A compact, low-cost and high sensitive LIF based on pinhole metal-capillary and direct laser-diode excitation

Highlights

• A compact and low-cost LIF based on pinhole metal-capillary (PMC) was proposed

• The background signal (noise) can be effectively suppressed in PMC

• The stringent requirement on laser beam-quality can be alleviated in PMC-LIF

• An overall ~480 fold improvement on SNR was realized by using PMC and laser diode

• A detection limit of 0.4 pM was realized for rhodamine B detection

Abstract

For laser-induced fluorescence (LIF), the SNR and related concentration detectability was severely limited by background signal (noise). Herein, an axial-excitation LIF based on pinhole metal-capillary (PMC) and direct laser-diode excitation was proposed. An overall ~480 fold improvement on SNR was realized, because the laser-sidewall interaction (LSI) and related background signal (noise) can be effectively suppressed in PMC by avoidance of laser leakage and laser contact with capillary sidewall. For rhodamine B detection without sample enrichment, a detection limit of concentration (DLC) of 0.4 pM was obtained, which is 2.5 fold lower than the DLC (1 pM) of commercial LIFs equipped with bulky and expensive Ar⁺ laser. For selective detection of Cu²⁺, a DLC of 5 pM was realized, which is more than 40 fold lower than that of previous fluorimetry detection with similar quantum-dot probe. The PMC-LIF features compact (16 × 4 × 4 cm³), low-cost and high concentration detectability.

Introduction

LIF is one of the most sensitive techniques for analysis of flowing sample in capillary [1], [2], [3], [4], [5], [6], [7], [8], [9], [10], [11], [12], [13], [14]. As shown in Fig. 1, according to the excitation-direction of capillary, there are mainly two kinds of LIFs, i.e., transverse-excitation of capillary (TE-LIF) [2], [3], [4], [5], [6], [7], [8], [9], axial-excitation along capillary (AE-LIF) [10], [11], [12], [13], [14]. For TE-LIF (e.g., confocal LIF), an ultra-low detection-limit of mass (DLM) can be realized (even a few molecules), owning to the attainable ultra-small detection-volume [2,[15], [16], [17]]. However, for some applications, such as food safety inspection and environmental monitor, the criterion of noxious or innoxious was based on concentration level rather than mass [18], [19], [20], [21], so low detection-limit of concentration (DLC) is also urgently required. Moreover, compactness and low-cost is also required for personal use.

As for improving SNR and related DLC, suppressing background (noise) becomes more important than simply enhancing signal, because the signal can be easily increased by increasing laser power or optical-path [9], [10]. Previous reports indicated that the background signal (noise) of TE-LIF was mainly resulted from laser sidewall-reflection, which can be considered as a kind of laser-sidewall interaction (LSI) [[8], [9],[13], [14],[22], [23], [24], [25], [26]]. Many noteworthy methods have been proposed to suppress the sidewall-reflection, such as (1) deviating laser reflection-direction from fluorescence collection by using deflected reflection [8], shifting focusing-point [9] or employing sheath-flow cell [14,[22], [23], [24]]; (2) reducing sidewall reflectivity by immersing into index-matched oil [25] or opening a slit (i.e., removing sidewall) at excitation point [26]; and (3) employing pulsed-excitation to separate longer-lived signal from background [23].

However, for pulsed-excitation, the apparatus is bulky, complicated and expensive [23]. As for continuous excitation, the sidewall reflection is still inevitable except removing the sidewall [[8], [9],22,[24], [25]]. Moreover, due to the stringent requirement on optical alignment and laser focusing, high beam-quality laser was required, such as bulky Ar⁺ laser [[22], [23],27] and solid state laser [2,5,24], which are more bulky and expensive than laser-diode. Currently, for detecting molecular dyes without sample enrichment, the DLCs are in a range of 1 ~ 100 pM with continuous Ar⁺ laser excitation [6,[11], [12],27], and the DLC can be improved to 0.089 pM with pulsed Ar⁺ laser excitation [23]. In comparison, by using direct (also continuous) laser-diode excitation, whose beam-quality is lower than that of Ar⁺ laser and solid state laser [28], [29], the DLCs are much higher, i.e., for molecular dye, most DLCs are higher than 400 pM [[3], [4],[30], [31]], and a few DLCs can reach 3 pM by using complicated sheath-flow cell [14].

As for AE-LIF (Fig. 1b), the signal can be enhanced as much as 100 ~ 3000 fold compared with TE-LIF, owing to the long optical-path (as long as meters) [32], [33], [34], [35], [36], [37], [38], [39]. However, the LSI and related background signal (noise) were also greatly enhanced, which makes the improvement on SNR very limited [34], [35]. For AE-LIF, the DLC (1.6 pM, 6 pM and 100 pM for molecular-dyes [10], [11], [12]) is still higher than that of TE-LIF. Moreover, the lowest DLC (1.6 pM) was realized by using ultra-long capillary (12 m) and special solvent (CS₂) [10].

To sum up, in order to obtain a DLC as low as 1 pM (for molecular dye), high beam-quality Ar⁺ laser is required [6,27]. Further improvement of DLC to ~0.1 pM can be realized by using complicated pulsed-excitation, which consists of Ar⁺ laser, electrooptic modulator, high-speed pulse generator, and lock-in amplifier, etc [23]. However, for direct (also continuous) laser-diode excitation, which is compact and low-cost, it is still a challenge to realize a DLC lower than 3 pM [14]. Thus, new method is needed to suppress the LSI in LIF (especially in AE-LIF).

Meanwhile, as one of essential heavy-metal, Cu²⁺ can function as catalyzing cofactor for ~20 kinds of enzymes and involve in bone formation, cellular respiration, etc [40], [41]. Therefore, several methods have been developed for detecting the concentration of Cu²⁺, such as colorimetry [42], electrochemistry [43] and fluorimetry [41,[44], [45], [46], [47], [48], [49]]. For fluorimetry, it features high sensitivity and selectivity, and the DLC is normally in the range of 0.1 nM ~ 6.9 nM by using quantum dot (QD) as fluorescent probe [41,[44], [45], [46], [47], [48], [49]].

In this paper, an AE-LIF based on pinhole metal-capillary (PMC) and direct laser-diode excitation was proposed (i.e., PMC-LIF). The PMC was constructed by placing a pinhole-mirror at the inlet of metal-capillary. An overall ~480 fold improvement on SNR was realized, because the LSI and related background signal (noise) can be most effectively suppressed in PMC by avoidance of laser leakage and laser contact with capillary sidewall. Moreover, a compact and low-cost PMC-LIF with a 7 cm long MC, direct laser-diode excitation, and axial close-coupling collection (Si photodetector) was fabricated. For RhB detection, a DLC of 0.4 pM was realized, which is 2.5 fold lower than that of commercial LIFs equipped with bulky Ar⁺ laser excitation. For selective detection of Cu²⁺, a DLC of 5 pM was realized, which is more than 40 fold lower than that of previous fluorimetry detection with similar QD probe.