Device and method for detecting fluorescence lifetime of optical fiber optical tweezers

Abstract

The invention provides optical fiber optical tweezer fluorescence lifetime detection equipment and a detection method. A light beam emitted by a light source is divided into two paths, two paths of received optical signals are coupled by using a first optical fiber coupler, and the coupled optical signals are transmitted to a connected optical fiber circulator and are transmitted to a displacement control assembly through the optical fiber circulator. The displacement control assembly captures target tiny particles by using one path of light signals, excites the target tiny particles by using the light signals corresponding to the other path of light beams to generate excitation fluorescence signals, and performs photon number detection on the excitation fluorescence signals by using the detection assembly to determine the fluorescence lifetime of the target tiny particles. According to the method, the fluorescence lifetime of the tiny particles can be detected while the tiny particles are flexibly controlled, and the method is simpler, more convenient and more efficient, and is suitable for ultrafast signal detection of a precise large instrument.

Description

Technical Field

[0001] The present invention relates to the field of microscale manipulation technology, and in particular to a fiber optic tweezers fluorescence lifetime detection device and a detection method. Background Art

[0002] Fiber tweezers fluorescence lifetime detection is a technology that combines fiber tweezers technology with fluorescence lifetime detection. While using fiber tweezers to manipulate particles, a laser of a specific wavelength is used to excite the particles to produce fluorescence. The fluorescence decay process is then measured through a detection system to obtain fluorescence lifetime information.

[0003] The current fiber optic tweezers fluorescence lifetime detection technology usually requires the use of microchannel or microfluidic technology for analysis in terms of multi-dimensional controllable operation. However, the preparation of microfluidic control components is complex, and as independent devices from the optical fiber, the system's optical coupling efficiency and integration are low, making it impossible to achieve efficient particle or cell manipulation. In addition, the method of using multiple single-mode optical fibers or splicing multiple optical fibers for optical capture has low manipulation flexibility due to the need to operate multiple optical fibers. Therefore, the methods in the existing technology cannot achieve flexible manipulation of tiny particles or cells to achieve fluorescence lifetime detection.

[0004] Therefore, the prior art needs to be further improved. Summary of the Invention

[0005] In view of the above-mentioned deficiencies in the prior art, the purpose of the present invention is to provide a fiber optic tweezers fluorescence lifetime detection device and detection method, which adopts a multi-mode fiber loop combination to realize multi-wavelength incidence, and uses multi-wavelength light beams to simultaneously realize fluorescence lifetime detection and microscale optical manipulation.

[0006] In a first aspect, the present application provides a fiber optic tweezers fluorescence lifetime detection device, comprising:

[0007] A light source assembly for emitting a first light beam and a second light beam with different wavelengths;

[0008] a first objective lens and a second objective lens, respectively disposed on the optical paths of the first light beam and the second light beam, for collecting optical signals of the first light beam and the second light beam, and transmitting the collected optical signals to a first optical fiber coupler;

[0009] The first optical fiber coupler is configured to couple the received optical signals corresponding to the first and second optical beams, and transmit the coupled optical signals to a connected optical fiber circulator;

[0010] The optical fiber circulator is used to transmit the optical signal corresponding to the first light beam and the optical signal corresponding to the second light beam to the displacement control component;

[0011] The displacement control component has an optical fiber end for capturing target microparticles, wherein the target microparticles are captured by a high-order focused beam formed by the optical signal corresponding to the first light beam at the optical fiber end, and the target microparticles generate an excitation fluorescence signal under the irradiation of the optical signal corresponding to the second light beam, and the excitation fluorescence signal is transmitted to the second optical fiber coupler via the optical fiber circulator;

[0012] The detection component is connected to the output end of the second optical fiber coupler, and is used to receive the excitation fluorescence signal output by the second optical fiber coupler, and detect the number of photons of the excitation fluorescence signal to determine the fluorescence lifetime of the target tiny particle.

[0013] Optionally, the light source assembly includes: a light source, a parallel light transmission system, a frequency doubling crystal element and a half-reflecting half-mirror lens;

[0014] The light source is used to emit a femtosecond laser beam;

[0015] The parallel light transmission system includes a first lens and a second lens, wherein the first lens and the second lens form a 4f system, the frequency doubling crystal element is located between the first lens and the second lens, and the first lens, the frequency doubling crystal element and the second lens are sequentially arranged on the optical path of the femtosecond laser beam, and are used to parallelize and frequency-double the femtosecond laser beam to output a mixed beam containing two different wavelengths;

[0016] The half-reflecting half-mirror is arranged on the optical path of the mixed light beam, and is used to separate the two optical signals of different wavelengths in the mixed light beam into two light beams by transmission and reflection respectively, to obtain a first light beam and a second light beam.

[0017] Optionally, the system is further provided with a reflector;

[0018] The reflecting mirror is arranged on the optical path of the first light beam after passing through the half-reflecting half-mirror lens, and reflects the first light beam to the first objective lens.

[0019] Optionally, the displacement control component includes an optical fiber end, a microscopic imaging component and a position control platform;

[0020] The microscopic imaging component is used to convert the optical signal corresponding to the first light beam output from the optical fiber circulator into an electrical signal to capture image information within the target area;

[0021] The position control platform is used to control the movement of the optical fiber end in the target area so that the optical fiber end captures target tiny particles in the target area.

[0022] Optionally, the position control platform is a six-axis precision translation platform, and the end face of the optical fiber end is a three-dimensional structure.

[0023] Optionally, the detection assembly includes: a photomultiplier tube and a time-correlated single photon counter;

[0024] The photomultiplier tube is used to amplify the received excitation fluorescence signal;

[0025] The time-correlated single-photon counter is used to measure the number of photons in the received excitation fluorescence signal, so as to determine the fluorescence lifetime of the target tiny particle according to the number of received photons.

[0026] Optionally, the wavelength of the femtosecond laser beam is 800 nm; the wavelength corresponding to the first beam is 800 nm, and the wavelength corresponding to the second beam is 400 nm.

[0027] Optionally, the focal length of the first lens and the second lens is 15 mm.

[0028] In a second aspect, the present application provides a method for detecting fluorescence lifetime using the optical fiber tweezers fluorescence lifetime detection device, which includes:

[0029] The first objective lens and the second objective lens respectively collect optical signals of the first light beam and the second light beam emitted by the light source assembly, and transmit the collected optical signals to the first optical fiber coupler; the first light beam and the second light beam have different wavelengths;

[0030] The first optical fiber coupler couples the received optical signals corresponding to the first and second optical beams, and transmits the coupled optical signals to the displacement control component via the optical fiber circulator;

[0031] The displacement control component uses the optical signal corresponding to the first light beam to capture the target tiny particles. The target tiny particles generate an excitation fluorescence signal under the irradiation of the optical signal corresponding to the second light beam. The excitation fluorescence signal is transmitted to the detection component via the optical fiber circulator and the second optical fiber coupler.

[0032] The detection component is used to detect the number of photons of the excitation fluorescence signal to determine the fluorescence lifetime of the target tiny particles.

[0033] Optionally, before the step of respectively collecting the light signals of the first light beam and the second light beam emitted by the light source assembly through the first objective lens and the second objective lens, the method further comprises:

[0034] The light source emits a femtosecond laser beam;

[0035] The parallel light transmission system arranged on the optical path of the femtosecond laser beam parallelizes and frequency-doubles the femtosecond laser beam, and outputs a mixed beam containing two different wavelengths;

[0036] A half-reflective half-mirror mirror arranged on the optical path of the mixed light beam separates the optical signals of two different wavelengths in the mixed light beam into two light beams to obtain a first light beam and a second light beam.

[0037] Beneficial effects:

[0038] The present invention provides a fiber optic tweezers fluorescence lifetime detection device and detection method. The device comprises the following steps: a light beam emitted by a light source component is divided into a first light beam and a second light beam of different wavelengths; a first objective lens and a second objective lens are used to collect the optical signals of the first and second light beams, and the collected optical signals are transmitted to a first optical fiber coupler; the two optical signals are coupled using the first optical fiber coupler, and the coupled optical signals are transmitted to a connected optical fiber circulator; the optical fiber circulator transmits the optical signals corresponding to the two light beams to a displacement control component, which uses the optical signal corresponding to the first light beam to capture target microparticles and uses the optical signal corresponding to the other light beam to excite the target microparticles to generate an excitation fluorescence signal. The excitation fluorescence signal is transmitted to the second optical fiber coupler via the optical fiber circulator, and the detection component detects the number of photons in the excitation fluorescence signal to determine the fluorescence lifetime of the target microparticles. The method of the present invention can detect the fluorescence lifetime of microparticles while flexibly manipulating them. The method is simpler and more efficient, and is suitable for ultrafast signal detection in large-scale precision instruments. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 This is a structural principle diagram of the optical fiber tweezers fluorescence lifetime detection device provided by the present invention;

[0040] Figure 2 It is a structural schematic diagram of a specific embodiment of the optical fiber tweezers fluorescence lifetime detection device provided by the present invention;

[0041] Figure 3 This is a flow chart of the steps of a method for realizing fluorescence lifetime detection by the optical fiber tweezers fluorescence lifetime detection device provided by the present invention. DETAILED DESCRIPTION

[0042] In order to make the purpose, technical solutions and advantages of the present invention more clear and distinct, the present invention is further described in detail below with reference to the accompanying drawings and examples. It should be understood that the specific embodiments described herein are only used to illustrate the present invention and are not intended to limit the present invention.

[0043] Fluorescence lifetime detection, a highly sensitive and time-resolution analytical technique, has demonstrated unique application value in a wide range of fields. Its core principle is to reveal molecular dynamics, interactions, and environmental information by measuring the time distribution of fluorescent molecules returning from an excited state to their ground state. It provides a powerful analytical tool in fields such as biomedicine and life sciences, materials science and nanotechnology, environmental monitoring and chemical analysis, and food and agriculture.

[0044] Optical fiber tweezers are a fiber-optic tweezer system that uses optical fibers to transmit and focus laser beams, enabling contactless capture and manipulation of tiny particles. Compared to traditional microscope-based optical tweezers, fiber tweezers offer advantages such as compactness, flexibility, and ease of integration. They hold broad application prospects in biomedicine, micro- and nanomanipulation, and optomechanics.

[0045] The core of fiber optic tweezers is to use optical fiber to transmit the laser beam to a tiny area, and through the special design of the optical fiber end face (such as tapering, lensing, etc.), focus the laser beam to the micrometer or even nanometer scale, forming a high-intensity gradient light field. When tiny particles (such as cells, bacteria, microspheres, etc.) enter the light field, they will be affected by the light radiation pressure, including scattering force and gradient force. The gradient force pulls the particles to the area with the greatest light intensity, while the scattering force tends to push the particles along the direction of light beam propagation. By precisely controlling the parameters of the light field (such as power, wavelength, polarization, etc.), stable capture and three-dimensional manipulation of particles can be achieved.

[0046] Many research institutions at home and abroad are currently conducting research on fiber-optic tweezers. Some researchers have successfully manipulated mammalian cells using a single, fine optical fiber. Some research teams have used two specially manufactured, mutually opposing optical waveguide rings to stably capture small particles and red blood cells. Some teams have used tapered optical fibers to create simple single-fiber optical tweezers systems, capturing polystyrene spheres and active yeast cells in a fluid at the focal point of the light beam, respectively, allowing the manipulated and captured objects to move freely and synchronously with the fiber-optic tweezers. Domestic micro-nano optics teams were the first to conduct research on optical tweezers and were the first to develop a three-dimensional fiber-optic tweezers system. Therefore, the scientific question of how to use femtosecond lasers to establish a three-dimensional fiber-optic tweezers system with built-in two-photon polymerization to achieve ultrafast information detection while simultaneously performing microscale manipulation is of paramount importance.

[0047] Currently, fiber optic tweezers need to be analyzed in conjunction with microchannel or microfluidic technology for multi-dimensional controllable operations. The preparation of microfluidic devices is complex, and as independent devices from optical fibers, the system's optical coupling efficiency and integration are low. Therefore, a simpler, more efficient, and highly integrated method for microparticle or cell manipulation is needed. In addition, researchers also use multiple single-mode optical fibers or multiple optical fiber splicing for optical capture. However, although multiple single-mode optical fibers or multiple optical fiber splicing can use light scattering forces to directly capture and manipulate tiny particles and cells, their manipulation flexibility is limited and their functions are single.

[0048] In order to overcome the above-mentioned problems in the prior art, the present application provides a fiber optic tweezers fluorescence lifetime detection device and detection method, which divides the light beam emitted by the light source into two paths with different wavelengths, and uses a fiber optic circulator to transmit the two light beams to the displacement control component, and the displacement control component uses the light signal corresponding to the first light beam to capture the target tiny particles. The captured target tiny particles generate an excitation fluorescence signal under the irradiation of the light signal corresponding to the second light beam, and the detection component detects the number of photons in the excitation fluorescence signal to determine the fluorescence lifetime of the target tiny particles. The fluorescence lifetime detection device and detection method provided by the present application can not only achieve the uplink of various excitation light captures, but also realize the backtransmission of multiple detection signals, that is, simultaneously realize the manipulation of tiny particles and the detection of fluorescence lifetime, with high operational flexibility, and suitable for ultrafast signal detection in precision large instruments.

[0049] The following is a more detailed description of the optical fiber tweezers fluorescence lifetime detection device and detection method provided by the present application in conjunction with the accompanying drawings.

[0050] In the first aspect, the present application provides a fiber optic tweezers fluorescence lifetime detection device, such as Figure 1 Shown, including:

[0051] The light source assembly 10 is configured to emit a first light beam and a second light beam having different wavelengths.

[0052] The light source assembly is used to emit two light beams, and the wavelengths of the two light beams are different. The two light beams of different wavelengths have different functions. In order to emit a light beam, the light source assembly includes a light source that emits a light beam. Since the light beam emitted by the light source is used to form optical tweezers for fluorescence lifetime detection, the light source used in this application is a femtosecond laser light source. Femtosecond laser is an ultrashort pulse laser with a pulse duration in the order of femtoseconds. It has the characteristics of high peak power, short pulse width, and small thermal effect, so it can achieve clear imaging. In this embodiment, an 800nm ​​femtosecond laser light source is preferably used. Since the light source assembly needs to emit two light beams with different wavelengths, the light source assembly can also use nonlinear optical crystal frequency doubling to achieve a mixed light beam containing two different wavelengths, and then use a filter or grating for splitting.

[0053] Furthermore, the laser corresponding to the femtosecond laser light source used in this embodiment is a femtosecond pulse laser with a pulse width of 100fs, 80MHz, and a near-infrared band of 805nm. Since this type of femtosecond pulse laser has the characteristics of high repetition rate and short pulse width, it has the advantages of high time resolution, high signal-to-noise ratio and high measurement accuracy in ultrafast time scale detection.

[0054] The first objective lens 105 and the second objective lens 106 are respectively arranged on the optical paths of the first light beam and the second light beam, and are used to collect optical signals of the first light beam and the second light beam, and transmit the collected optical signals to the first fiber coupler 107 .

[0055] The first light beam and the second light beam emitted by the light source assembly are collected by the first objective lens and the second objective lens respectively, and the optical signals collected by the first objective lens and the second objective lens are both transmitted to the first optical fiber coupler.

[0056] Combine Figure 2 As shown, a first objective lens 105 is disposed on the optical path of the first light beam and is used to receive the optical signal corresponding to the first light beam. A second objective lens 106 is disposed on the optical path of the second light beam and is used to receive the optical signal corresponding to the second light beam. Both the first objective lens 105 and the second objective lens 106 are connected to a first fiber coupler 107. Therefore, when the first objective lens 105 and the second objective lens 106 receive the optical signal, they transmit the received optical signal to the first fiber coupler 107.

[0057] The first optical fiber coupler 107 is used to couple the received optical signals corresponding to the first light beam and the second light beam, and transmit the coupled optical signals to the connected optical fiber circulator 108 .

[0058] One end of the first optical fiber coupler is connected to the first objective lens and the second objective lens, and the other end is connected to the optical fiber circulator. When the optical signals of the first light beam and the second light beam are received, the two optical signals are coupled and the coupled optical signals are transmitted to the optical fiber circulator.

[0059] The optical fiber circulator 108 is used to transmit the optical signal corresponding to the first light beam and the optical signal corresponding to the second light beam to the displacement control component. The optical fiber circulator is provided with an input port and an output port for optical signals. The optical signal corresponding to the first light beam and the optical signal corresponding to the second light beam are transmitted to the displacement control component via the output port.

[0060] A fiber circulator is a multi-port, nonreciprocal optical device. Signal transmission direction is irreversible, meaning light can only propagate in one direction, enabling bidirectional optical signal transmission on a single optical fiber. While optical signals can be redirected during transmission within the circulator, they must pass through the ports sequentially in one direction. Fiber circulators typically have three ports. In this step, the optical signals corresponding to the first and second beams are input to the displacement control assembly through the same output port.

[0061] The displacement control component 111 is used to capture target microparticles using the optical signal corresponding to the first light beam, and the target microparticles generate an excitation fluorescence signal under the irradiation of the optical signal corresponding to the second light beam. The excitation fluorescence signal is transmitted to the second optical fiber coupler through the optical fiber circulator.

[0062] The displacement control assembly disclosed in this embodiment includes: an optical fiber end and a position control platform for controlling the movement position of the optical fiber end. The surface of the optical fiber end is configured as a three-dimensional structure (which may be conical in shape). The optical signal corresponding to the first light beam forms a high-order focused light beam on the three-dimensional structure of the end face of the optical fiber end. This high-order focused light beam can exert a specific optical force on tiny particles, thereby achieving the capture and manipulation of tiny particles. The position control platform can control the movement of the optical fiber end in different directions to achieve the capture of tiny particles or cells.

[0063] The displacement control assembly disclosed in this embodiment is used to precisely control the position of the optical fiber end to capture target microparticles. The captured target microparticles, stimulated by the optical signal corresponding to the second light beam, emit excitation light. The excitation light emitted by the target microparticles can be transmitted via a fiber circulator to a detection assembly, enabling the detection assembly to detect the lifetime of the fluorescence signal. Because the fiber circulator can have two output ports, the fluorescence signal can be output from the other output port and transmitted to the detection assembly via a second fiber coupler.

[0064] The detection component 110 is connected to the output end of the second optical fiber coupler 109 and is used to detect the number of photons of the excitation fluorescence signal to determine the fluorescence lifetime of the target tiny particles.

[0065] The optical signal corresponding to the second light beam is input to the area corresponding to the detection component as detection light, so that the detection component can detect the excited fluorescence signal.

[0066] Combine Figure 2 As shown, further, the light source assembly 10 includes: a light source 101, a parallel light transmission system 102, a frequency doubling crystal element 1022 and a half-reflecting half-mirror lens 103.

[0067] In detail, the light source 101 is used to emit a femtosecond laser beam.

[0068] The parallel light transmission system 102 includes a first lens 1021 and a second lens 1023, and the first lens 1021 and the second lens 1023 form a 4f system; the frequency doubling crystal element 1022 is located between the first lens 1021 and the second lens 1023, and the first lens 1021, the frequency doubling crystal element 1022 and the second lens 1023 are arranged in sequence on the optical path of the femtosecond laser beam, and are used to parallelize and frequency-double the femtosecond laser beam and output a mixed beam containing two different wavelengths.

[0069] The half-reflecting half-mirror 103 is arranged on the optical path of the mixed light beam, and is used to separate the two optical signals of different wavelengths in the mixed light beam into two light beams by transmission and reflection respectively, to obtain a first light beam and a second light beam.

[0070] Combine Figure 2 As shown, the light beam emitted by the light source, after passing through the frequency-doubling crystal element, becomes a mixed beam containing two different wavelengths. For example, if the wavelength of the light beam emitted by the light source is 800nm, the frequency-doubling crystal element will double the 800nm ​​light signal to produce a mixed beam containing 800nm ​​and 400nm. The mixed beam is reflected and transmitted through a half-reflecting half-mirror lens, resulting in two beams with different wavelengths. For example, a mixed beam containing 800nm ​​and 400nm passes through a 400-600nm half-reflecting half-mirror lens and is then split into two beams of 800nm ​​and 400nm.

[0071] Furthermore, in order to change the transmission direction of the light beam, the device is further provided with a reflector 104 .

[0072] The reflector 104 is disposed on the optical path of the first light beam after passing through the half-reflecting mirror 103 , and reflects the first light beam to the first objective lens 105 .

[0073] Furthermore, the displacement control assembly 111 includes an optical fiber end, a microscopic imaging assembly 1112, and a position control platform 1111. The microscopic imaging assembly 1112 is used to convert the optical signal corresponding to the first light beam output from the optical fiber circulator 108 into an electrical signal to capture image information within the target area; the position control platform 1111 is used to control the movement of the optical fiber end within the target area to capture target microparticles within the target area using a high-order focused light beam formed on the surface of the optical fiber end.

[0074] To achieve precise manipulation of tiny particles within a target area, this embodiment utilizes a position control platform to drive the end face of an optical fiber tip, capturing the tiny particles using a high-order focused beam formed on the surface of the optical fiber tip. In one implementation, the position control platform 1111 is a six-axis precision translation stage. This six-axis precision translation stage enables precise control and tiny displacement in the x, y, and z directions in three dimensions. The end face of the optical fiber tip has a three-dimensional structure. This three-dimensional structure on the end face of the optical fiber tip can create a specific light field distribution on the end face of the fiber, localizing the light field energy within a region far below the diffraction limit, forming a high-intensity electromagnetic field hotspot. This provides a significant optical gradient force, enabling high-precision capture of nanoscale particles and improving capture accuracy. The optical fiber tip with a three-dimensional structure can dynamically control the surface plasmon light field through spatial movement of the probe or optical fiber. This dynamic control capability enables more flexible manipulation of tiny particles using optical tweezers, meeting the needs of diverse experiments and applications. Furthermore, the three-dimensional structure design optimizes the interaction between the light field and the tiny particles, improving capture stability. For example, by designing a suitable three-dimensional structure, the light field can be distributed more evenly around the particle, thereby reducing the possibility of particle escape. Therefore, the displacement control component provided by this application can more accurately capture and control the movement of target tiny particles.

[0075] Furthermore, the microscopic imaging component includes a CCD and a high-magnification objective lens. The CCD acts as an image sensor, converting the light signal focused by the objective lens into an electrical signal. Its high sensitivity allows it to capture weak fluorescence signals. The high-magnification objective lens can magnify and focus the target tiny particles or cells onto the CCD target surface, improving imaging quality.

[0076] The target tiny particles may be tiny particles or cells in a solution environment. Therefore, the six-axis precision translation stage of this embodiment drives the controllable optical fiber to move to achieve the capture of the tiny particles.

[0077] In one implementation, the detection assembly 110 includes a photomultiplier tube 1101 and a time-correlated single-photon counter 1102. The photomultiplier tube 1101 is configured to amplify the received excitation fluorescence signal and the optical signal corresponding to the second light beam. The time-correlated single-photon counter 1102 is configured to measure the number of photons in the received excitation fluorescence signal to determine the fluorescence lifetime of the target microparticle based on the received photon count.

[0078] Photomultiplier tubes (PMTs) can amplify tiny signals, switching from low to high modes. Because they have high sensitivity, they can detect extremely weak light signals. They also have a fast response speed and can capture rapidly changing light signals. Therefore, the use of PMTs can improve detection accuracy.

[0079] A time-correlated single-photon counter (TCSPC) accurately measures fluorescence decay by recording the arrival time of individual fluorescence photons. In fluorescence lifetime detection, it captures weak fluorescence signals and converts them into time-correlated electrical signals, providing the basis for subsequent data processing and analysis.

[0080] Time-correlated single-photon counters (TCSPCs) are a high-precision technology for measuring periodic photon events. Their core principle is to periodically excite a sample with a pulsed light source. A detector captures individual photons, generating an analog signal. A processing unit determines the photon arrival time based on this signal and constructs a time histogram through discretized sampling to characterize the temporal distribution of the photon sequence. In a TCSPC system, the photon counter measures the detection time of individual photons and reconstructs the waveform based on these individual time measurements. A time-correlated single-photon counter is used to detect single photons in periodic optical signals. For low-level, high-repetition-rate signals, the light intensity is typically very low, and the probability of detecting a single photon within a signal cycle is far less than 1, making the detection of multiple photons negligible. When a photon is detected, the time of the corresponding detector pulse is measured, and the event is accumulated by adding a "1" to a memory address proportional to the detection time. After many photons have passed, a histogram of the detection times, representing the waveform of the light pulse, is obtained. Based on the waveform analysis of the light pulse, the average time it takes for the fluorescent molecule to return from the excited state to the ground state is determined, thereby determining the fluorescence lifetime. Since the time-correlated single-photon counter has the advantages of ultra-high time resolution and ultra-high sensitivity, it can achieve better measurement accuracy when applied to fluorescence lifetime measurement.

[0081] To achieve optimal detection results, the wavelength of the femtosecond laser beam in this embodiment is 800 nm. The wavelength corresponding to the first beam is 800 nm, and the wavelength corresponding to the second beam is 400 nm. Optionally, the focal lengths of the first and second lenses are 15 mm.

[0082] The method of this embodiment, combined with fiber-optic tweezers, offers advantages in high sensitivity and operational flexibility for a wide range of invasive detection applications, such as deep tissue exploration. Furthermore, captured particles and large particles can serve as quantitative drug carriers. The fluorescence lifetime of fluorescent molecules, highly sensitive to changes in various environmental factors, can serve as a localization signal. For example, the detection device disclosed in this embodiment can be used to implement a prototype medical device capable of both environmental location and targeted, quantitative drug delivery.

[0083] The present invention provides a fiber optic tweezers fluorescence lifetime detection device, which is configured to split a light beam emitted by a light source into two paths, couple the two received light signals using a first fiber optic coupler, and transmit the coupled light signals to a connected fiber optic circulator; and transmit the two light signals to a displacement control component using the fiber optic circulator. The displacement control component uses one light signal to capture target microparticles, and the target microparticles generate an excitation fluorescence signal under the irradiation of the light signal corresponding to the other light beam. The generated excitation fluorescence signal is transmitted to the detection component via the fiber optic circulator and the second fiber optic coupler. The detection component detects the number of photons in the excitation fluorescence signal to determine the fluorescence lifetime of the target microparticles. The device of the present invention can detect the fluorescence lifetime of microparticles while flexibly manipulating them. The present invention is more convenient and efficient, and is suitable for ultrafast signal detection in large-scale precision instruments.

[0084] In a second aspect, the present application provides a fluorescence lifetime detection method, which is applied to the above-mentioned fiber optic tweezers fluorescence lifetime detection device, such as Figure 3 As shown, the fluorescence lifetime detection method specifically includes:

[0085] Step S1: A first objective lens and a second objective lens respectively collect optical signals of a first light beam and a second light beam emitted by a light source assembly, and transmit the collected optical signals to a first optical fiber coupler; the first light beam and the second light beam have different wavelengths.

[0086] The light source assembly emits two beams of light, each with different wavelengths and functions. The first beam forms a high-order focused beam on the end face of the optical fiber to capture target microparticles, while the second beam excites the captured microparticles to emit fluorescence.

[0087] Specifically, in order to enable the light source assembly to emit two light beams with different wavelengths, further, before the step of respectively collecting the light signals of the first light beam and the second light beam emitted by the light source assembly through the first objective lens and the second objective lens, the method further includes:

[0088] Step S01: A light source emits a femtosecond laser beam.

[0089] Step S02 : A parallel light transmission system arranged on the optical path of the femtosecond laser beam performs parallel and frequency-doubled processing on the femtosecond laser beam, and outputs a mixed light beam containing two different wavelengths.

[0090] Step S03: A half-reflecting half-mirror mirror arranged on the optical path of the mixed light beam separates the optical signals of two different wavelengths in the mixed light beam into two light beams to obtain a first light beam and a second light beam.

[0091] Step S2: The first optical fiber coupler couples the received optical signals corresponding to the first light beam and the second light beam, and transmits the coupled optical signals to the displacement control component via the optical fiber circulator.

[0092] The first objective lens and the second objective lens are both connected to a first optical coupler. When the first and second objective lenses receive the optical signals corresponding to the first and second light beams, respectively, they input the received optical signals into the first optical coupler. A fiber circulator is connected to the first optical coupler, and the first optical coupler transmits the coupled light corresponding to the first and second light beams to the fiber circulator. The coupled light beams are then transmitted to the displacement control assembly via the fiber circulator.

[0093] Step S3: The displacement control component uses the optical signal corresponding to the first light beam to capture the target tiny particles. The target tiny particles generate an excitation fluorescence signal under the irradiation of the optical signal corresponding to the second light beam. The excitation fluorescence signal is transmitted to the detection component via the optical fiber circulator and the second optical fiber coupler.

[0094] The displacement control component area is composed of a CCD and a six-axis precision displacement stage. The displacement control system can control the movement of optical fibers with three-dimensional structures on the end face of the optical fiber to capture tiny particles or cells in a solution environment.

[0095] Step S4: Utilize a detection component to detect the number of photons in the excitation fluorescence signal to determine the fluorescence lifetime of the target microparticles.

[0096] The detection component includes a photomultiplier tube and a time-correlated single photon counter, which constitute an ultrafast signal detection system. The photomultiplier tube is used to amplify the excitation fluorescence signal and the time-correlated single photon counter is used to detect the fluorescence lifetime of tiny particles or cells.

[0097] The detection device and method provided in this application utilize fiber couplers to couple signals onto optical fibers, resulting in high system optical coupling efficiency. Furthermore, a fiber circulator is used to multiplex optical paths, enabling multi-point detection and simultaneous manipulation of tiny particles and fluorescence lifetime detection. Because the method utilizes a single multimode fiber loop to achieve multi-wavelength input, it overcomes the drawbacks of limited control flexibility associated with structures requiring multiple single-mode fibers or splicing multiple fibers, thereby improving operational efficiency and precision.

[0098] Other embodiments of the present invention will readily occur to those skilled in the art after considering the specification and practicing the invention disclosed herein. The present invention is intended to cover any variations, uses, or adaptations of the present invention that follow the general principles of the present invention and include common knowledge or customary techniques in the art not disclosed herein.

[0099] In the description of this specification, the description with reference to the terms "one embodiment", "some embodiments", "example", "specific example", or "some examples" means that the specific features, structures, materials or characteristics described in conjunction with the embodiment or example are included in at least one embodiment or example of the present application. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described can be combined in any one or N embodiments or examples in a suitable manner. In addition, those skilled in the art can combine and combine different embodiments or examples described in this specification and features of different embodiments or examples without contradiction.

[0100] It is understandable that the above embodiments are exemplary and should not be construed as limiting the present application. A person skilled in the art may change, modify, replace and modify the above embodiments within the scope of the present application.

Claims

1. A fiber optic tweezers fluorescence lifetime detection device, characterized in that: include: A light source assembly for emitting a first light beam and a second light beam with different wavelengths; a first objective lens and a second objective lens, respectively disposed on the optical paths of the first light beam and the second light beam, for collecting optical signals of the first light beam and the second light beam, and transmitting the collected optical signals to a first optical fiber coupler; The first optical fiber coupler is configured to couple the received optical signals corresponding to the first and second optical beams, and transmit the coupled optical signals to a connected optical fiber circulator; The optical fiber circulator is used to transmit the optical signal corresponding to the first light beam and the optical signal corresponding to the second light beam to the displacement control component; The displacement control component has an optical fiber end for capturing target microparticles, wherein the target microparticles are captured by a high-order focused beam formed by the optical signal corresponding to the first light beam at the optical fiber end, and the target microparticles generate an excitation fluorescence signal under the irradiation of the optical signal corresponding to the second light beam, and the excitation fluorescence signal is transmitted to the second optical fiber coupler via the optical fiber circulator; The detection component is connected to the output end of the second optical fiber coupler, and is used to receive the excitation fluorescence signal output by the second optical fiber coupler, and detect the number of photons of the excitation fluorescence signal to determine the fluorescence lifetime of the target tiny particle.

2. The optical fiber tweezers fluorescence lifetime detection device according to claim 1, characterized in that: The light source assembly includes: a light source, a parallel light transmission system, a frequency doubling crystal element and a half-reflecting half-mirror lens; The light source is used to emit a femtosecond laser beam; The parallel light transmission system includes a first lens and a second lens, wherein the first lens and the second lens form a 4f system, the frequency doubling crystal element is located between the first lens and the second lens, and the first lens, the frequency doubling crystal element and the second lens are sequentially arranged on the optical path of the femtosecond laser beam, and are used to parallelize and frequency-double the femtosecond laser beam to output a mixed beam containing two different wavelengths; The half-reflecting half-mirror is arranged on the optical path of the mixed light beam, and is used to separate the two optical signals of different wavelengths in the mixed light beam into two light beams by transmission and reflection respectively, to obtain a first light beam and a second light beam.

3. The optical fiber tweezers fluorescence lifetime detection device according to claim 2, characterized in that: The system is further provided with a reflector; The reflecting mirror is arranged on the optical path of the first light beam after passing through the half-reflecting half-mirror lens, and reflects the first light beam to the first objective lens.

4. The optical fiber tweezers fluorescence lifetime detection device according to claim 1, characterized in that: The displacement control component includes an optical fiber end, a microscopic imaging component and a position control platform; The microscopic imaging component is used to convert the optical signal corresponding to the first light beam output from the optical fiber circulator into an electrical signal to capture image information within the target area; The position control platform is used to control the movement of the optical fiber end in the target area so that the optical fiber end captures target tiny particles in the target area.

5. The optical fiber tweezers fluorescence lifetime detection device according to claim 4, characterized in that: The position control platform is a six-axis precision displacement platform, and the end face of the optical fiber end is a three-dimensional structure.

6. The optical fiber tweezers fluorescence lifetime detection device according to claim 1, characterized in that: The detection assembly includes: a photomultiplier tube and a time-correlated single photon counter; The photomultiplier tube is used to amplify the received excitation fluorescence signal; The time-correlated single-photon counter is used to measure the number of photons in the received excitation fluorescence signal, so as to determine the fluorescence lifetime of the target tiny particle according to the number of received photons.

7. The optical fiber tweezers fluorescence lifetime detection device according to claim 2, characterized in that: The wavelength of the femtosecond laser beam is 800 nm; the wavelength corresponding to the first beam is 800 nm, and the wavelength corresponding to the second beam is 400 nm.

8. The optical fiber tweezers fluorescence lifetime detection device according to claim 2, characterized in that: The focal length of the first lens and the second lens is 15 mm.

9. A method for detecting fluorescence lifetime using the optical fiber tweezers fluorescence lifetime detection device according to any one of claims 1 to 8, characterized in that: include: The first objective lens and the second objective lens respectively collect optical signals of the first light beam and the second light beam emitted by the light source assembly, and transmit the collected optical signals to the first optical fiber coupler; the first light beam and the second light beam have different wavelengths; The first optical fiber coupler couples the received optical signals corresponding to the first and second optical beams, and transmits the coupled optical signals to the displacement control component via the optical fiber circulator; The displacement control component uses the optical signal corresponding to the first light beam to capture the target tiny particles. The target tiny particles generate an excitation fluorescence signal under the irradiation of the optical signal corresponding to the second light beam. The excitation fluorescence signal is transmitted to the detection component via the optical fiber circulator and the second optical fiber coupler. The detection component is used to detect the number of photons of the excitation fluorescence signal to determine the fluorescence lifetime of the target tiny particles.

10. The detection method according to claim 9, characterized in that: Before the step of respectively collecting the light signals of the first light beam and the second light beam emitted by the light source assembly through the first objective lens and the second objective lens, the method further includes: The light source emits a femtosecond laser beam; The parallel light transmission system arranged on the optical path of the femtosecond laser beam parallelizes and frequency-doubles the femtosecond laser beam, and outputs a mixed beam containing two different wavelengths; A half-reflective half-mirror mirror arranged on the optical path of the mixed light beam separates the optical signals of two different wavelengths in the mixed light beam into two light beams to obtain a first light beam and a second light beam.

Images