method based on gas source purge described in the present invention, the adjustment of the gas output parameters includes, during the purge process, judging the spreading state of the cell suspension; if the spreading state is judged to be an aggregation state, adjusting the position and angle of the nozzle, and controlling the direction of the airflow to align with the aggregation area for directional purge.

[0011] If the spreading state is determined to be a local blank, the gas flow rate and pressure are adjusted to replenish the cell suspension in the local blank area.

[0012] As a preferred embodiment of the bone marrow smear preparation method based on air source purge described in the present invention, the bone marrow smear fixation includes placing the bone marrow smear in a fixed environment, applying a fixative to the surface of the bone marrow smear, and removing the residual fixative after a preset fixation time.

[0013] As a preferred embodiment of the bone marrow smear preparation method based on air source purge described in the present invention, the bone marrow smear staining includes applying a staining liquid to the surface of the bone marrow smear after fixation so that the staining liquid covers the bone marrow smear area, maintaining the primary staining time, applying a buffer solution to the surface of the bone marrow smear, maintaining the secondary staining time, rinsing the bone marrow smear, and drying the rinsed bone marrow smear.

[0014] Another object of the present invention is to provide a bone marrow smear preparation system based on air source purge, which can solve the problems of current bone marrow smear automatic preparation technology, such as complex structure, inflexible parameter adjustment and discontinuous process, by setting up a sample processing module, an air source purge module and a fixation and staining module.

[0015] As a preferred embodiment of the bone marrow smear preparation system based on air source purge of the present invention, it includes:

[0016] A computer device includes a memory and a processor, wherein the memory stores a computer program, and the processor executes the computer program to implement the steps of a bone marrow smear preparation method based on gas source purge.

[0017] A computer-readable storage medium stores a computer program, which, when executed by a processor, implements the steps of a bone marrow smear preparation method based on air source purge.

[0018] Beneficial effects of the present invention: The bone marrow smear preparation method based on air source purge provided by the present invention significantly reduces the fluctuation of smear quality caused by sample state differences by centrifuging the bone marrow cell sample to remove the supernatant, adding an appropriate amount of diluent for resuspending, and combining a preset number of pipette puffing operations. It also provides a uniform basic suspension condition for the stable development of subsequent air flow purge, optimizes the preparation consistency and reproducibility from the source, applies the cell suspension to the slide, and directionally treats the cell suspension by air source purge to form a highly uniform and highly consistent single-layer cell distribution layer. It is widely applicable to smear scenarios of multiple types of samples such as blood, bone marrow, and hematopoietic stem cells, and ultimately achieves improved smear regional distribution quality and stable acquisition of morphological analysis results. The bone marrow smear formed by air source purge is fixed and stained to complete the bone marrow smear preparation operation, further improving the staining uniformity and color consistency, and avoiding the staining deviation caused by uneven thickness in traditional staining. The present invention achieves better results in cell resuspension uniformity, cell spreading consistency, and preparation process standardization. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following briefly introduces the drawings required for use in the description of the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0020] Figure 1 This is an overall flow chart of the bone marrow smear preparation method based on air source purge provided in Example 1 of the present invention.

[0021] Figure 2 This is an overall schematic diagram of a bone marrow smear preparation system based on air source purge provided in Example 3 of the present invention. DETAILED DESCRIPTION

[0022] To make the above-mentioned objects, features, and advantages of the present invention more clearly understood, the following detailed description of the specific embodiments of the present invention is given in conjunction with the accompanying drawings. It is obvious that the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary persons in this field without creative work should fall within the scope of protection of the present invention.

[0023] Example 1, reference Figure 1 , as one embodiment of the present invention, provides a method for preparing bone marrow smears based on air source purge, comprising:

[0024] S1: Obtain and process bone marrow cell samples to form a cell suspension.

[0025] Furthermore, obtaining and processing the bone marrow cell sample includes centrifuging the collected bone marrow suspension to form a bottom sediment of the bone marrow cells in the centrifuge tube, removing the supernatant, adding physiological saline to the bottom sediment for resuspending to form a cell suspension, and blowing the cell suspension with a pipette.

[0026] It should be noted that the bone marrow suspension collected is placed in a centrifuge tube, centrifuged at a speed of 1000rpm to 2000rpm, and the centrifugation time is controlled between 5 minutes and 10 minutes so that bone marrow cells are fully settled to the bottom of the centrifuge tube, thereby forming a clearly visible cell precipitation layer. Use a pipette to gently draw and remove the clarified liquid (supernatant) on the upper strata of the centrifuge tube, avoid disturbing the bottom cell precipitation, and according to the precipitation volume, add a certain volume of normal saline or cell culture fluid to resuspend the cell precipitation. The diluent volume added is preferably 5 times to 10 times of the cell precipitation volume. For example, when the precipitation cell volume is 0.1mL, 0.5mL to 1mL of resuspension is added. By liquid dilution, a cell suspension suitable for cell concentration and stable in physical properties can be obtained.

[0027] It should also be noted that by centrifuging the bone marrow suspension sample at a set speed (1000rpm~2000rpm), the bone marrow cells are fully settled to the bottom of the centrifuge tube, and the supernatant is gently removed with a pipette to avoid disturbing the sediment structure, thereby achieving the standardization of the mechanical conditions for the initial separation operation of the bone marrow sample and the non-destructive extraction control of the sediment extraction process, ensuring the complete retention of the bottom sediment cells, and providing a basis for the complete cell morphology for subsequent resuspension processing. By adding physiological saline or cell culture fluid for dilution according to the ratio of the sediment volume (usually 5 to 10 times), and performing a preset number of pipetting operations with a pipette, the cells are fully and evenly mixed in the diluent and the concentration is adjusted accordingly. The above-mentioned resuspension process not only eliminates the aggregation and clumping phenomena between cells, but also avoids the difficulty of observation caused by cell damage or excessive dilution in a high concentration state. By adding physiological saline or cell culture fluid for dilution according to the ratio of the sediment volume (5 to 10 times), and performing a preset number of pipetting operations with a pipette, the cells are fully and evenly mixed in the diluent and the concentration is adjusted accordingly. The resuspension process not only eliminates the aggregation and clumping of cells, but also avoids cell damage at high concentrations or difficulty in observation caused by excessive dilution.

[0028] Furthermore, using a pipette to blow the cell suspension includes inserting a pipette tip equipped with a suction tip below the liquid surface of the cell suspension, performing suction and discharge operations in sequence, and fully mixing the cells and the liquid through a preset number of blows. The blown operation is performed until the cell suspension reaches a preset uniformity requirement, and the suction tip is avoided from contacting the container wall during the blown process.

[0029] It should be noted that the pipette tip equipped with a sterile tip is vertically inserted below the resuspension liquid level, go deep but do not touch the bottom of the tube, after drawing part of the cell mixture, then slowly discharged, so repeatedly draw and discharge operation, to realize the mixing and resuspension of cells in liquid. The action requirement of pipetting operation is stable, slowly, the preset number of times of pipetting is preferably 10 times to 20 times, avoid forming bubble or shearing force too large to cause cell rupture or death, in the pipetting process, special attention should be paid to controlling the tip position, avoid contacting with the centrifuge tube wall, to prevent cell from losing because of wall hanging, avoid the cell damage caused by electrostatic adhesion or shearing friction, a preferred embodiment of the preset uniformity requirement is, cell distribution state in diluent is without obvious deposition, agglomeration or floating layer separation phenomenon, and under microscope (low power objective lens), it can be observed that cells are uniformly suspended in liquid background, cell number fluctuation in unit volume (10 μ L) is controlled within ± 10%, and there is no visible aggregation agglomeration.

[0030] It should also be noted that the present invention forms a set of standardized cell resuspension and mixing technical solutions by vertically inserting the pipette tip below the liquid surface of the suspension and avoiding contact with the wall of the centrifuge tube, combining a preset 10 to 20 times of slow blowing operation, and setting a standard for judging the uniformity of cell mixing. This ensures that while ensuring the integrity of the cell structure, the cells are highly dispersed and have a stable concentration in the liquid, avoiding problems such as aggregation, sedimentation, and bubble interference, effectively improving the consistency and stability of the bone marrow cell sample preparation process, ensuring the uniformity of cell spreading during the subsequent gas source purge process, providing a homogeneous and physically damage-free sample basis for microscopic observation and staining operations, and improving the final smear quality and diagnostic reliability.

[0031] S2: Apply the cell suspension to the glass slide and perform direction-directed treatment of the cell suspension by air purge.

[0032] Furthermore, the gas source purge includes, after the cell suspension is dropped onto the slide, starting the gas source purge device and adjusting the gas output parameters according to the sample characteristics. By adjusting the gas output parameters, the airflow stability and coverage range are controlled to form a directional purge path. During the purge process, the gas output parameters are adjusted according to the spreading state of the cell suspension, and combined with the movement rhythm of the slide, the cell suspension is evenly spread along the surface to a predetermined area.

[0033] It should be noted that after the cell suspension is prepared, first use a pipette to absorb an appropriate amount of cell suspension (5-10 μL) and accurately drop it at the designated position at one end of a clean glass slide. The designated position is preferably at one end of the center in the length direction of the glass slide, so as to facilitate the subsequent propulsion of the airflow along the length of the glass slide and provide sufficient spreading distance. Start the gas source purge device and initialize the gas output parameters. The gas source can be oil-free compressed air or high-purity nitrogen to ensure that no droplet contamination or gas impurities are introduced. The initial gas pressure is set to 0.1-0.15 MPa. When the viscosity of the cell suspension is high, it can be appropriately increased to 0.15-0.2 MPa. If the cell state is relatively fragile, the pressure is reduced to 0.1-0.12 MPa to prevent the cell structure from rupturing. The gas flow rate is initially set to 1-2 L / min and adjusted in real time according to the droplet expansion speed and uniformity during the spreading process, with a maximum of no more than 3 L / min. The nozzle diameter is controlled within the range of 0.5 to 1 mm to form a needle-shaped directional airflow, which facilitates precise control of the airflow direction and concentration. The distance between the nozzle and the surface of the slide is adjusted to 1 to 1.5 cm, and the angle of inclination is maintained at 30° to 45°, so that the airflow forms a reasonable tangential incident angle with the surface of the slide, thereby driving the directional spreading of the droplet surface. After the air source parameters are set, the air source purge device is turned on to output the airflow from the nozzle, precisely aiming at the center of the droplet for directional purge operation. At the same time, the operator slowly moves the slide parallel to the glass slide at a speed of 1 to 2 cm / s, and controls the movement direction to be consistent with the airflow direction, so that the cell suspension gradually spreads on the slide surface along the propulsion path. During the purge process, the airflow direction should be kept stable and the flow rate should be uniform, and it should be adjusted synchronously with the slide movement rhythm to avoid coating interruption, blanks or accumulation due to excessive airflow or uneven movement. When the cell suspension is observed to form a stable, continuous and uniform liquid film structure on the surface of the glass slide, a preferred solution for the predetermined area is: the area length accounts for about 2 / 3 to 3 / 4 of the total length of the slide, and the width direction basically covers the effective observation area of ​​the slide. The preferred predetermined area can leave sufficient coating advancement distance to avoid insufficient blowing due to the front end being too close or the back end being too far causing dilution of cell concentration, matching the physical field of view of microscopic observation, facilitating continuous acquisition of cell images under low-power and medium-power objectives, controlling cell concentration and distribution density, maintaining uniform cell density in the preferred predetermined area, avoiding accumulation and blanks, and completing the gas source blowing operation when the cell suspension is evenly spread along the surface to the predetermined area.

[0034] It should also be noted that by precisely controlling the nozzle parameters and airflow stability, the cell suspension can smoothly transition along the preset path, avoiding undesirable phenomena such as faults, white spots, and liquid film tearing that occur during traditional blowing, thereby ensuring the integrity of the smear image quality and its diagnostic value. The gas parameters and operation rhythm are standardized and adjustable control items, which are suitable for cell samples of different concentrations and viscosities, broadening the scope of application of the system, supporting the integrated control of automated equipment, and clarifying the parameter specifications for nozzle layout, flow rate setting, and movement trajectory, reducing operational deviations caused by manual experience, which is particularly suitable for batch production and standardized application scenarios. By limiting the coating area and setting microscopic quality standards, it is ensured that the final smear has a continuous cell layer, appropriate density, and good staining state under the microscope, thereby improving image acquisition accuracy and analysis efficiency.

[0035] Furthermore, the adjustment of the gas output parameters includes, during the purge process, determining the spreading state of the cell suspension. If the spreading state is determined to be an aggregated state, the position and angle of the nozzle are adjusted to control the direction of the airflow to align with the aggregated area for directional purge.

[0036] If the spreading state is determined to be a local blank, the gas flow rate and pressure are adjusted to replenish the cell suspension in the local blank area.

[0037] It should be noted that during the purge process, if cells in a local area are clearly accumulated, forming a darker color or overlapping cell images, and the boundaries do not diffuse, it is determined to be in an aggregated state. If a transparent blank area visible to the naked eye appears on the slide surface or the liquid film is too thin and the coating is discontinuous, it is determined to be in a local blank state. When it is determined to be an aggregated state, the angle and position of the nozzle should be immediately adjusted to directly aim the airflow at the aggregated area, and the local enhanced airflow should be controlled to be slightly higher than the original setting to achieve local disaggregation. The angle adjustment range is fine-tuned within 30° to 45°, and the nozzle position can be moved forward 5-10mm to ensure that the airflow direction is precisely directed at the aggregation point. When it is determined to be in a local blank state, the gas flow rate and pressure should be reduced to prevent local over-purging from exacerbating cell rarefaction. At the same time, use a pipette to replenish the cell suspension in the blank area in an appropriate amount. The amount of replenishment should be limited to prevent the formation of new droplet accumulation. After replenishment, re-purge the area to restore the continuity of the liquid film and cell uniformity.

[0038] It should also be noted that through local enhanced purging or fluid replenishment correction, common accumulation areas and blank areas can be effectively eliminated, cells can be evenly spread on the entire glass slide surface, and the spatial continuity of the stained image can be improved. Compared with the traditional one-time purging method, the present invention allows fine-tuning and correction operations to be implemented during the preparation process, reducing the failure rate caused by sample differences or operational errors. By precisely controlling the purging area and gas intensity, the risk of cell rupture caused by excessive purging or high pressure is reduced, while avoiding sample consumption caused by overall re-spreading, thereby improving preparation efficiency and sample utilization.

[0039] S3: Fixing and staining the bone marrow smear formed by blowing through the gas source to complete the bone marrow smear preparation operation.

[0040] Furthermore, the bone marrow smear fixation includes placing the bone marrow smear in a fixation environment, applying a fixative to the surface of the bone marrow smear, and removing the residual fixative after a preset fixation time.

[0041] It should be noted that the bone marrow smear that has completed the purging operation is placed in a fixed environment at room temperature, and a fixative is applied to the surface of the bone marrow smear to keep the morphological structure of the cells on the slide stable. The fixative is preferably methanol, which has good permeability and cell structure preservation ability, can effectively penetrate the cell membrane and promote rapid cell fixation. Use a dropper or pipette to draw an appropriate amount of methanol and evenly add the methanol to the surface of the bone marrow smear. The coverage should be sufficient to form a complete liquid film structure to ensure that the entire smear area is evenly affected by the fixative. The volume of methanol required for each smear is generally 3 to 5 mL. After adding the fixative, the smear is kept on a fixed stand and allowed to react and fix for 3 to 5 minutes at room temperature. If the fixation time is too short, the cell structure will fall off or the morphology will be unstable during the staining process. If the fixation time is too long, the cell structure will degenerate or the staining effect will decrease.

[0042] Furthermore, the bone marrow smear staining includes applying a staining solution to the surface of the bone marrow smear after fixation so that the staining solution covers the bone marrow smear area, maintaining the primary staining time, applying a buffer solution to the surface of the bone marrow smear, maintaining the secondary staining time, rinsing the bone marrow smear, and drying the rinsed bone marrow smear.

[0043] It should be noted that after the bone marrow smear is naturally dried after fixation, the smear is placed horizontally on the staining rack. The staining solution is preferably Wright's stain. Use a dropper to absorb the pre-diluted Wright's stain and add it dropwise to the smear surface so that the stain completely covers the smear area without forming flow or droplet accumulation. The amount of Wright's stain required for each smear is generally 3 to 5 mL. The dye dilution ratio is 1:10 to 1:15 (volume ratio), and can be flexibly adjusted according to the cell density of the bone marrow smear to achieve the best staining effect. After maintaining the primary staining state for 3 to 5 minutes, continue with the secondary staining operation, that is, add an appropriate amount of buffer to the smear surface. The buffer is preferably phosphate buffer (pH between 6.4 and 6.8), and the amount is 1 to 2 times the volume of the dye, and gently shake the slide to fully mix the dye and buffer. Continue to maintain the staining reaction for 5 to 10 minutes, so that the various cell components on the smear can bind to the alkaline and acidic dyes according to the difference in affinity, which is manifested as: the cell nucleus is stained purple-red, and the cytoplasm is stained pink to light blue. After the staining is completed, the smear is rinsed with tap water or distilled water. The rinsing method should be a slow downward flow of water to avoid direct impact on the surface of the smear and causing cell shedding. The rinsing process lasts for 1 to 2 minutes, until the clear water flowing down has no obvious staining or is slightly light blue. Place the rinsed smear on absorbent paper to dry naturally in the shade, avoiding high temperature or direct sunlight to prevent cell deformation or staining fading, and ensure that the cell morphology remains clear and the staining is accurately distinguished, providing a reliable basis for subsequent microscopic observation and morphological analysis.

[0044] Example 2 is an embodiment of the present invention, which provides a bone marrow smear preparation method based on air source purge. In order to verify the beneficial effects of the present invention, scientific demonstration is carried out through economic benefit calculation and simulation experiments.

[0045] First, an experimental group (bone marrow smears were prepared using air source purging technology) and a control group (bone marrow smears were prepared using traditional push-slide technology) were set up. Fifty bone marrow smears were prepared in each group, and the experimental subjects were bone marrow samples from healthy rats.

[0046] Experimental instruments and materials:

[0047] Instruments: air purge device (with precise pressure adjustment), slides required for traditional slide pushing, micropipette, optical microscope, cell counting chamber.

[0048] Materials: Rat bone marrow cell suspension, methanol fixative, Wright's stain, phosphate buffer.

[0049] Experimental process:

[0050] Bone marrow smears were prepared using air purge technology and traditional push-slide technology, and the preparation time of each smear was recorded.

[0051] The prepared smears were fixed and stained, and the operation steps and parameters were kept consistent.

[0052] Under an optical microscope, 10 high-power fields were randomly selected from each smear to observe the cell distribution and cell morphology, and to count the cells.

[0053] Experimental results:

[0054] Homogeneity: Use a cell counting chamber to determine the initial cell concentration of the cell suspension in each smear, which is recorded as C0.

[0055] Under a microscope, count the number of cells in each high-power field (C). i i=1,2,3,4,5,6,7,8,9,10, the coefficient of variation CV of cell distribution is calculated according to the formula:

[0056]

[0057] Where CV represents the coefficient of variation, which is used to indicate the degree of discreteness of the number of cells per unit area, n represents the number of sampling units, that is, the total number of cell statistical sub-areas divided within the predetermined observation area, and C i represents the number of cells in the ith sub-region, represents the arithmetic mean of the number of cells in all sub-regions, and the coefficient of variation of cell distribution is shown in Table 1.

[0058] Table 1 Cell distribution coefficient of variation

[0059]

[0060] Normalize data:

[0061] The qualified rates of smears prepared by different operators in the experimental and control groups were statistically analyzed. The qualified criteria were uniform cell distribution, complete morphology, and good staining effect. The qualified rate was calculated as follows:

[0062]

[0063] The qualified rates of smears are shown in Table 2.

[0064] Table 2 Smear qualified rate

[0065]

[0066] Efficiency improvement data:

[0067] The total time for preparing 50 smears in each group was recorded, and the average preparation time for a single smear was calculated. The efficiency improvement times of the two groups were compared. The smear time is shown in Table 3.

[0068] Table 3 Smear time

[0069]

[0070] Diagnostic Accuracy Data:

[0071] Calculate the percentage of effective cells in the microscope field of view. Effective cells refer to cells with complete morphology and clearly observable structures. The percentage of effective cells is expressed as:

[0072]

[0073] The two groups of smears were observed and diagnosed blindly, and the cell missed detection rate and misjudgment rate were calculated, as shown in Table 4.

[0074] Table 4 Smear effective cell percentage, cell missed detection rate, and misjudgment rate

[0075] Technology Type Average effective cell percentage Cell missed detection rate False positive rate Air source purge technology 86.70％ 11.80％ 7.90％ Traditional film pushing technology 64.30％ 27.60％ 17.80％

[0076] Example 3, reference Figure 2 , which is an embodiment of the present invention, provides a bone marrow smear preparation system based on air source purge, including a sample processing module 100, an air source purge module 200, and a fixation and staining module 300.

[0077] Among them, S4: the sample processing module 100 includes a cell centrifugation and resuspension unit 101 and a cell mixing control unit 102. The cell centrifugation and resuspension unit 101 is used to centrifuge the collected bone marrow suspension and remove the supernatant, and then add liquid to resuspend the cells to form a processable initial cell suspension. The cell mixing control unit 102 is used to use a pipette to blow the cell suspension to evenly disperse the cells and adjust it to a suspension state suitable for smearing.

[0078] It should also be noted that the cell centrifugation and resuspension unit 101 outputs processed cell suspension data, which is passed as input to the cell mixing control unit 102 to perform blowing and mixing processing in subsequent links. The cell suspension state parameters after processing by the cell mixing control unit 102 are passed to the gas parameter control unit 201 as a basis for adjusting the initial purge parameters.

[0079] S5: The gas source purge module 200 includes a gas parameter control unit 201 and a spreading state feedback control unit 202. The gas parameter control unit 201 is used to set and adjust the gas parameters to generate a directional airflow suitable for cell spreading. The spreading state feedback control unit 202 is used to determine whether there are aggregations or blank areas, and automatically adjust the gas parameters or control the fluid replenishment behavior to achieve uniform spreading.

[0080] It should also be noted that the output control parameters such as air pressure, flow rate, nozzle angle, etc. set by the gas parameter control unit 201 are sent to the nozzle actuator, and feedback data from the spreading state feedback adjustment unit 202 is received in real time, including the location of the gathering area or the range information of the blank area, for dynamic correction of the gas output control instructions.

[0081] S6: The fixation and staining module 300 includes a fixative application control 301 and a staining process management unit 302. The fixative application control 301 is used to control the uniform application and timing of the fixative on the smear surface to complete the fixation process of the cell structure. The staining process management unit 302 is used to sequentially control the application of the staining solution and buffer solution, as well as the staining time, rinsing and drying process to complete the staining process of the smear.

[0082] It should also be noted that the fixation application control unit 301 outputs a fixation completion signal and slide status information to the staining process management unit 302. This information includes staining liquid volume estimation parameters and fixation time tags for driving the liquid control sequence of the staining stage.

Claims

1. A bone marrow smear preparation method based on air source purge, characterized in that: include: Obtain and process bone marrow cell samples to form cell suspensions; The cell suspension is applied to a glass slide and directional treated by air purge; The bone marrow smear formed by the air source is fixed and stained to complete the bone marrow smear preparation operation; The gas source purge includes starting the gas source purge device after the cell suspension is dropped onto the glass slide, adjusting the gas output parameters to form a directional airflow path, so that the cell suspension spreads on the surface of the glass slide, and dynamically adjusting the gas parameters according to the spreading state of the cell suspension during the purge process, so that the cell suspension spreads evenly to a preset area; The bone marrow smear fixation and bone marrow smear staining include placing the bone marrow smear in a fixed environment after forming a bone marrow smear by blowing an air source, applying a fixative to the surface of the bone marrow smear and maintaining it for a preset fixation time, applying a staining solution to the surface of the fixed bone marrow smear and maintaining it for a primary staining time, applying a buffer solution and maintaining it for a secondary staining time, and then rinsing and drying the stained bone marrow smear.

2. The method for preparing bone marrow smears based on air source purge according to claim 1, wherein: The method of obtaining and processing the bone marrow cell sample includes centrifuging the collected bone marrow suspension to form a bottom sediment of the bone marrow cells in the centrifuge tube, removing the supernatant, adding physiological saline to the bottom sediment for resuspending to form a cell suspension, and pipetting the cell suspension with a pipette.

3. The method for preparing bone marrow smears based on air source purge according to claim 2, wherein: The method of blowing the cell suspension with a pipette includes inserting a pipette tip equipped with a pipette tip below the liquid surface of the cell suspension, performing suction and discharge operations in sequence, and fully mixing the cells and the liquid by blowing a preset number of times. The blowing operation is performed until the cell suspension reaches a preset uniformity requirement, and the pipette tip is prevented from contacting the container wall during the blowing process.

4. The method for preparing bone marrow smears based on air source purge according to claim 3, wherein: The gas source purge includes starting the gas source purge device after dropping the cell suspension onto the slide, and adjusting the gas output parameters according to the sample characteristics. By adjusting the gas output parameters, the airflow stability and coverage range are controlled to form a directional purge path. During the purge process, the gas output parameters are adjusted according to the spreading state of the cell suspension, and combined with the movement rhythm of the slide, the cell suspension is evenly spread along the surface to a predetermined area.

5. The method for preparing bone marrow smears based on air source purge according to claim 4, characterized in that: The adjusting of the gas output parameters includes, during the purge process, determining the spreading state of the cell suspension, and if the spreading state is determined to be an aggregated state, adjusting the position and angle of the nozzle to control the direction of the airflow to align with the aggregated area for directional purge; If the spreading state is determined to be a local blank, the gas flow rate and pressure are adjusted to replenish the cell suspension in the local blank area.

6. The method for preparing bone marrow smears based on air source purge according to claim 5, characterized in that: The bone marrow smear fixation includes placing the bone marrow smear in a fixation environment, applying a fixative to the surface of the bone marrow smear, and removing the residual fixative after a preset fixation time.

7. The method for preparing bone marrow smears based on air source purge according to claim 6, characterized in that: The bone marrow smear staining includes applying a staining solution to the surface of the bone marrow smear after fixation so that the staining solution covers the bone marrow smear area, maintaining a primary staining time, applying a buffer solution to the surface of the bone marrow smear, maintaining a secondary staining time, rinsing the bone marrow smear, and drying the rinsed bone marrow smear.

8. A bone marrow smear preparation system based on air source purge, using the bone marrow smear preparation method based on air source purge according to any one of claims 1 to 7, characterized in that: It includes a sample processing module (100), an air source purge module (200), and a fixation and staining module (300); The sample processing module (100) includes a cell centrifugation and resuspension unit (101) and a cell mixing control unit (102). The cell centrifugation and resuspension unit (101) is used to centrifuge the collected bone marrow suspension and remove the supernatant, and then add liquid to resuspend the cells to form a processable initial cell suspension. The cell mixing control unit (102) is used to use a pipette to blow the cell suspension to evenly disperse the cells and adjust it to a suspension state suitable for smearing. The gas source purge module (200) comprises a gas parameter control unit (201) and a spreading state feedback control unit (202), wherein the gas parameter control unit (201) is used to set and adjust gas parameters to generate a directional airflow suitable for cell spreading, and the spreading state feedback control unit (202) is used to determine whether there are aggregated or blank areas, and automatically adjust gas parameters or control fluid replenishment behavior to achieve uniform spreading; The fixing and staining module (300) includes a fixing solution application control (301) and a staining process management unit (302). The fixing solution application control (301) is used to control the uniform application and timing of the fixing solution on the smear surface to complete the cell structure fixation process. The staining process management unit (302) is used to sequentially control the application of the staining solution and the buffer solution, as well as the staining time, rinsing and drying processes to complete the staining process of the smear.

9. A computer device comprising a memory and a processor, wherein the memory stores a computer program, wherein: When the processor executes the computer program, the steps of the bone marrow smear preparation method based on gas source blowing according to any one of claims 1 to 7 are implemented.

10. A computer-readable storage medium having a computer program stored thereon, characterized in that: When the computer program is executed by a processor, the steps of the bone marrow smear preparation method based on gas source purging according to any one of claims 1 to 7 are implemented.

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