1. Introduction

Single cell analysis has driven the development of particle trapping techniques, exploiting various physical mechanisms, including optical trapping [1], acoustic trapping [2] and dielectrophoresis [3]. More recently, optical fibers have been used for cell trapping, where a force is created by optical means. For example, fiber-based optical tweezers [4–7] and optical stretchers [8,9] were demonstrated. In addition, cell trapping by photothermal effects [10] and evanescent fields near the fiber tip [11] were exploited. One main motivation for the use of fibers is that the potential combination of cell trapping and optical detection in a single device may lead to advanced catheters, with fiber probes that can extend the functionalities of lab-on-a-chip to in-vivo applications. Various other fields could also benefit from a simple trapping technique that simultaneously enables optical analysis. For example, micron-size insoluble drug particles play an important role in pharmaceutical research [12], and the optical characterization of trapped particles of a given size could be valuable for inspection and process control. One could also envisage the characterization of trapped particles in a laser-induced breakdown spectroscopy system [13,14]. In the present paper, an optical fiber probe is presented that can simultaneously trap, excite and detect fluorescently labeled micro-particles in a liquid medium. In contrast to previous work, the trapping mechanism is based on the simple use of fluidic forces and liquid aspiration through a micro-structured optical fiber with a constricted tip. This allows for simultaneous particle trapping and optical analysis without the use of an external microscope.

2. Optical fiber probe

2.1 Optical fiber tip for fluidic trapping

The optical fiber probe presented in this work makes use of the micro-structured optical fiber shown in Fig. 1(a). The fiber has an 8-µm diameter Ge-doped core and 5 holes along the cladding of diameter 20 µm. The outer diameter of the fiber is 125 µm. Laser light can be guided either in the 8-µm core seen in the photograph shown in Fig. 1(a, top), or in the entire central area delimited by the 5 holes, which has a diameter of 18 µm, as seen in Fig. 1(a, bottom). A schematic of the fiber probe is depicted in Fig. 1(b). A capillary tip with an opening smaller than the size of the particles of interest is collapsed onto the 5-holes fiber, aligned coaxially with its core. Liquid flows into the opening of the capillary tip when the probe is immersed in a suspension of particles and negative differential pressure is applied to the remote end of the fiber. This causes a suspended particle to be dragged towards the opening, where it is trapped. Applying positive differential pressure to the remote end releases the trapped particle. Laser light is coupled to the 8-µm core and travels towards the fiber tip for particle excitation. The optical signal from the particle is collected and guided back in the wider 18-µm waveguiding region delimited by the 5 holes towards a detector. In this work, 10-µm diameter fluorescently labeled particles were used as a proof-of-principle demonstration.

 

Fig. 1 (a) Photographs of the illuminated 5-holes micro-structed optical fiber. Excitation laser light is confined in the 8-µm core (top) and collected fluorescence light in the 18-µm area delimited by the holes (bottom). (b) Schematic of the fiber probe for fluidic trapping and optical detection. The dashed black arrows indicate fluid flow and the red arrows the fluorescence light. Silica glass is represented in blue. (c) Microscope images of the fiber probe fabrication (left) showing the spliced capillaries (top) and the etched 5-holes fiber (bottom), and final component immersed in water after partial hole collapse (right).

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The fiber probe is fabricated using a Vytran GPX-3000 glass processing station. A 90/125 µm (inner/outer) diameter capillary is spliced to a 25/125 µm diameter capillary which is cut to a length of 50 µm, as shown in Fig. 1(c, 1). A 5-mm section of the 5-holes fiber is etched by using hydrofluoric acid to 80 µm and cleaved. (Fig. 1(c, 2)). The etched section is introduced inside the 90/125 µm capillary which is cut at 3 mm to the left side. The arrangement is fixed by heating the 90 µm capillary, collapsing it onto the 5-holes fiber, as shown schematically in Fig. 1(b). A microscope image of the final component is shown in Fig. 1(c, right). In order to trap 10-µm particles, the inner diameter of capillary tip is reduced from 25 µm hole to 8 µm by partially collapsing the hole with heat. Fine adjustment (submicron) of the hole-size can be achieved with successive low-current electrical discharges.

The ability to perform sensitive optical analysis of trapped particles is limited by the finite distance d between the particle and the light collecting 5-hole fiber end, Fig. 1(a). For the component used here, d = 100 µm. The normalized collection efficiency (i.e. fraction collected of the total light emitted by a particle) defined as η = Ω/4π [15,16] for an illumination solid-angle Ω and a particle with isotropic light emission is discussed in relation to Fig. 2.

 

Fig. 2 Simulation of collection efficiency η as a function of distance d between trapped particles and fiber end-face. Solid blue and dashed orange curves correspond to the 5-holes fiber with NA 0.6 and a fiber with NA 0.2, respectively. The diameter of the collection region is 18 µm for both fibers. The black point corresponds to the parameters of the fiber probe shown in Fig. 1 (d = 100 µm and η = 0.002).

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The 18-µm diameter region that collects light has a numerical aperture NA = (n²_silica – n²_holes)^1/2 = 0.6 when the holes are filled with water (n_holes = 1.33, n_silica = 1.46). In principle, if d could be reduced arbitrarily, this high NA would enable collection and guidance of light within a maximum incidence angle θ_max = sin⁻¹(NA/n_holes) = 27°. In this case, the collection efficiency is limited by the solid angle defined by the fiber NA, Ω_NA = 2π(1-cos θ_max), and saturates at 5.4% (η = 0.054). This value compares favorably with the fraction 0.5% that would be collected with a fiber with NA 0.2 (typical value for a multimode fiber) and same core diameter. In the case when d is sufficiently large (d>17.7 µm for NA 0.6), the maximum collection angle θ is limited by the diameter of the collection region (18 µm) to θ = tan⁻¹(18 µm/2d) < θ_max, and the collection efficiency η decreases, since Ω = 2π(1-cosθ). Figure 2 shows the collection efficiency η calculated as a function of d for an 18-µm diameter core and a numerical aperture 0.6 and 0.2. The black point in the graph corresponds to the parameters of the component shown in Fig. 1 (d = 100 µm, η = 0.002). Although the normalized collection efficiency here is relatively low, it is sufficient for sensitive detection of particles. Figure 2 also shows how the collection efficiency can be increased, if needed, by reducing d. However, this was not found necessary in the present experiment.

2.2 Liquid and light combiner

A liquid-light combiner is built to simultaneously allow for light coupling into and out of the 5-holes fiber and for controlling the pressure in the holes for particle trapping [17]. A schematic of this component is shown in Fig. 3. A double-clad optical fiber is used, with outer diameter of 125 µm, a large outer core of 105 µm and a smaller inner core of 8 µm, Fig. 3(inset 1). The fiber is tapered down to a diameter of 25 µm, Fig. 3(inset 2), by simultaneously pulling and heating it in the Vytran GPX-3000 processing station. The 25-µm taper is then spliced to the central region of the 5-holes fiber without blocking the holes, Fig. 3(inset 3). The tapered region and the splice are placed inside a large silica housing capillary of 0.8/1 mm diameter. One end of a piece of 90/125 µm capillary is also inserted in the housing capillary to provide access to the holes, enabling particle trapping and releasing by applying negative or positive differential pressure, respectively. The housing capillary is sealed on both sides using UV-curing glue. Excitation light is coupled to the 8-µm core of the double-clad fiber and is guided through the down-taper to the 5-holes fiber and the fiber probe. Emission from trapped particles collected by the area enclosed by the 5 holes is guided back through the taper (seen by the emission light as an up-taper) to the 105-µm core of the double-clad fiber and towards the optical system for detection.

 

Fig. 3 Schematic of the liquid-light combiner for coupling light to and from the 5-holes fiber core and accessing the holes for trapping and releasing particles. Microscope images are included, which illustrate different parts of the component.

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The fiber taper also works as a numerical aperture converter [18]. As mentioned above, emission light reaching the fiber tip at angles up to 27° could be collected and guided back due to the high NA (0.6) of the 18-µm collection region. As the collected light travels back through the fiber up-taper, it experiences a trade between light spot diameter and numerical aperture. The numerical aperture after the up-taper NA_out can be calculated by

2.3 Experimental setup

The experimental setup is illustrated in Fig. 4. A solid-state laser emitting at 491 nm wavelength with a power of 2 mW (Cobolt Inc) is used for particle excitation. The excitation beam is reflected by a dichroic mirror (Thorlabs DMLP505) and coupled to the 8-µm core of the double-clad fiber (Thorlabs DCF13) by using a 10x objective lens. The 491-nm laser beam is guided in the double-clad fiber towards the liquid-light combiner and subsequently to the 5-holes fiber probe. The collected emission light, which travels back in the 105-µm core, passes through the dichroic mirror and is analyzed in a spectrometer (Ocean Optics QE65000). A bandpass filter is used to remove the undesired excitation light. A syringe is connected to the input capillary of the liquid-light combiner to control the differential pressure used for trapping. Before operation, the fiber component is filled with water to prevent creation of bubbles. Subsequently, the fiber probe is introduced in a diluted suspension containing a mixture of 10-µm fluorescent particles with emission in the green spectral region (ThermoScientific, max. excitation at 468 nm and max. emission 508 nm) and in the yellow (Beckman Coulter max. excitation at 488, emission 515-660 nm)

 

Fig. 4 Lay-out of the experimental setup. (A) corresponds to the liquid-light combiner described in section 2.2.

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3. Results

Suction with the syringe makes water flow into the fiber probe, dragging particles towards the opening, as shown in Fig. 5(a). Small particles flow freely into the probe. A sufficiently large particle reaching the constrained opening blocks it and stops the flow of the liquid medium, Fig. 5(b). The negative differential pressure causes the liquid columns filling the holes of the 5-holes fiber to empty, as can be seen in the two dark regions of Fig. 5(b). The excitation beam illuminates the trapped particle and its spectrum is measured and recorded with an integration time of 100 ms. Fig. 5(c) and 5(d) show a part of the emission spectrum (from 530 nm to 650nm) for the different types of particles present in the suspension. These results indicate that particles can be readily identified and optically characterized by the fiber probe. Finally, the trapped particle is released by positive differential pressure. A video showing continuous trapping, excitation, detection, and releasing of particles can be found in Visualization 1.

 

Fig. 5 Simultaneous trapping and optical detection of 10-µm fluorescent particles (Visualization 1). (a) Particle flowing towards the fiber probe. (b) Particle trapped in the opening of the fiber probe. (c,d) Excitation (left) and recorded spectrum (left) of a green fluorescence particle (c) and a yellow fluorescent particle (d). A video can be found in Visualization 1.

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4. Discussion and conclusion

The device presented here integrates micropipette aspiration [20] for cell trapping and optical detection in a single-ended optical fiber probe, allowing for optical studies to be carried out without relying on an external microscope. This opens the possibility of extending the trapping capability of micropipettes to the simultaneous identification of cells through their optical signature in in-vivo settings. By exploiting label-free techniques such as auto-fluorescence and Raman spectroscopy, the probe could be incorporated readily into a smart catheter for in-vivo studies. Additionally, for in-vitro applications, this device could be used as a dip-probe for rapid characterization of tagged cells, which may be applicable, for instance, in circulating tumor cell isolation [21] or selective collection of cells in biological assays [22,23]. The trapping mechanism used here, based on fluid suction, is independent of particle concentration, since the liquid medium is being constantly sucked into the fiber. This in an advantage compared to optical tweezers-based approaches that rely on particles passing near the fiber end. In addition, the present fiber probe avoids high power illumination typically required for trapping with optical-forces. This eliminates the risk of burning and damaging healthy tissue due to excessive light levels, thus making its implementation more likely under in-vivo conditions. Furthermore, the fluidic trapping mechanism is size-selective and, by properly choosing the inner diameter of the constricted capillary tip, particles as small as 1 µm, such as bacteria, could be potentially trapped. In addition, morphology based cell analysis could be performed. For instance, deformability of a trapped cell could be characterized by accurately controlling the suction force [20], while carrying out optical measurements. This could enable retrieving mechanical properties of cells, which has been proven as an efficient method for label-free detection of cancer [24,25]. Finally, the device demonstrated here could also be used in other fields, for example, optical analysis of micron-size particulates for pharmaceutical research [12] or water quality testing [10].