Using an insulating fiber as the sampling probe and ionization substrate for ambient ionization-mass spectrometric analysis of volatile, semi-volatile, and polar analytes.

A sharp metal needle used as the ionization emitter in conventional atmospheric pressure chemical ionization (APCI) mass spectrometry (MS) is usually required for analyte ionization through corona discharge (i.e., gas discharge). Nevertheless, we herein demonstrate that an insulating fiber (tip diameter: 10-60 µm; length: ~ 1 cm) made of glass or bamboo can function as an APCI-like ionization emitter. Although no direct electric contact is made on the fiber, the ionization of volatiles and semi-volatiles occurs when the fiber is placed close (~ 1 mm) to the inlet of the mass spectrometer. No analyte ion signals can be observed without placing the insulating fiber in front of the mass spectrometer. The generation of ion species mainly relies on the electric field provided by the mass spectrometer. Presumably, owing to the high electric field provided by the mass spectrometer, the dielectric breakdown voltages of gas molecules in the air and the fiber are overcome, leading to the ionization of analytes in gas phase. In addition, the insulating fiber can function as a holder for sample solutions. Electrospray ionization-like processes derived from polar analytes such as amino acids, peptides, and proteins can readily occur when the insulating fiber deposited with a sample droplet is placed close to the inlet of the mass spectrometer. The feasibility of using the current approach for the detection of nonpolar and polar analytes from complex fetal bovine serum samples without tedious sample pretreatment is demonstrated in this work. The main advantage of using the suggested fiber is that the fiber can be used as the sampling probe to pick up samples and placed in front of a mass spectrometer for direct MS analysis. The application of using a robust, insulating, and disposable probe to pick up samples from real samples such as onion, honey, and pork samples followed by direct MS analysis is also demonstrated.

Functional magnetic nanoparticle-based affinity probe for MALDI mass spectrometric detection of ricin B.

The use of lactosylated Fe3O4 magnetic nanoparticles (MNP@LAC) has been explored as affinity probes against ricin B based on galactose-ricin B binding interactions. Lactose was bound onto the surface of aminated MNPs through the Maillard reaction. The enrichment of ricin B took ~1 h by incubating MNP@LAC with samples under shaking at room temperature, followed by magnetic isolation. The resultant MNP@LAC-ricin B conjugates were characterized by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). The limit of detection toward ricin B was ~3 nM by using the developed method. It was possible to detect the peptides derived from the tryptic digest of trace ricin B (~0.39 nM) enriched by the MNP@LAC probes followed by tryptic digestion and MALDI-MS analysis. The feasibility of using the developed method for detection of ricin B from complex white corn starch samples spiked with trace ricin B was demonstrated.

Using lactosylated cysteine functionalized gold nanoparticles as colorimetric sensing probes for rapid detection of the ricin B chain.

A new colorimetric method that can be used to rapidly detect toxic ricin is demonstrated. Lactosylated cysteine-functionalized gold nanoparticles (Au@LACY NPs) were prepared by a one-pot reaction and employed as optical probes for determination of ricin B chain. It is found that the Au@LACY NPs undergo aggregation in the presence of ricin B chain. This leads to surface plasmon coupling effects of the particles and a color change from red to blue, with absorption maxima at 519 and 670 nm, respectively. The feasibility of using the current approach for quantitative analysis of ricin B chain is also demonstrated. The calibration plot is generated by plotting the ratio of the absorbance at the wavelength of 634 to 518 nm versus the concentration of the ricin B chain. The spectrophotometric method has a ~29 pM (~ 0.91 ng·mL-1) detection limit, and the sample with the concentration of ~ 400 pM (~ 13 ng·mL-1) can be detected visually. Graphical abstractSchematic representation of using lactosylated cysteine capped gold nanoparticles (Au@LACY NPs) as colorimetric probes for the ricin B chain through surface plasmon coupling effects. Sample solution turns from red to blue in the presence of ricin B chain.

Tail Fiber Protein-Immobilized Magnetic Nanoparticle-Based Affinity Approaches for Detection of Acinetobacter baumannii.

Acinetobacter baumannii (A. baumannii) strains are common nosocomial pathogens that can cause infections and can easily become resistant to antibiotics. Thus, analytical methods that can be used to rapidly identify A. baumannii from complex samples should be developed. Tail fiber proteins derived from the tail fibers of bacteriophages can recognize specific bacterial surface polysaccharides. For example, recombinant tail proteins, such as TF2 and TF6 derived from the tail fibers of bacteriophages ϕAB2 and ϕAB6, can recognize A. baumannii clinical isolates M3237 and 54149, respectively. Thus, TF2 and TF6 can be used as probes to target specific A. baumannii strains. Generally, TF2 and TF6 are tagged with a hexahistidine (His6) for ease of purification. Given that His6 possesses specific affinity toward alumina through His6-Al chelation, TF2- and TF6-immobilized alumina-coated magnetic nanoparticles (Fe3O4@Al2O3 MNPs) were generated through chelation under microwave heating (power, 900 W) for 60 s in this study. The as-prepared TF2-Fe3O4@Al2O3 and TF6-Fe3O4@Al2O3 MNPs were used as affinity probes to trap trace A. baumannii M3237 and 54149, respectively, from sample solutions. Matrix-assisted laser desorption/ionization mass spectrometry capable of identifying bacteria on the basis of the obtained fingerprint mass spectra of intact bacteria was used as the detection tool. Results demonstrated that the current approach can be used to distinguish A. baumannii M3237 from A. baumannii 54149 by using TF2-Fe3O4@Al2O3 and TF6-Fe3O4@Al2O3 MNPs as affinity probes. Furthermore, the limits of detection of the current method for A. baumannii M3237 and 54149 are ∼105 and ∼104 cells mL-1, respectively. The feasibility of using the developed method to selectively detect A. baumannii M3237 and 54149 from complex serum samples was demonstrated.

Functionalized gold nanoparticles as affinity nanoprobes for multiple lectins.

Glycan-lectin interactions are commonly observed in nature. Analytical methods, which are used to detect lectins that rely on the use of glycan ligand-modified nanoprobes as affinity probes, have been developed. Most of the existing methods are focused on the use of synthetic glycan ligands. Nevertheless, naturally available glycoproteins, such as ovalbumin in chicken egg whites, are good sources for fabricating glycan-immobilized nanoprobes. In this study, we generated functionalized gold nanoparticles (Au NPs) from a one-pot reaction by reacting chicken egg white (cew) proteins with aqueous tetrachloroaurate. The generated Au@cew NPs are mainly encapsulated by ovalbumin, in which the surface is decorated by abundant hybrid mannose and Galß(1→4)GlcNAc-terminated glycan ligands. Thus, the generated Au@cew NPs containing hybrid mannose and Galß(1→4)GlcNAc have the capability to selectively bind with their corresponding lectins. Lectins including concanavalin A, banana lectin, and ricin B that have binding moieties toward specific sugars were used as the model samples. Our results showed that the generated AuNPs can be used as multiplex affinity probes for these model lectins. Lectins can be selectively released from the Au@cew NP-lectin conjugates by using specific sugars, such as mannose, glucose, and ß-lactose, as the releasing agents to release specific lectins from the conjugates. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) was used as the tool to characterize the released species from the nanoprobes. The limit of detection of these model lectins using the current approach was low (in nM). The feasibility of using the Au@cew NP-based affinity MALDI-MS to selectively detect specific lectins from complex samples was also demonstrated.

Selective Detection of Shiga-like Toxin 1 from Complex Samples Using Pigeon Ovalbumin Functionalized Gold Nanoparticles as Affinity Probes.

Escherichia coli O157:H7 is a foodborne pathogen. This bacterial strain can generate Shiga-like toxins (SLTs), which can cause serious sickness and even death. Thus, it is important to develop effective and sensitive methods that can be used to rapidly identify the presence of SLTs from complex samples. Pigeon egg white (PEW) contains abundant glycoproteins, including pigeon ovalbumin (POA) (∼60%). POA possesses Gal-α(1→4)-Gal-ß(1→4)-GlcNAc termini, which can recognize the B subunits in SLT type 1 (SLT-1B). Thus, POA is a suitable probe for trapping SLT-1B. In this work, we used PEW proteins as starting materials to react with aqueous tetrachloroauric acid for generation of PEW-protein-immobilized gold nanoparticles (AuNPs@PEW) via one-pot reactions. We demonstrated that the generated AuNPs@PEW were mainly dominated by POA-immobilized Au NPs. The as-prepared AuNPs@PEW were used as affinity probes to selectively probe SLT-1B from complex cell lysates derived from E. coli O157:H7. The selective trapping step can be completed within ∼90 s under microwave heating (power = 450 W) to enrich sufficient SLT-1B for matrix-assisted laser desorption/ionization (MALDI) mass spectrometric analysis. Furthermore, this approach can be used to detect SLT-1B at a concentration as low as ∼40 pM. The feasibility of using the proposed method to selectively detect SLT-1B from ham contaminated by E. coli O157:H7 was also demonstrated.

Detection of ricin by using gold nanoclusters functionalized with chicken egg white proteins as sensing probes.

Ricin produced from the castor oil plant, Ricinus communis, is a well-known toxin. The toxin comprises A and B chains. Ricin A chain can cause toxicity by inhibiting protein synthesis, and ricin B can bind to the galactose ligand on the cell membrane of host cells. Inhalation or ingestion of ricin may even lead to death. Therefore, rapid and convenient sensing methods for detecting ricin in suspicious samples must be developed. In this study, we generated protein encapsulated gold nanoclusters (AuNCs@ew) with bright photoluminescence by using chicken egg white proteins as starting materials to react with aqueous tetrachloroaurate. The generated nanoclusters, which were mainly composed of chicken ovalbumin-encapsulated AuNCs, can recognize ricin B because of the presence of Galß(1→4)GlcNAc ligands on chicken ovalbumin. The generated conjugates of AuNCs@ew and ricin B were heavy and readily settled down under centrifugation (13,000rpm, 60min). Thus, bright spots resulting from the conjugates at the bottom of the sample vials were easily visualized by the naked eye under ultraviolet light illumination. The limit of detection (LOD) was ~4.6µM. The LOD was reduced to ~400nM when fluorescence spectroscopy was used as the detection tool, while the LOD can be further improved to ~7.8nM when using matrix-assisted laser desorption/ionization mass spectrometry as the detection method. We also demonstrated the feasibility of using the proposed approach to selectively detect ricin B chain in complex samples.

Using protein-encapsulated gold nanoclusters as photoluminescent sensing probes for biomolecules.

In this study, we generated gold nanoclusters (AuNCs) using inexpensive chicken egg white proteins (AuNCs@ew) as reagents. AuNCs@ew were generated by reacting aqueous tetrachloroauric acid with diluted chicken egg white under microwave heating (90W) through subsequent heating cycles (5 min/cycle). Within 10 cycles, red photoluminescent AuNCs@ew with maximum emission wavelength at ~640 nm (λex=370 nm) were obtained. The quantum yield of the as-generated AuNCs was ~6.6%. The intact and the tryptic digest of AuNCs@ew were characterized by mass spectrometry. The results showed that the AuNCs@ew were mainly derived from ovalbumin, i.e., the major protein in egg white, encapsulated AuNCs. The AuNCs@ew also has the common features found in AuNCs@protein, which is sensitive to the presence of heavy metal ions such as Cu(2+). The photoluminescence of the AuNCs@ew was quenched with the addition of Cu(2+). Furthermore, the photoluminescence of the quenched AuNCs@ew can be restored in the presence of the molecules containing phosphate functional groups because of the strong binding affinity between Cu(2+) and phosphates. We used the AuNCs@ew-Cu(2+) conjugates as switch-on sensing probes for the detection of phosphate containing metabolites such as adenosine-5'-triphosphate (ATP) and pyrophosphate (PPi). The results showed that the photoluminescence of the sensing probes increased as the concentration of the phosphate-containing molecules in the sample solution increased. The limits of detection achieved using the AuNCs@ew-Cu(2+) for ATP and PPi were ~19 and ~5 µM, respectively. Additionally, we also demonstrated the feasibility of using the AuNCs@ew as the sensing probes for lectins such as concanavalin A (Con A) based on the molecular recognitions between the glycan ligands on the AuNCs@ew and glycan binding sites on Con A.