Detection Of Food Allergens Research Articles

Specific detection of crustacean allergens in food: Development of indirect competitive and sandwich ELISA targeting sarcoplasmic calcium binding protein

To overcome the problem of cross-reactivity between crustaceans, mollusks, insects, and mites in current crustacean allergen immunodetection, the significant stable crustacean allergen, sarcoplasmic calcium binding protein (SCP) was innovatively selected as the target of ELISA detection. An indirect competitive ELISA (icELISA) and a sandwich ELISA (sELISA) were constructed and evaluated with spiked model foods. The sELISA (LOD 0.001 mg/kg and LOQ 0.003 mg/kg) is more sensitive than icELISA (LOD 0.11 mg/kg and LOQ 0.56 mg/kg). But the icELISA presented a wider standard range (10–10000 ng/mL) and greater recoveries of SCP in matrices (84.04%–118.94%). Sufficient precision (repeatability and reproducibility <15%), excellent specificity distinguishing crustaceans from others, and adequate detectability in diverse commercial processed foods were observed from the SCP-based icELISA. These results demonstrated the feasibility and applicability of SCP-targeted icELISA and provided a new direction to improve the capacity for crustacean allergen detection.

Sesame Detection in Food Using DNA-Functionalized Gold Nanoparticles: A Sensitive, Rapid, and Cost-Effective Colorimetric Approach.

Food safety control is a key issue in the food and agriculture industries. For such purposes, developing miniaturized analytical methods is critical for enabling the rapid and sensitive detection of food supplements, allergens, and pollutants. Here, a novel bioanalytical methodology based on DNA-functionalized gold nanoparticles (AuNPs) and colorimetric detection was developed to detect the presence of sesame (a major allergen) through sesame seed DNA as a target, in food samples. The presence of sesame DNA induces controlled nanoparticle aggregation/desegregation, resulting in a color change (from blue to red) proportional to sesame DNA concentration. The incorporation of multicomponent nucleic acid enzymes (MNAzymes) in this strategy has been carried out to perform an isothermal signal amplification strategy to improve the sensitivity of detection. Also, open-source software for color analysis was used to ensure an unbiased visual color-change detection, enhancing detection accuracy and sensitivity and opening the possibility of performing a simple and decentralized analyte detection. The method successfully detected the presence of sesame DNA in sesame seed, sesame oil, olive oil, and sunflower oil. In brief, the developed approach constitutes a simple and affordable alternative to perform a highly sensitive detection of DNA in food without complex methodologies or the requirement of expensive instrumentation.

Application of MOF Nanoenzyme in Rapid Detection of Food Allergens System

Food allergies are a common immune response that can lead to serious health problems. Rapid and accurate detection of allergens in food is crucial for protecting people’s health and quality of life. In this study, β-lactoglobulin in milk serves as a model allergen, and nucleic acid aptamer-loaded MN patches are utilized as rapid capture elements. Moreover, metal organic frameworks loaded with complementary aptamer sequences are employed as signal amplification elements. The combination of these two elements establishes a rapid visualization detection system for allergens, employing specific microneedle array patches-MOF nanoenzymes. The results show that the combination of TMB+H2O2 +MOF-RC has the highest peroxidase like activity of 1.732. Compared to TMB+H2O2 +MOF, it has increased by 0.547. Meanwhile, the MN-MOF detection method has excellent specificity for the detection of β-Lg. This specific microneedle array patches and MOF nanoenzymes-based allergen rapid detection system demonstrates practical application benefits, providing a viable solution for swiftly detecting food allergens and safeguarding people’s health and quality of life.

The differences of characteristics and allergenicity between natural and recombinant tropomyosin of Macrobrachium nipponense

Tropomyosin (TM) is the main allergen of Macrobrachium nipponense. Recombinant allergens have great prospects in the detection, diagnosis, and treatment of food allergens. The purpose of this study was to compare the differences in structure and allergenicity between natural TM and recombinant TM. Recombinant TM of M. nipponense with a molecular weight of 38 kDa was successfully expressed in the Escherichia coli system. The amino acid sequence as well as secondary structure between natural and recombinant TM were similar, which were verified by mass and CD spectrometry, respectively. Studies showed that both natural TM and recombinant TM had strong allergenicity, and recombinant TM was more allergenic, which could be used as a substitute for natural TM in the diagnosis and treatment of shrimp allergy. This study provided stable and reliable allergen components for the detection of crustacean allergens and the diagnosis and treatment of food allergies caused by crustacean allergens.

Rapid and sensitive detection of shrimp allergens within 30 min using a new food nucleic acid release agent combined with a fast real-time PCR assay

To achieve fast on-site detection of shrimp allergens in food, this study established a new direct, fast, and real-time quantitative polymerase chain reaction (PCR) technology. The technology completed nucleic acid extraction in 4 min using a new nucleic acid releaser, and the DNA samples were detected using fast qPCR within 25 min (total detection was approximately 30 min). The results indicated that the direct fast qPCR can specifically detect shrimp, with a limit of detection of 0.0001% of shrimp in artificially simulated meat samples and a detection precision variation coefficient (CV value) was less than 5%. It successfully identified shrimp in different types of commercial food samples. As a simple, fast, and sensitive nucleic acid testing technology, it has broad application prospects for the on-site detection of shrimp allergens in food.

A sensitive CRISPR/Cas12a-assisted fluorescent aptasensor for rapid detection of food allergens

Food allergens elicit abnormal immune system responses among allergic individuals and sensitive detection for allergenic ingredient is greatly significant. To address this need, a novel fluorescent aptasensor, assisted by Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR), have been developed for food allergens. In this study, aptamer offers distinctive recognition capabilities in binding specific targets, while CRISPR-associated-12a protein (Cas12a) holds precise cis-cleavage for cutting fluorescent signal probes. Notably, the utilization of Cas12a cis-cleavage activity, rather than trans-cleavage, eliminates the necessity for additional fluorescent probes, thus reducing interference between substances and enhancing sensitivity. Throughout the process, complementary DNA (cDNA) plays a crucial dual role in target recognition conversion and signal presentation, representing a key challenge and innovative aspect of this study. To evaluate the performance of the aptasensor, lysozyme (LYS) is employed as a representative model target of food allergens. Under optimal conditions, the developed aptasensor could achieve an exceptional low limit of detection (LOD) of 6.10 pM with a dynamic detection range of 10 pM-320 pM. The aptasensor demonstrates high selectivity and great recovery rates. This strategy yields promising outcomes, holding the potential to serve as a valuable reference for various food allergens detection.

Rapid and accurate detection of fish allergen within 30 min using a food nucleic acid releaser combined with rapid real time PCR assay

To achieve rapid on-site nucleic acid detection of fish allergens in food, this study established a new direct and rapid qPCR detection technology for fish. The technology was able to complete nucleic acid extraction in 4 min using a food nucleic acid releaser, and the DNA samples were amplified by rapid qPCR within 25 min. The results showed that direct-rapid qPCR can specifically detect fish, and it can also detect 0.00001 % of fish components in artificially simulated samples, with high detection precision. This method can be used to effectively detect fish in various deep-processed foods. As a simple, rapid, and accurate molecular detection method for fish allergens, this method has broad application prospects for on-site food safety detection.

Non-destructive Near-infrared Hyperspectral Imaging in Food Technologies with a Focus on Monitoring Seed Viability

Seed qualities including viability and germination significantly influence the quantity and the quality of harvest. Technological means to assess seed qualities and attributes of seed-derived food products are varied. This paper highlights the use of infrared hyperspectral imaging (NIR- HSI) in food quality control, authentication, safety, process monitoring, shelf-life prediction, ingredients analysis, allergens detection and food sorting and grading. It also shows a particular application of NIR-HSI for the monitoring of nutrient content of sprouts and germinated seeds for industrial processing of foods with high nutritional values. The paper further reviews the applications of NIR-HSI to predict seed viability and germination. The non-destructive, rapid, and high-throughput capability of NIR-HSI were demonstrated through research works combining the NIR-HSI technology with chemometrics tools to reach more than 90% prediction rate. These relatively high rates may depend on the storage conditions or the stringency of the artificial aging conditions applied to parts of the seeds. However, the NIR-HSI has also proven efficient using naturally aged seeds with the prediction rates up to 90% correct classification, demonstrating the high capability of the technology. In combination with advanced chemometrics tools, some components of emerging technologies such as traditional machine learning and deep learning models have been added to increase the efficiency of NIR_HSI. Overall, the research works reviewed in this paper and which cover several food crops and food products showed that NIR-HSI is set to reach new heights in monitoring seed viability for improved seed stock management, crop production and innovation in the food industry.

Ensuring food safety: Microfluidic-based approaches for the detection of food contaminants.

Detecting foodborne contamination is a critical challenge in ensuring food safety and preventing human suffering and economic losses. Contaminated food, comprising biological agents (e.g. bacteria, viruses and fungi) and chemicals (e.g. toxins, allergens, antibiotics and heavy metals), poses significant risks to public health. Microfluidic technology has emerged as a transformative solution, revolutionizing the detection of contaminants with precise and efficient methodologies. By manipulating minute volumes of fluid on miniaturized systems, microfluidics enables the creation of portable chips for biosensing applications. Advancements from early glass and silicon devices to modern polymers and cellulose-based chips have significantly enhanced microfluidic technology, offering adaptability, flexibility, cost-effectiveness and biocompatibility. Microfluidic systems integrate seamlessly with various biosensing reactions, facilitating nucleic acid amplification, target analyte recognition and accurate signal readouts. As research progresses, microfluidic technology is poised to play a pivotal role in addressing evolving challenges in the detection of foodborne contaminants. In this short review, we delve into various manufacturing materials for state-of-the-art microfluidic devices, including inorganics, elastomers, thermoplastics and paper. Additionally, we examine several applications where microfluidic technology offers unique advantages in the detection of food contaminants, including bacteria, viruses, fungi, allergens and more. This review underscores the significant advancement of microfluidic technology and its pivotal role in advancing the detection and mitigation of foodborne contaminants.

Development and validation of a targeted mass spectrometry method for the quantification of milk protein allergens in multiple complex food matrices using matrix-independent calibration

Quantitative detection of food allergens is crucial in assessing the risk of allergic responses in susceptible individuals and ensuring regulatory compliance. Targeted mass spectrometry methods provide an opportunity to analyze allergens from complex, processed food matrices. However, probable interferences of diverse food matrices mandates preparation of matrix-matched calibration curves, challenging adaptation of methods into multiple matrices. In this study, we have developed a novel matrix-independent calibration strategy employing exogenous carrier proteins as background and demonstrated its adaptability in quantification of milk from multiple food matrices at concentrations relevant for risk assessment applications. Peptides representing casein or whey fractions of milk were detectable from concentrations as low as 1 ppm nonfat dry milk (NFDM) in baked cookie matrix at 46–99% accuracy, 0.5 ppm total milk protein (TMP) in pasteurized beverages at 94–138% accuracy, and 1 ppm casein in chocolate at 35–51% accuracy. Quantitative estimations of milk from matrix-independent calibration curves in the background of egg white powder as exogenous carrier were comparable to those from a matrix-matched calibration curve, and the quantitative precision was within 30% CV across the inter-day replicates. The sensitivity, accuracy, and precision of this matrix-independent approach ensure robustness of the method for high throughput food allergen analysis as required in routine testing labs.

Integrated microfluidic platform for on-site qPCR analysis: food allergen detection from sample to result

Improving food safety is crucial in the contexte of “One Health” approach. To guarantee product quality and safety, food industry, having a very high turnover rate, needs short time-to-result analyses....

The detection and regulation advances of food allergens

The detection and regulation advances of food allergens

Comprehensive characterization and detection of nut allergens in bakery foods using Q-TOF mass spectrometry and bioinformatics

Abstract Food allergy is a growing health issue worldwide and the demand for sensitive, robust and high-throughput analytical methods is rising. In recent years, mass spectrometry-based methods have been established for multiple food allergen detection. In the present study, a novel method was developed for the simultaneous detection of almond, cashew, peanut, and walnut allergens in bakery foods using liquid chromatography–mass spectrometry. Proteins unique to these four ingredients were extracted, followed by trypsin digestion, quadrupole time-of-flight (Q-TOF) mass spectrometry and bioinformatics analysis. The raw data were processed by de-novo sequencing module plus PEAKS DB (database search) module of the PEAKS software to screen peptides specific to each nut species. The thermal stability and uniqueness of these candidate peptides were further verified using triple quadrupole mass spectrometry (QQQ-MS) in multiple reaction monitoring (MRM) mode. Each nut species was represented by four peptides, all of which were validated for label-free quantification (LFQ). Calibration curves were constructed with good linearity and correlation coefficient (r2) greater than 0.99. The limits of detection (LODs) were determined to range from 0.11 to 0.31 mg/kg, and were compared with the reference doses proposed by Voluntary Incidental Trace Allergen Labelling (VITAL). The recoveries of the developed method in incurred bakery food matrices ranged from 72.5% to 92.1% with relative standard deviations (RSD) of &amp;lt;5.2%. The detection of undeclared allergens in commercial bakery food samples confirmed the presence of these allergens. In conclusion, this method provides insight into the qualitative and quantitative detection of trace levels of nut allergens in bakery foods.

Assessment of crustacean allergen detection methods: cross reactivity with edible insect samples

Increased interest in consumption of insects in recent years has led to an increased focus on associated food safety concerns, and allergy is one of the most relevant. In the United States, crustacean shellfish are regulated as a major allergenic food group per the Food, Drug, and Cosmetic (FD&C) Act. Insects and crustacean shellfish are both arthropods, and clinical cross-reactivity between the two groups has been demonstrated. The goal of this work was to establish whether that clinical cross-reactivity translates into analytical cross-reactivity with detection assays targeting crustacean shellfish allergens. Edible insect samples were analyzed using four different crustacean allergen detection methods: Multi-Analyte Profiling Food Allergen Detection Assay (xMAP FADA), enzyme-linked immunosorbent assay (ELISA), western blot, and real-time polymerase chain reaction (PCR). Results indicate that the immunoassay-based xMAP FADA, ELISA, and western blot were susceptible to cross-reactivity, while the DNA-based PCR methods had minimal reactivity with insect samples. These results confirm that edible insects show analytical cross-reactivity with the immunoassays which may result in false positive detection of crustacean allergens in insect samples. Confirmation using DNA-based PCR, which shows little to no cross-reactivity, clarifies ambiguous results.

Standardization of a Mass Spectrometry-Based Workflow for Food Allergen Quantification.

In this chapter, the analytical workflow typically used for the development and validation of an analytical method tailored to food allergen detection and quantification is presented. The main steps defining the workflow are herein described and commented with specific notes about the critical issues that can be faced and common solutions to be adopted. References to guidelines and/or recommendation available from official bodies, as well as main papers from international consortia operating on the specific research field, are also reported, whenever possible. As such, this chapter may represent a practical guide to drive method development in the standardization of analytical methods for food allergen detection.

Biosensors for the Detection of Food Allergens.

Biosensors enable fast and specific detection of various molecules, including allergens. Surface plasmon resonance (SPR) aptasensors combine the capabilities of SPR detection, i.e., sensitive, fast, label-free, and real-time monitoring with the specificity and stability of aptamers for the biorecognition of various ligands. Compared to other toxic compounds in food, allergens pose specific analytical challenges. Among allergens, lysozyme, also named Gal d 4, is a food additive used in cheese, wine, beer, sausages, etc., to control bacterial activity. In this chapter, we describe an SPR aptasensor that is applicable for lysozyme analysis in wines. Besides detecting residual allergen amounts, the aptasensor can be used to monitor the interaction of lysozyme with phenolic compounds in wine.

Validation Procedures for Quantification of Food Allergens by Enzyme-Linked Immunosorbent Assay (ELISA).

Enzyme-linked immunosorbent assay (ELISA) is a widely used analytical technique for food allergen detection and quantification. Validating ELISA protocols is important for both assay developers and end users as it ensures method reliability. This chapter describes the protocols for validating the sensitivity, specificity, precision, accuracy, robustness, and ruggedness of an ELISA. Example procedures are also provided for sample preparation, allergen extraction, and ELISA operation.

Food allergen detection by mass spectrometry: From common to novel protein ingredients.

Food allergens are molecules, mainly proteins, that trigger immune responses in susceptible individuals upon consumption even when they would otherwise be harmless. Symptoms of a food allergy can range from mild to acute; this last effect is a severe and potentially life-threatening reaction. The European Union (EU) has identified 14 common food allergens, but new allergens are likely to emerge with constantly changing food habits. Mass spectrometry (MS) is a promising alternative to traditional antibody-based assays for quantifying multiple allergenic proteins in complex matrices with high sensitivity and selectivity. Here, the main allergenic proteins and the advantages and drawbacks of some MS acquisition protocols, such as multiple reaction monitoring (MRM) and data-dependent analysis (DDA) for identifying and quantifying common allergenic proteins in processed foodstuffs are summarized. Sections dedicated to novel foods like microalgae and insects as new sources of allergenic proteins are included, emphasizing the significance of establishing stable marker peptides and validated methods using database searches. The discussion involves the in-silico digestion of allergenic proteins, providing insights into their potential impact on immunogenicity. Finally, case studies focussing on microalgae highlight the value of MS as an effective analytical tool for ensuring regulatory compliance throughout the food control chain.

Multifold improvement in allergen detection capability of dipstick-type immunosensors via macromolecular crowding

Dipstick-type lateral flow immunosensors are used widely for on-site detection of food allergens. The weakness of the immunosensors of this type, however, is their low sensitivity. Contrary to current methods, that focus on improving detection capability through the introduction of novel labels or multistep protocols, this work exploits macromolecular crowding to modify and regulate the microenvironment of the immunoassay, thus promoting the interactions that are responsible for allergen recognition and signal generation. The effect of 14 macromolecular crowding agents was explored using, as a model, commercially available and widely applied dipstick immunosensors, which are already optimized in terms of reagents and conditions for peanut allergen detection. An about 10-fold improvement in detection capability was achieved by using polyvinylpyrrolidone, Mr 29,000, as a macromolecular crowder without compromising simplicity and practicality. The proposed approach is complementary to other methods of improving the sensitivity by using novel labels. Because biomacromolecular interactions have a fundamental role in all types of biosensors, we foresee that the proposed strategy will also find applications in other biosensors and analytical devices.

Immunostick colorimetric assay for highly sensitive detection of food allergens by bio-nanocapsule-scaffolding technology.

The detection sensitivity of immunostick colorimetric assay has been increased by using a bio-nanocapsule as a scaffold for oriented immobilization of immunoglobulin Gs. This immunostick produced ∼82-folds stronger coloration in the detection of food allergens and reduced detection time by a factor of 5.