JPH0143257B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a particle detection device for detecting particles such as blood cells suspended in a liquid. It is an object of the present invention to provide a particle detection device that can effectively detect particles such as particles, especially smaller particles such as platelets.

Conventionally, when detecting and counting microparticles such as red blood cells and platelets, a suspension of particles is passed through micropores provided below the liquid level, and detection is performed based on the difference in electrical impedance between the liquid and the particles. This is converted into an electrical pulse signal and counted. Normally, when detecting and counting red blood cells and platelets, micropores from several tens of microns to about 100 microns are used, and the blood is diluted in advance to a concentration that allows each blood cell to pass through the micropores before counting and measuring. It's summery. When counting only red blood cells or separating platelets and counting only platelets, counting can be performed relatively easily by simply removing the noise signal in the detection device, but for platelets, red blood cells This method requires separation from large particles, and this separation requires conversion to the number of platelets in the original blood, which has disadvantages such as being relatively troublesome and time-consuming, and with large errors. Ta.

In addition, in the whole blood method, which detects and counts blood by simply diluting it, the number of red blood cells per ^mm3 is around 5 million, and the number of platelets is around 200,000, which is 25 times the number of platelets. In addition to having to measure and subtract at the same time, there are factors that have a significant impact on platelet counts. This is a phenomenon in which the blood cells that have passed through the micropores are entangled in the detection area again and are detected as if they had passed through the micropores, causing an error in the counted value. This phenomenon mainly involves red blood cells that have already passed through the micropores, producing a pulse with the same height as the platelet signal. For this reason, even if a so-called window-type threshold circuit removes the large pulses caused by red blood cells and allows only platelet-sized signals to pass through, the pulses caused by the involvement of red blood cells will be added, making it difficult to measure by separating only platelets. The drawback was that the measured value was much higher than the platelet count converted to blood.

Methods to overcome this drawback include (1) narrowing the detection area to prevent entangled particles from being detected, and (2) preventing particles that have passed through the micropores from coming close to the micropores again. There is a method. For method (1), the electric field distribution for detection is limited to a very limited area by arranging electrodes near the micropores, and for example, electrodes are provided near the micropores by vapor deposition or plating. In reality, metals such as platinum, which have excellent corrosion resistance, are used as electrodes, and even if these are applied to the surface of the detector by vapor deposition or plating, they easily peel off or are susceptible to corrosion due to electrolysis. It is not practical due to corrosion. Incidentally, platinum is chemically stable but has low mechanical strength. In addition, as a method for (2), it is possible to provide a means to suck all the particle suspension after passing through the micropores, or to blow away the particle suspension with a liquid that does not contain particles. Because the structure is complex and it becomes impossible to quantify the particle suspension that has passed through the micropores, it is not possible to simply connect a liquid quantification device inside the detector as in the past. However, this method has the drawback of requiring a complicated method with many error factors, such as converting the number of particles passing per unit time to the number of particles per unit volume.

The present invention was developed in view of the above points, and has a structure that is simple and inexpensive to manufacture, prevents particles such as red blood cells from returning, and allows for easy correction. The object of the present invention is to provide a particle detection device that can easily detect, classify, and count particles.

The particle detection device of the present invention has a pellet 1 with a fine hole 2 formed in the lower part thereof, and a spherical substance 5, a fibrous substance 24,
A detector filled with a buffer material made of a porous material 25 or a combination thereof to form a particle return section, and a sample chamber 6 provided so that the lower part of the detector is immersed in the particle suspension 7. , a liquid quantification device 17 connected to the upper part of the detector to quantify the particle suspension sucked through the fine hole 2, a suction pressure source 20 connected to this liquid quantification device, Particle detection electrode 2 arranged in
1 and 22.

Hereinafter, the configuration of the present invention will be explained based on embodiments shown in the drawings. FIGS. 1 and 2 show an example of a detector used in the particle detection device of the present invention. 1 is a pellet 1 having micropores 2 with a diameter of about 50 to 100 μm, and this pellet 1 is made of ruby or sapphire, etc., and is formed into a disk shape with a thickness of about 10 to 100 μm, and is made of glass, synthetic resin, or ceramic, and has a lower end. is fitted and fixed to the lower side or bottom of the sealed cylindrical detector main body 3. 4
is a large part for fixing provided on the upper part of the detector main body 3. In addition, from the bottom of the inside of the detector body 3 to 5 to 15 mm above the fine hole 2, a spherical substance 5 or approximately spherical substance such as glass beads, stainless steel balls, or synthetic resin balls with a diameter of about 1 mm or less is filled. has been done. Therefore, from the micropore 2 to the upper surface of the spherical material 5, a large number of intricate passages of several tens to hundreds of micrometers are formed, and the sample after passing through the micropore 2 is sequentially transported through these passages. The ejected flow after passing through the micropores moves as a gentle flow, exerting a buffering effect similar to a tetrapod for dissipating waves on the coastline, and as a result, the return of particles and entrainment phenomena are extremely will be relaxed. This is also clear from the data shown in FIG. 4 obtained by the particle detection device shown in FIG. 3, which will be described later.

Next, one embodiment of a particle detection device using a detector configured as described above will be described based on FIG. 3. Reference numeral 6 denotes a sample chamber for containing a particle suspension, for example, a suspension of blood cells, and a suspension of blood cells 7 is introduced into the sample chamber 6 through a sample inlet 8 and discharged through a sample outlet 10. . The detector body 3 is fixed such that its lower part is immersed in the suspension 7.
The upper part of the detector body 3 has two liquid reservoir spaces 12 and 13 (the lower space is 12 and the upper space is 13) vertically adjacent to each other via a flexible diaphragm 11. Upper part of the main body 14
The inside of the detector main body 3 is fixed so as to communicate with the space 12 below. This lower space 12 is connected to a drainage reservoir and a suction pressure source 16 via a discharge pipe 15 having a solenoid valve, etc., and is configured to discharge unnecessary samples inside the detector and the fluid reservoir space 12. There is. On the other hand, the upper space 13
is connected to a liquid metering device 17, and this liquid metering device 17 is further connected to a suction pressure source 20 via a control valve 18 for controlling air pressure and the like. For example, the liquid metering device 17 is of a type that optically detects the movement of the liquid level and measures a predetermined volume. The diaphragm 11 isolates the liquid metering device 17 from the interior of the detector, and has the effect of preventing sample from entering the liquid metering device 17 and causing contamination. Also, the liquid reservoir space 12
An internal electrode 21 is disposed in the suspension in the sample chamber 6, and an external electrode 22 is disposed in the suspension in the sample chamber 6.These electrodes 21 and 22 are connected to a detection circuit 23 to separate the particles passing through the micropores 2 from the liquid. It is configured to detect based on the difference in electrical impedance between the two. Note that atmospheric pressure can also be introduced into the liquid metering device 17 by switching the control valve 18.

In the particle detection device configured as described above, a sample (particle suspension) is introduced into the sample chamber 6 from the sample introduction port 8, and the liquid level is adjusted to the fine hole 2 of the detector body 3.
Fill until it reaches above. At this time, the solenoid valve of the discharge pipe 15 is closed and in a cutoff state, and the control valve 18 is opened and suction pressure is introduced into the liquid metering device 17. At the same time, the liquid level in the liquid metering device 17 rises and the diaphragm 11 is curved upward, and the sample is introduced into the detector through the fine hole 2. At this time, an electrical change occurs between the internal electrode 21 and the external electrode 22 and is detected as a particle signal. The liquid level of the liquid metering device 17 is detected, a start/stop signal is sent to the counting device, and the number of pulses counted during this period is counted and displayed as the number of particles per unit volume. When the counting is completed, the control valve 18 is switched, atmospheric pressure is introduced into the liquid metering device 17, and the diaphragm 11 returns to its original position. On the other hand, suction pressure is introduced into the discharge pipe 15, and the sample is discharged into the liquid reservoir space 12 through the discharge pipe 15.

The particle signals detected by the electrodes 21 and 22 are
A window-type threshold circuit with two levels divides the pulse into three regions depending on the height of the pulse. Signals below the lower limit level are removed as unnecessary signals such as noise, and pulse signals above the upper limit level are removed as large signals such as red blood cells. Particle signals and intermediate level signals between the upper and lower limits are classified as signals of small particles such as platelets, and each is counted. In order to detect pulses that exceed the upper limit even at intermediate levels using a window type comparator and prevent them from being counted, a circuit configuration such as a one-shot multivibrator, delay circuit, etc. is used to generate a prohibition signal. , as an example of the circuit.

In a particle counting device using a particle detection device configured as described above, in order to investigate the influence of red blood cell entrainment, measurements were performed under the following conditions while varying the particle size of the spherical material 5 in the detector body 3. Ta. The results were as shown in Figure 4.

As the spherical substance 5, glass beads with a diameter of 4 mm,
Experiments were conducted using glass beads with a diameter of 3 mm, glass beads with a diameter of 2 mm, glass beads with a diameter of 1 mm, and cases where no glass beads were used at all. As a sample, the red blood cell count measurement value is 1500.
A sample about three times darker than a normal sample was used so as to show a count of 10,000 pieces/mm ³ . Further, the counting time was repeated for a total of 15 seconds, including a set time of about 1 second, a counting time of about 12 seconds, and a reset time of about 2 seconds. The diameter of the micropores was 80 μm, and the suction pressure of the suction pressure source connected to the liquid metering device was approximately 130 mmHg/cm ² . The measurement method was to fill each of the above-mentioned glass beads to a depth of about 1 cm or more above the micropores, and first aspirate a diluted blood sample into the detector. Subsequently, a blank solution containing no blood (a solution containing only a diluent and no blood cells suspended) was repeatedly drawn into the detector, and counting measurements were performed at platelet sensitivity (threshold level). That is, at the 0th measurement, a normal counting measurement was carried out, at the 1st measurement, a counting measurement of the blank liquid was carried out, and at the 10th measurement, the counting measurement of the blank liquid was carried out.

By doing the above, the red blood cells sucked into the detector during the 0th measurement are
From the first measurement, it can be seen that the blank liquid is diluted one after another and gradually stops being rolled up. In other words, in Figure 4, the number of red blood cells that are counted as platelets due to return (entrainment) (at this time, only liquid passes through the micropores and particles do not pass through) is the number of red blood cells counted without glass beads or with a diameter of 4 mm in the first round. 85 for glass beads
The 10,000-odd particles gradually become diluted with repeated counting, and after 10 measurements, 1 to 1000
The number has decreased to 20,000. This tendency increases as the diameter of the glass beads decreases, and
The ejected flow from the micropores inside the detector, which causes the entrainment phenomenon, is relaxed, and there is no return.
In the case of glass beads with a diameter of 1 mm, the result corresponding to the 10th test without using glass beads is
It was found that this phenomenon occurred noticeably from the first measurement and exhibited a very large effect. By setting the diameter of the glass beads to about 1 mm in this way, the sample that has passed through the micropore does not stagnate on the back side of the micropore, and while preventing the phenomenon of rebound or entrainment, It became clear that the particles were quickly caught in the gaps between the glass beads and transported. In addition, the red blood cells in the previous sample and the sample before the previous sample that have already been aspirated will not affect the platelet count in each batch.
However, the influence of the red blood cells in the sample being aspirated this time (equivalent to the 0th time) cannot be determined in the above experiment; for example, the red blood cells immediately after passing through the micropores may make a U-turn and become entangled right behind the micropores. In some cases, pulses equivalent to platelets are generated, giving an error to the platelet count. However, this value is much smaller than the conventional one.

The above phenomenon can be expressed mathematically as follows.

PL (number of platelets) = PL ₀ (count value equivalent to platelets) - {a
₀ + a ₁ × RBC ₁ (current red blood cell count) PL (platelet count) = PL ₀ (count value equivalent to platelets) − {a
₀ +a ₁ ×RBC ₁ (current red blood cell count) +a ₂ ×RBC ₂ (previous red blood cell count) +…+a _o ×
RBC _o (number of red blood cells before n-1 times)} Note that a ₀ , a ₁ , a _{2 .} . . a _o are constants.

As is clear from Figure 4, by filling the bottom of the detector with glass beads, the value of the constant was required to be up to n = 11, and at the same time the value of the constant became smaller, and at the same time, the correction was made. If the number of items to be performed can be reduced and the diameter of the glass beads is about 1 mm, it is no longer necessary to correct the number of red blood cells and a predetermined constant, and the above equation can be simplified as shown below.

PL = PL ₀ - (a ₀ + a ₁ × RBC ₁ ) As a result of measuring a large number of samples at a dilution concentration 50,000 times the normal concentration, the correlation with the conventional method was good because a ₀ =
8.5, and a ₁ =0.027. In addition, PL,
The units of PL ₀ , a ₀ , and RBC ₁ are all 10,000 pieces/mm ³ . The values of a ₀ and a ₁ must be changed somewhat depending on the diameter of the fine pores of the detector and the shape of the detector body, and furthermore, it is necessary to correct the simultaneous passage between platelets and red blood cells. That is, as the respective concentrations of platelets and red blood cells increase, platelets and platelets and platelets and red blood cells pass through the micropores almost simultaneously and are counted as one particle, so it is necessary to correct the measured values.

In the above embodiment, the spherical material 5 is used as a buffer material to suppress the jet of the aspirated sample, but instead of the spherical material, as shown in FIG. 5, a passage with a diameter of several tens of microns is used. A large number of filter-like fibrous substances 24 can be used. Suitable fibrous materials include cellulose, glass fiber, fluorine fiber, carbon fiber, and other synthetic resin fibers.

Further, as shown in FIG. 6, a porous material 25 made of relatively coarse sponge or foamed synthetic resin can be used.

Furthermore, as shown in FIG. 7, a spherical substance 5 with a diameter of 1 mm or less is filled in the lower part of the detector main body 3, and the upper side of this spherical substance 5 is covered with the fibrous substance 24 and porous substance 25. Almost good results were obtained. In addition, when the detectors shown in FIGS. 5, 6, and 7 were used, almost the same results regarding constants were obtained as when the detector shown in FIG. 1 was used. In addition, if the position where these cushioning materials are filled is lower than about 3 mm above the position of the micro-hole, a rebound phenomenon will occur, but if the position where the buffer material is filled is higher than 3 mm from the micro-hole, that is, 5 mm to 10 mm, or if it is safe. It is preferable to set it to approximately 15 mm.

As explained above, the particle detection device of the present invention prevents particles such as red blood cells from returning, easily performs correction, has a simple structure, can be manufactured at low cost, and detects particles such as red blood cells and platelets. This has the effect that two types of particles, large and small, can be easily detected and classified and counted.

[Brief explanation of drawings]

Fig. 1 is a front view showing an example of a detector used in the particle detection device of the present invention, and Fig. 2 is an A in Fig. 1.
3 is an explanatory diagram showing one embodiment of the particle detection device of the present invention, and FIG. 4 is a cross-sectional view taken along the line A, and FIG. Graphs showing experimental results when not used, and FIGS. 5 to 7 are explanatory cross-sectional views showing other examples of detectors using the particle detection device of the present invention. 1... Pellet, 2... Micropore, 3... Detector body, 4... Ampulla, 5... Spherical substance, 6... Sample chamber, 7... Suspension, 8... Sample introduction port, 10...
...Sample outlet, 11...Diaphragm, 12,1
3... Space, 14... Upper part of the detector body, 15...
Discharge pipe, 16... Suction pressure source, 17... Liquid quantitative device, 18... Control valve, 20... Suction pressure source, 21... Internal electrode, 22... External electrode, 23
...detection circuit, 24 ... fibrous material, 25 ... porous material.

Claims (1)

[Claims] 1. It has a pellet with micropores in its lower part, and the interior is filled with a buffer material made of spherical material, fibrous material, porous material, or a combination thereof up to 5 to 15 mm above the micropores to prevent particles from returning. a sample chamber in which the lower part of the detector is immersed in the particle suspension; and a sample chamber connected to the upper part of the detector for quantifying the particle suspension aspirated through the micropores. A particle detection device comprising: a liquid quantification device that performs the following: a suction pressure source connected to the liquid quantification device; and a particle detection electrode disposed within a detector and a sample chamber. .