Sample Preparation Free Mass Spectrometry Using Laser-Assisted Rapid Evaporative Ionization Mass Spectrometry: Applications to Microbiology, Metabolic Biofluid Phenotyping, and Food Authenticity.

Mass spectrometry has established itself as a powerful tool in the chemical, biological, medical, environmental, and agricultural fields. However, experimental approaches and potential application areas have been limited by a traditional reliance on sample preparation, extraction, and chromatographic separation. Ambient ionization mass spectrometry methods have addressed this challenge but are still somewhat restricted in requirements for sample manipulation to make it suitable for analysis. These limitations are particularly restrictive in view of the move toward high-throughput and automated analytical workflows. To address this, we present what we consider to be the first automated sample-preparation-free mass spectrometry platform utilizing a carbon dioxide (CO2) laser for sample thermal desorption linked to the rapid evaporative ionization mass spectrometry (LA-REIMS) methodology. We show that the pulsatile operation of the CO2 laser is the primary factor in achieving high signal-to-noise ratios. We further show that the LA-REIMS automated platform is suited to the analysis of three diverse biological materials within different application areas. First, clinical microbiology isolates were classified to species level with an accuracy of 97.2%, the highest accuracy reported in current literature. Second, fecal samples from a type 2 diabetes mellitus cohort were analyzed with LA-REIMS, which allowed tentative identification of biomarkers which are potentially associated with disease pathogenesis and a disease classification accuracy of 94%. Finally, we showed the ability of the LA-REIMS system to detect instances of adulteration of cooking oil and determine the geographical area of production of three protected olive oil products with 100% classification accuracy.

Critical evaluation of ambient mass spectrometry coupled with chemometrics for the early detection of adulteration scenarios in Origanum vulgare L.

Nowadays, most of the screening methods in food manufacturing are based on spectroscopic techniques. Ambient Mass Spectrometry is a relatively new field of analytical chemistry which has proven to offer similar speed and ease-of-use when compared to other fingerprinting techniques, alongside the advantages of good selectivity, sensitivity and chemical information. Numerous applications have been explored in food authenticity, based either on the target detection of adulteration markers or, less frequently, on the development of multivariate classification models. The aim of the present work was to evaluate and compare the capabilities of Direct Analysis in Real Time (DART) and Atmospheric Solid Analysis Probe (ASAP) Mass Spectrometry (MS) for the high-throughput authenticity screening of commercial herbs and spices products. The gross addition of bulking material to dried Mediterranean oregano was taken as case study. First, a pilot sample set, constituted by authentic dried oregano, olive leaves (a frequently reported adulterant) and mixtures thereof at different levels (i.e. 10, 20, 30 and 50% w/w) was used. Each sample was fingerprinted by both ambient-MS techniques. After appropriate pre-processing, the whole mass spectra were used for the subsequent multivariate data analysis. Soft Independent Modelling of Class Analogy was adopted as classification algorithm and the model was challenged with both new authentic oregano and in-house prepared blends. To the best of our knowledge, this is the first report of DART-MS and ASAP-MS used in full scan mode and coupled to chemometric modelling as rapid fingerprinting approach for food authentication. Although both the techniques provided satisfactory results, ASAP-MS clearly showed greater potential, leading to reproducible, diagnostic feature-rich mass spectra. For this reason, ASAP-MS was further tested under a more convoluted scenario, where the training and validation sets were enlarged with additional authentic oregano samples and a wider range of adulterant species, respectively. Overall good results were achieved, with 93% model predictive accuracy, and screening detection capability estimated between 5-20% (w/w) addition, depending on the adulterant considered with the only exception of majorana. Investigation of Q residuals could highlight the statistically-relevant chemical markers which could be tentatively annotated by coupling the ASAP probe with a high resolution mass analyser. The results from the validation study confirmed the great potential of ASAP-MS in combination with chemometrics as fast MS-based screening solution and demonstrated its feasibility for classification model building.

Making complex measurements of meat composition fast: Application of rapid evaporative ionisation mass spectrometry to measuring meat quality and fraud.

Increasing demands are being placed on meat producers to verify more about their product with regards to safety, quality and authenticity. There are many methods that can detect aspects of these parameters in meat, yet most are too slow to keep up with the demands of modern meat processing plants and supply chains. A new technology, Rapid Evaporative Ionisation Mass Spectrometry (REIMS), has the potential to bridge the gap between advanced laboratory measurements and technology that can screen for quality, safety and authenticity parameters in a single measurement. Analysis with REIMS generates a detailed mass spectral fingerprint representative of a meat sample without the need for sample processing. REIMS has successfully been used to detect species fraud, detect use of hormones in meat animals, monitor meat processing and to detect off flavours such as boar taint. The aim of this review is to summarize these and other applications to highlight the potential of REIMS for meat analysis. Sampling methods and important considerations for data analysis are discussed as well as limitations of the technology and remaining challenges for practical adoption.

Interlaboratory and Interplatform Study of Steroids Collision Cross Section by Traveling Wave Ion Mobility Spectrometry.

Collision cross section (CCS) databases based on single-laboratory measurements must be cross-validated to extend their use in peak annotation. This work addresses the validation of the first comprehensive TWCCSN2 database for steroids. First, its long-term robustness was evaluated (i.e., a year and a half after database generation; Synapt G2-S instrument; bias within ±1.0% for 157 ions, 95.7% of the total ions). It was further cross-validated by three external laboratories, including two different TWIMS platforms (i.e., Synapt G2-Si and two Vion IMS QToF; bias within the threshold of ±2.0% for 98.8, 79.9, and 94.0% of the total ions detected by each instrument, respectively). Finally, a cross-laboratory TWCCSN2 database was built for 87 steroids (142 ions). The cross-laboratory database consists of average TWCCSN2 values obtained by the four TWIMS instruments in triplicate measurements. In general, lower deviations were observed between TWCCSN2 measurements and reference values when the cross-laboratory database was applied as a reference instead of the single-laboratory database. Relative standard deviations below 1.5% were observed for interlaboratory measurements (<1.0% for 85.2% of ions) and bias between average values and TWCCSN2 measurements was within the range of ±1.5% for 96.8% of all cases. In the context of this interlaboratory study, this threshold was also suitable for TWCCSN2 measurements of steroid metabolites in calf urine. Greater deviations were observed for steroid sulfates in complex urine samples of adult bovines, showing a slight matrix effect. The implementation of a scoring system for the application of the CCS descriptor in peak annotation is also discussed.

Rapid evaporative ionization mass spectrometry coupled with an electrosurgical knife for the rapid identification of Mediterranean Sea species.

The topic of food analysis and safety has attracted increasing interest in recent decades owing to recent scandals concerning fraudulent activities (mislabeling, sophistication, adulteration, etc.) that can undermine human health. Among them, seafood fraud has probably the strongest relationship with food safety, an activity that goes beyond economic interests. This article explores the capabilities of an innovative instrumental setup, called the "iKnife," as a powerful tool in this specific research area, where until now genomics and proteomics have been the workhorses in analytical approaches. iKnife, which means "intelligent knife," is the name of a recent technology based on rapid evaporative ionization mass spectrometry (REIMS). REIMS is an emerging technique able to characterize different samples rapidly, affording a comprehensive profile usable as a fingerprint, without the need for preliminary extraction or cleanup procedures. In detail, a REIMS source is coupled to a high-resolution tandem mass spectrometer; such coupling allows one to maximize the amount of information (discriminant features) collected for a single analysis, as well as to focus on target analytes to achieve enhanced sensitivity and selectivity. A database was created from 18 marine species typical of the Mediterranean Sea, all caught in the very small area of the Strait of Messina, and reliable identification was achieved for each species with confidence higher than 99%. One big model and three submodels were built by principal component analysis and linear discriminant analysis for unambiguous key variable identification within each class (e.g., Cephalopoda), order (e.g., Perciformes), or family (e.g., Carangidae). Graphical abstract.

Investigations into the performance of travelling wave enabled conventional and cyclic ion mobility systems to characterise protomers of fluoroquinolone antibiotic residues.

RATIONALE: Fluoroquinolones (FLQs) have been shown to form protomers with distinctive fragment profiles. Experimental parameters affect protomer formation, impacting observed conventional tandem mass spectrometric (MS/MS) dissociation and multiple reaction monitoring (MRM) transition reproducibility. Collision cross section (CCS) measurement can provide an additional identification metric and improved ion mobility (IM) separation strategies could provide further understanding of fluctuations in fragmentation when using electrospray ionisation (ESI). METHODS: Porcine muscle tissue was fortified with nine fluoroquinolone antibiotics. Extracts were cleaned using QuEChERS dispersive extraction. Separation was achieved via ultra-high-performance liquid chromatography (UHPLC) and analysis performed using positive ion ESI coupled with linear T-wave IM (N2 and CO2 drift gas) and cyclic IM-MS (calibrated to perform accurate mass and CCS measurement). RESULTS: IM-resolved protomeric species have been observed for nine FLQs (uniquely three for danofloxacin). Long-term reproducibility and cross-platform T-wave/cIM studies have demonstrated CCS metric errors <1.5% when compared with a FLQ protomer reference CCS library. When comparing FLQ protomer separation using a standard, linear T-wave IM separator (N2 /CO2 ) and using a high-resolution cyclic T-wave device (N2 ), protomer peak-to-peak resolution ranged between Rs = 1 to Rs = 6 for the IM strategies utilised. CONCLUSIONS: CCS is a reliable cross platform metric; specific FLQ CCS identification fingerprints have been produced, illustrating the potential to compliment MS/MS specificity or provide an alternative identification metric. Using cIM there is opportunity to correlate the erratic nature of protomer formation with the analytical conditions used and to gain further understanding of ionisation/dissociation mechanisms taking place during routine analyses.

Rapid evaporative ionisation mass spectrometry and chemometrics for high-throughput screening of growth promoters in meat producing animals.

In a proof of concept perspective, Rapid Evaporative Ionisation Mass Spectrometry (REIMS) was explored for the direct analysis of meat samples from ß-agonist treated livestock. In this context, the combination of REIMS with untargeted metabolomics was investigated to identify carcasses from treated animals on the basis of a modification of indirect metabolites profile. The REIMS analysis generated specific lipid profiles which enabled the differentiation of meat samples collected from pigs treated with ractopamine via their feeding regime. Furthermore, the strategy was found successful when tested on different muscle types (loin, shoulder and thigh), which further expands its applicability. Classification performances were greater than 95% accurate which fully answers requirements of a screening strategy. This research indicates that REIMS implemented in an untargeted-metabolomics workflow can be considered as a high-throughput and accurate strategy for real-time meat classification in relation to ractopamine (and wider ß-agonists) treatment in pig production. This approach may subsequently be implemented as a rapid screening test, at the slaughterhouse or at border inspection points, to detect such practice.

A real time metabolomic profiling approach to detecting fish fraud using rapid evaporative ionisation mass spectrometry.

INTRODUCTION: Fish fraud detection is mainly carried out using a genomic profiling approach requiring long and complex sample preparations and assay running times. Rapid evaporative ionisation mass spectrometry (REIMS) can circumvent these issues without sacrificing a loss in the quality of results. OBJECTIVES: To demonstrate that REIMS can be used as a fast profiling technique capable of achieving accurate species identification without the need for any sample preparation. Additionally, we wanted to demonstrate that other aspects of fish fraud other than speciation are detectable using REIMS. METHODS: 478 samples of five different white fish species were subjected to REIMS analysis using an electrosurgical knife. Each sample was cut 8-12 times with each one lasting 3-5 s and chemometric models were generated based on the mass range m/z 600-950 of each sample. RESULTS: The identification of 99 validation samples provided a 98.99% correct classification in which species identification was obtained near-instantaneously (≈ 2 s) unlike any other form of food fraud analysis. Significant time comparisons between REIMS and polymerase chain reaction (PCR) were observed when analysing 6 mislabelled samples demonstrating how REIMS can be used as a complimentary technique to detect fish fraud. Additionally, we have demonstrated that the catch method of fish products is capable of detection using REIMS, a concept never previously reported. CONCLUSIONS: REIMS has been proven to be an innovative technique to help aid the detection of fish fraud and has the potential to be utilised by fisheries to conduct their own quality control (QC) checks for fast accurate results.

Prediction of Collision Cross-Section Values for Small Molecules: Application to Pesticide Residue Analysis.

The use of collision cross-section (CCS) values obtained by ion mobility high-resolution mass spectrometry has added a third dimension (alongside retention time and exact mass) to aid in the identification of compounds. However, its utility is limited by the number of experimental CCS values currently available. This work demonstrates the potential of artificial neural networks (ANNs) for the prediction of CCS values of pesticides. The predictor, based on eight software-chosen molecular descriptors, was optimized using CCS values of 205 small molecules and validated using a set of 131 pesticides. The relative error was within 6% for 95% of all CCS values for protonated molecules, resulting in a median relative error less than 2%. In order to demonstrate the potential of CCS prediction, the strategy was applied to spinach samples. It notably improved the confidence in the tentative identification of suspect and nontarget pesticides.

Rapid evaporative ionization mass spectrometry for high-throughput screening in food analysis: The case of boar taint.

Boar taint is a contemporary off-odor present in meat of uncastrated male pigs. As European Member States intend to abandon surgical castration of pigs by 2018, this off-odor has gained a lot of research interest. In this study, rapid evaporative ionization mass spectrometry (REIMS) was explored for the rapid detection of boar taint in neck fat. Untargeted screening of samples (n=150) enabled discrimination between sow, tainted and untainted boars. The obtained OPLS-DA models showed excellent classification accuracy, i.e. 99% and 100% for sow and boar samples or solely boar samples, respectively. Furthermore, the obtained models demonstrated excellent validation characteristics (R2(Y)=0.872-0.969; Q2(Y)=0.756-0.917), which were confirmed by CV-ANOVA (p<0.001) and permutation testing. In conclusion, in this work for the first time highly accurate and high-throughput (<10s) classification of tainted and untainted boar samples was achieved, rendering REIMS a promising technique for predictive modelling in food safety and quality applications.

Identification of the Species of Origin for Meat Products by Rapid Evaporative Ionization Mass Spectrometry.

Increasingly abundant food fraud cases have brought food authenticity and safety into major focus. This study presents a fast and effective way to identify meat products using rapid evaporative ionization mass spectrometry (REIMS). The experimental setup was demonstrated to be able to record a mass spectrometric profile of meat specimens in a time frame of <5 s. A multivariate statistical algorithm was developed and successfully tested for the identification of animal tissue with different anatomical origin, breed, and species with 100% accuracy at species and 97% accuracy at breed level. Detection of the presence of meat originating from a different species (horse, cattle, and venison) has also been demonstrated with high accuracy using mixed patties with a 5% detection limit. REIMS technology was found to be a promising tool in food safety applications providing a reliable and simple method for the rapid characterization of food products.

Use of atmospheric pressure solids analysis probe time-of-flight mass spectrometry to screen for plasticisers in gaskets used in contact with foods.

RATIONALE: Plasticisers are used in the PVC gaskets of metal closures on glass jars and bottles used for foods and beverages. They may migrate and so contaminate the packed foodstuff. The plasticisers are present in a high proportion and are often a complex mixture of substances leading to time-consuming analytical methodologies. This work describes a rapid screening method to identify the plasticisers used. METHODS: Analysis was carried out by direct sampling of the gaskets using atmospheric pressure solids analysis probe (ASAP) with time-of-flight (TOF) mass spectrometry (MS) using a SYNAPT G2 HDMS system. The accurate mass information collected was then compared to a user-prepared database of plasticisers to aid identification. RESULTS: The rapid identification approach was shown to be successful for 24 gasket samples previously analysed by alternative more lengthy gas chromatographic (GC) methods. Quantification by dissolution followed by standard addition was also demonstrated to be reliable. CONCLUSIONS: The ASAP-TOFMS method is a useful technique for rapidly screening gaskets for the presence of plasticisers. It can be used to identify specific gaskets deserving of further quantitative analysis by chromatographic methods, saving time and money by avoiding unnecessary analyses. Copyright © 2015 John Wiley & Sons, Ltd.

Evaluation and validation of an accurate mass screening method for the analysis of pesticides in fruits and vegetables using liquid chromatography-quadrupole-time of flight-mass spectrometry with automated detection.

This study reports the development and validation of a screening method for the detection of pesticides in 11 different fruit and vegetable commodities. The method was based on ultra performance liquid chromatography-quadrupole-time of flight-mass spectrometry (UPLC-QTOF-MS). The objective was to validate the method in accordance with the SANCO guidance document (12571/2013) on analytical quality control and validation procedures for pesticide residues analysis in food and feed. Samples were spiked with 199 pesticides, each at two different concentrations (0.01 and 0.05 mg kg(-1)) and extracted using the QuEChERS approach. Extracts were analysed by UPLC-QTOF-MS using generic acquisition parameters. Automated detection and data filtering were performed using the UNIFI™ software and the peaks detected evaluated against a proprietary scientific library containing information for 504 pesticides. The results obtained using different data processing parameters were evaluated for 4378 pesticide/commodities combinations at 0.01 and 0.05 mg kg(-1). Using mass accuracy (± 5 ppm) with retention time (± 0.2 min) and a low response threshold (100 counts) the validated Screening Detection Limits (SDLs) were 0.01 mg kg(-1) and 0.05 mg kg(-1) for 57% and 79% of the compounds tested, respectively, with an average of 10 false detects per sample analysis. Excluding the most complex matrices (onion and leek) the detection rates increased to 69% and 87%, respectively. The use of additional parameters such as isotopic pattern and fragmentation information further reduced the number of false detects but compromised the detection rates, particularly at lower residue concentrations. The challenges associated with the validation and subsequent implementation of a pesticide multi-residue screening method are also discussed.

Toggled RNA aptamers against aminoglycosides allowing facile detection of antibiotics using gold nanoparticle assays.

We have used systematic evolution of ligands by exponential enrichment (SELEX) to isolate RNA aptamers against aminoglycoside antibiotics. The SELEX rounds were toggled against four pairs of aminoglycosides with the goal of isolating reagents that recognize conserved structural features. The resulting aptamers bind both of their selection targets with nanomolar affinities. They also bind the less structurally related targets, although they show clear specificity for this class of antibiotics. We show that this lack of aminoglycoside specificity is a common property of aptamers previously selected against single compounds and described as "specific". Broad target specificity aptamers would be ideal for sensors detecting the entire class of aminoglycosides. We have used ligand-induced aggregation of gold-nanoparticles coated with our aptamers as a rapid and sensitive assay for these compounds. In contrast to DNA aptamers, unmodified RNA aptamers cannot be used as the recognition ligand in this assay, whereas 2'-fluoro-pyrimidine derivatives work reliably. We discuss the possible application of these reagents as sensors for drug residues and the challenges for understanding the structural basis of aminoglycoside-aptamer recognition highlighted by the SELEX results.

Development and validation of a fast monoclonal based disequilibrium enzyme-linked immunosorbent assay for the detection of triphenylmethane dyes and their metabolites in fish.

Malachite Green (MG), Crystal Violet (CV) and Brilliant Green (BG) are antibacterial, antifungal and antiparasitic agents that have been used for treatment and prevention of diseases in fish. These dyes are metabolized into reduced leuco forms (LMG, LCV, LBG) that can be present in fish muscles for a long period. Due to the carcinogenic properties they are banned for use in fish for human consumption in many countries including the European Union and the United States. HPLC and LC-MS techniques are generally used for the detection of these compounds and their metabolites in fish. This study presents the development of a fast enzyme-linked immunosorbent assay (ELISA) method as an alternative for screening purposes. A first monoclonal cell line producing antibodies to MG was generated using a hybridoma technique. The antibody had good cross-reactivates with related chromatic forms of triphenylmethane dyes such as CV, BG, Methyl Green, Methyl Violet and Victoria Blue R. The monoclonal antibody (mAb) was used to develop a fast (20 min) disequilibrium ELISA screening method for the detection of triphenylmethanes in fish. By introducing an oxidation step with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) during sample extraction the assay was also used to detect the presence of the reduced metabolites of triphenylmethanes. The detection capability of the assay was 1 ng g(-1) for MG, LMG, CV, LCV and BG which was below the minimum required performance limit (MRPL) for the detection method of total MG (sum of MG and LMG) set by the Commission Decision 2004/25/EC (2 ng g(-1)). The mean recoveries for fish samples spiked at 0.5 MRPL and MRPL levels with MG and LMG were between 74.9 and 117.0% and inter- and intra-assay coefficients of variation between 4.7 and 25.7%. The validated method allows the analysis of a batch of 20 samples in two to three hours. Additionally, this procedure is substantially faster than other ELISA methods developed for MG/LMG thus far. The stable and efficient monoclonal cell line obtained is an unlimited source of sensitive and specific antibody to MG and other triphenylmethanes.

An RNA-aptamer-based assay for the detection and analysis of malachite green and leucomalachite green residues in fish tissue.

A robust screening assay employing solid phase extraction (SPE) followed by a novel aptamer-based procedure is presented for the rapid detection and semiquantitation of the triphenylmethane dye, Malachite Green (MG) and its primary metabolite Leucomalachite Green (LMG) in fish tissue. To the authors' knowledge, this is the first reported use of an RNA aptamer for the development of a diagnostic assay for the detection of chemical residues in food. The aptamer based screening assay is found to be highly specific for MG; but has negligible affinity for the LMG metabolite. However, because the LMG metabolite is lipophilic and known to be highly persistent in tissues, an oxidation step has been incorporated within the sample cleanup procedure to ensure that all LMG residues are converted to MG prior to measurement. This article provides evidence that an oligonucleotide aptamer can be used as an alternative recognition element to conventional antibodies with application to the detection of residues in food. Furthermore, this finding has the future potential to reduce the number of animals currently being used in the production of antibodies for immunodiagnostic kits.

A generic method for the quantitative analysis of aminoglycosides (and spectinomycin) in animal tissue using methylated internal standards and liquid chromatography tandem mass spectrometry.

Aminoglycosides (AGs) are a large and diverse group of antibiotics. Although AGs may cause side effects of nephrotoxicity and ototoxicity, they are still occasionally being used for the treatment of serious infections. In this study the development of a method is described for the quantitative determination and confirmation of seven aminoglycosides (and relevant isomers) and spectinomycin in animal tissues. The extraction was based on an extraction followed by a concentration and clean-up step using weak cation exchange solid phase extraction. The separation was performed by ion-pair liquid chromatography on a C(18) column followed by mass spectrometric detection. The method was validated according to the EU requirements for a quantitative confirmatory method. Permethylated aminoglycosides (in-house synthesised internal standards) were used for accurate quantification. The accuracy of the analyses of AGs in kidney ranged from 94 to 111%, intra-day precision ranged between 2.5 and 7.4% (R.S.D.(r)) and inter-day precision ranged between 2.2 and 17.3% (R.S.D.(RL), n=21, MRL level). Accuracy (muscle tissue) varied from 83 to 128% with an intra-day precision between 2.2 and 17.3% (R.S.D.(r), n=7, MRL level). From the results it was concluded that the method was able to monitor MRL levels which ranged from 750 to 20,000 microgkg(-1) for kidney and from 50 to 10,000 microgkg(-1) for muscle tissue.

A rapid microbial inhibition-based screening strategy for fluoroquinolone and quinolone residues in foods of animal origin.

A rapid, high-throughput antimicrobial screening assay for the detection of fluoroquinolone and 4-quinolone residues in foods of animal origin has been developed in ampoule format. The assay employs a single Escherichia coli species sensitive to those Gram-negative inhibitiory antimicrobial compounds and is presented in a comparable format to the existing commercially available Premi Test and Delvotest ampoule-based microbial inhibition tests (DSM, Delft, The Netherlands). In the novel E. coli assay the microorganism, in vegetative state, is inoculated into a nutrient agar pellet containing a pH sensitive acid-base indicator dye. A simple extraction protocol that is selective for fluoroquinolone and quinolone compounds was developed to recover, cleanup and concentrate the target analyte(s) from a variety of tissue types and matrices prior to screening analysis. The method detected 16 target compounds at concentrations equal to or below the maximum residue limits (where applicable). The method has been validated using the prototype assay in accordance with the 2002/657/EC guidelines for the validation of qualitative screening assays. False positive and false negative responses rates for the procedure have been determined as less than 5%. The stability of a selection of representative target analytes has been demonstrated for a 20-week period under a variety of storage conditions both in tissue and in extract.

Development and validation of an optical SPR biosensor assay for tylosin residues in honey.

In recent years there has been an increase in the use of tylosin in apiculture as bacterial brood diseases become resistant to oxytetracycline. Confirmatory mass spectrometry based methods have been developed but up until now there has been no complementary screening method available capable of sub 10 microg kg(-1) detection limits. In this paper the development and validation of a screening method using optical biosensor technology is presented. The honey was first dissolved in a phosphate buffer and following solid-phase extraction (SPE) cleanup was analyzed using a Biacore Q instrument. Using the criteria specified in European Commission Decision 2002/657/EC for qualitative screening methods, the detection capability (CCbeta) of the method was determined to be 2.5 microg kg(-)(1). Honey samples containing trace residue levels of tylosin were analyzed by both the biosensor screening method and a LC-MS/MS confirmatory procedure; the results were in good agreement.