WO2024259548A1 - Biological aerosol monitoring apparatus and monitoring method - Google Patents

Abstract

A biological aerosol monitoring apparatus, comprising a light path, a gas path and a signal processing system, wherein the light path comprises a laser irradiation light path configured to irradiate particles to be tested, a scattered light collection light path configured to receive scattered light signals, and a fluorescence collection light path configured to receive fluorescence signals. A focal point (M) of an excitation light source (101) configured to irradiate said particles is located before or behind a photosensitive area (G) on an optical axis of the laser irradiation light path, such that the cross section of an excitation light spot in the photosensitive area (G) that is perpendicular to the optical axis of the laser irradiation light path is a rectangular shape, and when a single aerosol particle passes through the photosensitive area (G), the single aerosol particle is continuously irradiated by excitation light to obtain an intrinsic fluorescence bleaching speed. Information such as an intrinsic fluorescence bleaching speed, a scattered light peak value and a relative fluorescence intensity of aerosol particles is integrated, such that preliminary classification of biological particles is realized, instrument alerts caused by non-target microorganisms are significantly reduced, and the capability of an instrument for identifying the types of biological aerosols is improved, thereby reducing the frequency and cost of subsequent biological detection.

Description

Bioaerosol monitoring device and monitoring method Technical Field

The invention relates to real-time monitoring of bioaerosol, in particular to a bioaerosol monitoring device and a monitoring method.

Background Art

Real-time monitoring of bioaerosols has important application value. Excitation light induced intrinsic fluorescence detection technology has many advantages in the field of real-time monitoring of bioaerosols, such as fast speed, high sensitivity, no consumables, and non-invasiveness. Its technical principle is that bioaerosol particles contain organic molecules such as tryptophan, reduced coenzyme I (ie NADH) and riboflavin. These components can produce intrinsic fluorescence under the induction of ultraviolet light, while non-biological aerosol particles are generally difficult to produce intrinsic fluorescence. Based on this, it can be distinguished whether aerosol particles have biological properties.

However, the existing technologies currently have the following major problems: First, cigarette particles, kaolin, and dust containing polycyclic aromatic hydrocarbons in airborne particles can also generate intrinsic fluorescence signals. Most existing technologies are unable to distinguish these interferents, which may cause false detection or false alarms of the instrument; Second, there are many types of bioaerosol particles. In practical applications, it is often necessary to monitor only a few or a dozen biological targets, while most existing technologies can only monitor the total number of fluorescent particles in general and cannot classify aerosol particles. For example, pollen or some harmless microbial species do not need to be monitored. Therefore, preliminary screening of biological particles is required to reduce the frequency and cost of subsequent biological detection.

Patent document CN108375530A discloses a real-time bioaerosol detection method and detection device, including an optical path, an air path intersecting with the optical path, and a signal processing system connected to the optical path. The optical path includes a laser emission optical path for irradiating the measured particles, a scattered light collection optical path for receiving scattered light signals, and a fluorescence collection optical path for receiving fluorescence signals. The air path is used to sample the measured particles, and the minimum detection resolution is one microbial particle. The optical signal processing system is used to analyze and process signals, including a scattered light preamplifier and a fluorescence preamplifier, which can simultaneously monitor the concentration of microbial particles and non-microbial particles in the air. By identifying and distinguishing microbial particles and non-microbial particles, real-time monitoring of the concentration and quantity of microbial particles in the air is achieved and the monitoring accuracy is improved. Patent document CN103940709 discloses a real-time microbial particle counter, which determines the particle size and biological properties of the measured particles by detecting the scattered light and fluorescence intensity emitted by a single particle under the irradiation of excitation light. However, the above two patent documents can only detect the peak value of scattered light pulse and fluorescence pulse, the microbial determination index is single, and the fluorescence bleaching information cannot be tested. Many interference substances are detected, and the false alarm rate of the instrument is high.

Patent document CN110411995A discloses a bioaerosol monitoring device and method based on intrinsic fluorescence bleaching characteristics, which detects fluorescence bleaching information and distinguishes the types of aerosol particles according to the fluorescence bleaching characteristics. However, the instrument first enriches and samples multiple aerosol particles together, and then irradiates the collected particle group for a long time to obtain the fluorescence bleaching characteristics. The disadvantages are: ① It measures the cumulative fluorescence of multiple particles, and cannot obtain the scattered light, intrinsic fluorescence and fluorescence bleaching characteristics of a single aerosol particle. For samples containing multiple aerosol particles, it is impossible to accurately determine the test results; ② It needs to enrich a certain number of aerosol particles before testing, the detection sensitivity is low, and it is not real-time monitoring, and the result determination has a certain lag.

Summary of the invention

In view of the above existing problems, the present invention provides a bioaerosol monitoring device and a monitoring method, which utilizes that the intrinsic fluorescence bleaching rates generated by organic molecules in bioaerosol particles are different under the irradiation of the same excitation light, such as riboflavin and other components bleaching faster, while NADH and other components bleach slower or almost no bleaching. By analyzing the area, amplitude and other information of the intrinsic fluorescence pulse signal generated by aerosol particles passing through a light-sensitive area under continuous irradiation, and combining the information such as the elastic scattering light pulse signal of the particles, characteristic information of different types of aerosol particles can be collected, providing a basis for the preliminary classification of aerosol particles.

The technical solution of the present invention is as follows:

On one hand, the present invention provides a bioaerosol monitoring device, comprising an optical path, an air path intersecting with the optical path, and a signal processing system connected to the optical path; the optical path comprises a laser irradiation optical path for irradiating measured particles, a scattered light collection optical path for receiving scattered light signals, and a fluorescence collection optical path for receiving fluorescence signals; the air path is used to sample measured particles; the signal processing system is used to analyze and process signals; the intersection area of the laser irradiation optical path and the air path is a light sensitive area; the device is characterized in that the focus of the excitation light source for irradiating measured particles is located before or after the light sensitive area on the optical axis of the laser irradiation optical path, so that the cross section of the excitation light spot perpendicular to the optical axis of the laser irradiation optical path in the light sensitive area is rectangular, and when a single aerosol particle passes through the light sensitive area, it is continuously irradiated by the excitation light to obtain an intrinsic fluorescence bleaching rate.

Preferably, the laser irradiation optical path for irradiating the measured particles includes an excitation light source, the light emitted by the excitation light source is parallel to the X-axis direction and collimated by an aspherical mirror to form a parallel light beam, and then focused by a cylindrical mirror and passes through the light sensitive area to reach the light trap. In the Y-axis direction of the light sensitive area, there are a curved reflector and a dichroic mirror in sequence, and the dichroic mirror is set at 45° to the Y-axis. The dichroic mirror has high reflectivity to elastic scattered light and high transmittance to the fluorescent band.

Preferably, the scattered light collecting optical path for receiving the scattered light signal comprises a focusing mirror, a scattering aperture and a scattered light receiver which are sequentially placed along the transmission direction of the elastic scattered light.

Preferably, the fluorescence collection optical path for receiving the fluorescence signal comprises a fluorescence filter, a fluorescence focusing lens, a fluorescence diaphragm and a fluorescence receiver which are sequentially placed along the fluorescence transmission direction.

Preferably, the air circuit is composed of an air inlet nozzle, an air outlet nozzle and an air pump, and the air inlet direction of the air circuit is parallel to the Z axis and passes through the light sensitive area.

Preferably, the signal processing system includes a scattered light amplifying circuit, a scattered light AD conversion circuit, a fluorescence amplifying circuit, a fluorescence AD conversion circuit and an FPGA circuit; the elastic scattered light pulse signal collected by the scattered light receiver is sequentially amplified by the scattered light amplifying circuit, converted into a digital signal by the scattered light AD conversion circuit and reaches the FPGA circuit; the intrinsic fluorescence pulse signal collected by the fluorescence receiver is sequentially amplified by the fluorescence amplifying circuit, converted into a digital signal by the fluorescence AD conversion circuit and reaches the FPGA circuit, and the FPGA circuit is used to calculate the relative fluorescence intensity of the measured aerosol particles, the ratio of the scattering peak values and the intrinsic fluorescence bleaching rate.

On the other hand, the present invention also provides a method for monitoring bioaerosols using the above-mentioned bioaerosol monitoring device, comprising the following steps:

S1. Construct a three-dimensional coordinate system for the peak value, relative fluorescence intensity and intrinsic fluorescence bleaching rate of elastic scattered light pulse signals of different bioaerosol particles:

S1.1 When bioaerosol particles pass through the light-sensitive area, they stimulate elastic scattered light pulse signals and intrinsic fluorescence pulse signals, which are received by the scattered light receiver and the fluorescence receiver respectively, and are processed by the signal amplification circuit and the AD conversion circuit and then transmitted to the FPGA circuit;

S1.2 Calculate the peak value h ₀ of the elastic scattered light pulse signal of the bioaerosol particles to be tested, the peak value h ₁ of the intrinsic fluorescence pulse signal, and the area s ₁ of the intrinsic fluorescence pulse signal through the FPGA circuit;

S1.3. Calculate the relative fluorescence intensity, i.e., the ratio of the peak value _h1 of the intrinsic fluorescence pulse signal to the peak value _h0 of the elastic scattered light pulse signal, by using the FPGA circuit;

S1.4. Calculate the intrinsic fluorescence bleaching rate through the FPGA circuit, that is, the ratio of the area s ₁ of the intrinsic fluorescence pulse signal to the peak value h ₁ of the intrinsic fluorescence pulse signal;

S1.5 Repeat steps S1.1-S1.4 using different bioaerosol particles to construct a three-dimensional coordinate system of the peak value, relative fluorescence intensity and intrinsic fluorescence bleaching rate of the elastic scattered light pulse signal of different bioaerosol particles;

S2. When the bioaerosol particles to be tested pass through the light-sensitive area, elastic scattered light pulse signals and intrinsic fluorescence pulse signals are stimulated, which are received by the scattered light receiver and the fluorescence receiver respectively and transmitted to the FPGA circuit;

S3. The peak value, relative fluorescence intensity and intrinsic fluorescence bleaching rate of the elastic scattered light pulse signal of the biological aerosol particles to be tested are calculated by the FPGA circuit, and the type of the aerosol particles to be tested is determined by comparing the landing area in the three-dimensional coordinate system.

Compared with the prior art, the present invention has the following beneficial effects:

1) The focus of the excitation light is not located at the photosensitive area, but before or after the photosensitive area on the optical axis of the illumination light path, so that the cross section of the excitation light spot perpendicular to the X-axis in the photosensitive area is approximately rectangular, rather than a thin line. When a single aerosol particle passes through the photosensitive area, it will be continuously irradiated by the excitation light for a period of time, and the fluorescence bleaching characteristics of a single aerosol particle can be obtained.

2) Due to the different components of different aerosol particles, the bleaching speed of the fluorescence signal is not the same, and the fluorescence pulse waveform will also be significantly different. For aerosol particles that do not contain bleaching components in the fluorescence component, the fluorescence pulse waveform is approximately rectangular; for aerosol particles that only contain a small amount of bleaching components in the fluorescence component, the fluorescence pulse waveform is approximately trapezoidal; for aerosol particles whose fluorescent substances are mainly bleaching components, the fluorescence pulse waveform is approximately triangular. Since the airflow velocity and the size of the illumination beam are fixed, the time it takes for the aerosol particles to pass through the photosensitive area, that is, the duration of the fluorescence pulse signal, is basically fixed. The ratio of the area to the peak value of the fluorescence pulse signal of a single aerosol particle is calculated by FPGA, reflecting the fluorescence bleaching speed of the particle.

3) The intrinsic fluorescence bleaching rate can effectively distinguish between interferents and different types of bioaerosols, greatly reducing or even avoiding false detection of the instrument.

4) Through comprehensive analysis of the intrinsic fluorescence bleaching rate, scattered light peak, relative fluorescence intensity and other information of aerosol particles, preliminary classification of biological particles can be achieved, which can significantly reduce instrument alarms caused by non-target microorganisms and improve the instrument's ability to identify bioaerosol types, thereby reducing the frequency and cost of subsequent biological detection.

5) By setting a dichroic mirror in the optical path to separate the fluorescence signal into two or more fluorescence bands, a multi-channel fluorescence pulse signal is obtained, and the bleaching speed of fluorescence in different bands is calculated respectively, which increases the judgment index of aerosol particle classification, thereby achieving more accurate and precise judgment.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG1 is a schematic diagram of the optical path of an embodiment of a bioaerosol monitoring device of the present invention.

FIG. 2 is a schematic diagram of the illumination light path and the gas path in the embodiment of the bioaerosol monitoring device of the present invention.

FIG. 3 is a schematic diagram of a signal processing system of an embodiment of a bioaerosol monitoring device of the present invention.

FIG. 4 is a schematic diagram of the fluorescence signal pulse waveform of aerosol particles with different bleaching speeds according to the present invention.

DETAILED DESCRIPTION

The present invention will be further described below in conjunction with the accompanying drawings and embodiments, but the protection scope of the present invention should not be limited thereto.

Please refer to Figures 1 and 2. Figure 1 is a schematic diagram of the optical path of an embodiment of the bioaerosol monitoring device of the present invention, and Figure 2 is a schematic diagram of the illumination optical path and the gas path in the embodiment of the bioaerosol monitoring device of the present invention. As shown in the figure, the bioaerosol monitoring device of this embodiment includes an optical path, a gas path and a signal processing system. The optical path includes an excitation light source 101. The excitation light beam emitted by the excitation light source 101 is collimated by an aspherical mirror 102 in parallel with the X-axis direction to form a parallel light beam, and then focused by a cylindrical mirror 103 and passes through a light sensitive area G to reach a light trap 105. The focus M of the excitation light beam is located between the light sensitive area G and the light trap 105 on the optical axis of the illumination optical path. The light sensitive area G is approximately rectangular in cross section perpendicular to the X-axis. In the Y-axis direction of the light sensitive area G, there are curved reflectors 10 4. Photosensitive area G, dichroic mirror 106, the dichroic mirror 10 is set at 45 with the Y axis, the dichroic mirror 106 has high reflectivity to elastic scattered light and high transmittance to the fluorescent band, along the direction of elastic scattered light are scattered light focusing mirror 111, scattered light diaphragm 112, scattered light receiver 113, along the direction of fluorescence are fluorescent filter 107, fluorescent focusing mirror 108, fluorescent diaphragm 109, fluorescent receiver 110, the scattered light receiver 113 and the fluorescent receiver 110 are synchronously reached to the FPGA circuit (not shown) after passing through the signal amplification circuit and AD conversion circuit; the air path is composed of an air inlet nozzle 201, an exhaust nozzle 20 and an air pump, the air inlet direction of the air path is parallel to the Z axis and passes through the photosensitive area G; the intersection of the illumination light path and the air path channel is the photosensitive area G.

In the XOY plane, the light emitted by the excitation light source 101 is collimated by the aspheric mirror 102 to form a parallel beam, and then focused by the cylindrical mirror 103 and passes through the photosensitive area G to reach the light trap 105. The focus of the excitation light is between the photosensitive area G and the light trap 105 on the optical axis of the illumination light path. At this time, the photosensitive area is approximately rectangular in the cross section perpendicular to the X axis. The elastic scattered light and intrinsic fluorescence (if any) generated by the aerosol particles passing through the photosensitive area after being excited are collected by the curved reflector 104 of the collection unit and then converted into parallel light to reach the dichroic mirror 106. The dichroic mirror 106 has high reflection for elastic scattered light and high transparency for the fluorescence band. Therefore, the elastic scattered light is reflected by the dichroic mirror 106 to the focusing mirror 111 and then passes through the scattering aperture 112 to reach the scattered light receiver 113. The fluorescence is transmitted by the dichroic mirror 106 and passes through the fluorescence filter 107, the focusing mirror 108 and the fluorescence aperture 109 in sequence to reach the fluorescence receiver 110. The scattered light pulse signal obtained by the scattered light receiver 113 reaches the FPGA circuit 303 after passing through the first signal amplification circuit 301 and the first AD conversion circuit 302, and the fluorescent pulse signal obtained by the fluorescent receiver 110 reaches the FPGA circuit 303 synchronously after passing through the second signal amplification circuit 304 and the second AD conversion circuit 305, as shown in FIG. 3 .

FIG2 is a schematic diagram of the relative positions of the illumination optical path and the gas path. In the XOZ plane, the gas path is composed of an air inlet nozzle 201, an exhaust nozzle 202, and an air pump. The air inlet direction of the gas path is parallel to the Z axis and passes through the light sensitive area G.

The fluorescence pulse waveforms of different aerosol particles will also be different, as shown in Figure 4. For aerosol particles whose fluorescent components do not contain bleaching components, their fluorescence pulse waveform is approximately rectangular, as shown in Figure 4 (a); for aerosol particles whose fluorescent components contain only a small amount of bleaching components, their fluorescence pulse waveform is approximately trapezoidal, as shown in Figure 4 (b); for aerosol particles whose fluorescent substances are mainly bleaching components, their fluorescence pulse waveform is approximately triangular, as shown in Figure 4 (c).

The monitoring method of the present invention is as follows:

(1) After the monitoring device is started, the aerosol particles to be measured will emit elastic scattered light pulse signals, and some particles will also emit intrinsic fluorescence pulse signals at the same time. If the measured particle only has elastic scattered light signals, it is determined to be a non-biological particle. Otherwise, the elastic scattered light signal and intrinsic fluorescence pulse signal of the measured particle are simultaneously transferred to the FPGA circuit after passing through the weak signal amplification circuit and the AD conversion circuit. The FGPA circuit obtains the peak value h0 of the scattered pulse signal of the measured aerosol particle, the area s1 of the fluorescence pulse signal, and the peak value h1 of the fluorescence pulse signal through calculation;

(2) FPGA further calculates the relative fluorescence intensity of the aerosol particles being measured, that is, the ratio of the fluorescence peak to the scattering peak h1/h0, and the intrinsic fluorescence bleaching rate s1/h1.

(3) According to step (1) and step (2), the scattered light peak value h0, relative fluorescence intensity h1/h0 and intrinsic fluorescence bleaching rate s1/h1 of various aerosol particles that can emit intrinsic fluorescence are tested, and then a three-dimensional coordinate system is established with these three detection values, the coordinates of different types of aerosol particles are marked, and the regions corresponding to the different types of aerosol particles are divided in the coordinate system.

(4) When testing unknown fluorescent aerosol particles, the monitoring device detects and calculates the scattered light peak, relative fluorescence intensity and intrinsic fluorescence bleaching rate of the aerosol particles to be tested, thereby obtaining the landing point position of the particles to be tested in the three-dimensional coordinate system, and then determining the type of the aerosol particles to be tested according to the corresponding division area in step (3) corresponding to the landing point position.

Experiments show that the present invention collects characteristic information of different types of aerosol particles by analyzing the area, amplitude and other information of the intrinsic fluorescence pulse signal generated when aerosol particles pass through the photosensitive area under continuous irradiation, and combining it with the elastic scattering light pulse signal of the particles, thereby providing a basis for the preliminary classification of aerosol particles.

Claims (7)

A bioaerosol monitoring device comprises an optical path, an air path intersecting the optical path, and a signal processing system connected to the optical path; the optical path comprises a laser irradiation optical path for irradiating measured particles, a scattered light collection optical path for receiving scattered light signals, and a fluorescence collection optical path for receiving fluorescence signals; the air path is used to sample measured particles; the signal processing system is used to analyze and process signals; the intersection area of the laser irradiation optical path and the air path is a light sensitive area (G); it is characterized in that the focus (M) of the excitation light source for irradiating measured particles is located before or after the light sensitive area (G) on the optical axis of the laser irradiation optical path, so that the cross section of the excitation light spot perpendicular to the optical axis of the laser irradiation optical path in the light sensitive area (G) is rectangular, and when a single aerosol particle passes through the light sensitive area, it is continuously irradiated by the excitation light to obtain an intrinsic fluorescence bleaching rate. The bioaerosol monitoring device according to claim 1 is characterized in that the laser irradiation light path for irradiating the measured particles includes an excitation light source (101), the light emitted by the excitation light source (101) is parallel to the X-axis direction and collimated by an aspheric mirror (102) to form a parallel light beam, and then focused by a cylindrical mirror (103) and passes through a light sensitive area (G) to reach a light trap (105), and in the Y-axis direction of the light sensitive area (G) are a curved reflector (104) and a dichroic mirror (106), the dichroic mirror (106) is arranged at 45° with the Y-axis, and the dichroic mirror (106) is highly reflective to elastic scattered light and highly transparent to the fluorescent band. The bioaerosol monitoring device according to claim 1 is characterized in that the scattered light collection optical path for receiving the scattered light signal includes a focusing mirror (111), a scattering aperture (112) and a scattered light receiver (113) arranged in sequence along the transmission direction of the elastic scattered light. The bioaerosol monitoring device according to claim 1 is characterized in that the fluorescence collection optical path for receiving the fluorescence signal includes a fluorescence filter (107), a fluorescence focusing lens (108), a fluorescence diaphragm (109) and a fluorescence receiver (110) arranged in sequence along the fluorescence transmission direction. The bioaerosol monitoring device according to claim 1 is characterized in that the air path is composed of an air inlet nozzle (201), an air exhaust nozzle (202) and an air pump, and the air inlet direction of the air path is parallel to the Z axis and passes through the light sensitive area (G). The bioaerosol monitoring device according to claim 1 is characterized in that the signal processing system comprises a scattered light amplifying circuit (301), a scattered light AD conversion circuit (302), a fluorescence amplifying circuit (303), a fluorescence AD conversion circuit (305) and an FPGA circuit (303); the elastic scattered light pulse signal collected by the scattered light receiver (113) is sequentially amplified by the scattered light amplifying circuit (301), converted into a digital signal by the scattered light AD conversion circuit (302) and reaches the FPGA circuit (303); the intrinsic fluorescence pulse signal collected by the fluorescence receiver (110) is sequentially amplified by the fluorescence amplifying circuit (303), converted into a digital signal by the fluorescence AD conversion circuit (305) and reaches the FPGA circuit (303), and the FPGA circuit is used to calculate the relative fluorescence intensity of the aerosol particles being measured, the ratio of the scattering peak value and the intrinsic fluorescence bleaching rate. A bioaerosol monitoring method is implemented using the bioaerosol monitoring device according to any one of claims 1 to 6, comprising the following steps: S1. Construct a three-dimensional coordinate system for the peak value, relative fluorescence intensity and intrinsic fluorescence bleaching rate of elastic scattered light pulse signals of different bioaerosol particles: S1.1 When the bioaerosol particles pass through the light-sensitive area, elastic scattered light pulse signals and intrinsic fluorescence pulse signals are stimulated, which are received by the scattered light receiver (113) and the fluorescence receiver (110) respectively, and are transmitted to the FPGA circuit after being processed by the signal amplification circuit and the AD conversion circuit; S1.2 Calculate the peak value h ₀ of the elastic scattered light pulse signal of the bioaerosol particles to be tested, the peak value h ₁ of the intrinsic fluorescence pulse signal, and the area s ₁ of the intrinsic fluorescence pulse signal through the FPGA circuit; S1.3. Calculate the relative fluorescence intensity by FPGA circuit, that is, the ratio of the peak value _h1 of the intrinsic fluorescence pulse signal to the peak value _h0 of the elastic scattered light pulse signal; S1.4. Calculate the intrinsic fluorescence bleaching rate through the FPGA circuit, that is, the ratio of the area s ₁ of the intrinsic fluorescence pulse signal to the peak value h ₁ of the intrinsic fluorescence pulse signal; S1.5 Repeat steps S1.1-S1.4 using different bioaerosol particles to construct a three-dimensional coordinate system of the peak value, relative fluorescence intensity and intrinsic fluorescence bleaching rate of the elastic scattered light pulse signal of different bioaerosol particles; S2. When the bioaerosol particles to be tested pass through the light-sensitive area, elastic scattered light pulse signals and intrinsic fluorescence pulse signals are stimulated, which are received by the scattered light receiver (113) and the fluorescence receiver (110) respectively and transmitted to the FPGA circuit; S3. The peak value, relative fluorescence intensity and intrinsic fluorescence bleaching rate of the elastic scattered light pulse signal of the biological aerosol particles to be tested are calculated by the FPGA circuit, and the type of the aerosol particles to be tested is determined by comparing the landing area in the three-dimensional coordinate system.