Detection of fish allergen by droplet digital PCR

Daga, Cinzia; Cau, Simona; Tilocca, Maria Giovanna; Soro, Barbara; Marongiu, Aldo; Vodret, Bruna

doi:10.4081/ijfs.2018.7264

Cited by 8 publications

(4 citation statements)

References 11 publications

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“…The RT-PCR-assay detection method was developed targeting the ribosomal 18S rRNA gene. Primers were designed according to Daga et al [ 47 ]: forward primer, 18S F (5′-GTACACACGGCCGGTACAGT-3′), reverse primer 18S R (5′-CATGGGTTTTGGGTCTGATAA-3′), and probe 18S P (FAM-5′-CCGTACTTGGATAACTGTGGCAATTC-3′) (Sigma-Aldrich, St. Louis, MO, USA).…”

Section: Methodsmentioning

confidence: 99%

“…Even though PCR methods have been mainly used for species authentication purposes, they are a useful tool to detect the potential presence of fish allergens in foods. Real-time PCR assays for the detection of fish have been proposed targeting either nuclear (e.g., parvalbumin, rhodopsin, Hoxc13, and 18s rRNA) and mitochondrial genes (12S rRNA and 16S rRNA) [ 38 , 41 , 46 , 47 , 48 , 49 ]. Real-time PCR commercial kits for the detection of different allergens are available on the market.…”

Section: Introductionmentioning

confidence: 99%

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Detection of Fish Allergens in Foods Using an In-House Real-Time PCR Targeting the Ribosomal 18S rRNA Gene

Cau

Daga

Spanu

et al. 2022

Foods

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Fish is one of the major food allergens which, in sensitised individuals, can cause life-threatening allergic reactions, even when present in small amounts. To protect consumers’ health, the correct labeling of foods is important. The objective of the present study was to validate an in-house real-time PCR method targeting the ribosomal 18S rRNA gene as universal DNA marker for the detection of fish in foods. The specificity of the primers was assessed on 20 fish species commonly marketed in the Mediterranean basin and other species of molluscs and crustaceans and foods of animal and plant origin. The absolute detection of the method was assessed using DNA extracted from a fish mixture and the SureFood® QUANTARD Allergen 40 reference material. The relative amount was assessed on a fish and béchamel sauce blend. Commercial food samples either labelled with or without fish in the ingredient list, were tested for the presence of fish DNA. The primer showed high specificity against the selected fish species. The limit of detection (LOD) and limit of quantification (LOQ) of the in-house method were 0.5 pg/µL and 5 pg/µL, respectively. The relative quantification in fish and béchamel blend samples detected a concentration as low as 0.000025%, corresponding to 0.25 mg/kg of fish, indicating the suitability of the method in a food matrix. The presence of fish DNA was always detected in commercial samples in which the presence of fish was listed in the ingredient list. The method was able to detect the presence of fish DNA also in samples in which the presence of fish was indicated as traces or was not declared on the label. The proposed method was demonstrated to be a reliable, specific, and sensitive method for the detection of fish allergens in foods. Therefore, the proposed real-time PCR method could be used as a useful instrument in the verification of compliance with allergen labelling regulations.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Detection of Fish Allergens in Foods Using an In-House Real-Time PCR Targeting the Ribosomal 18S rRNA Gene

Cau

Daga

Spanu

et al. 2022

Foods

View full text Add to dashboard Cite

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“…Information about DNA-based detection methods used for seafood allergen is summarized in table 2 below. [9] Real-time PCR…”

Section: Mechanism Of Seafood Allergymentioning

confidence: 99%

The immune response of seafood allergen and their detection methods: A review

Khushbu¹,

Gulati²,

Sushma³

2022

Pharma Innovation

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Seafood is highly nutritive and has an excellent source of proteins, vitamins, minerals, and essential polyunsaturated fatty acids such as Eicosatetraenoic acid (ETA) and Docosahexaenoic acid (DHA) from the n-3 series (omega-3). Due to great nutritive value and large health benefits, their consumption has been increasing continuously. Sometimes but significantly, in a small number of individuals consumption of seafood often leads to adverse reactions like a food allergy. It is a global public health issue in regards to food safety that decline the quality of life and may endanger the lives in severe cases. Seafood allergen can induce hypersensitive immune responses due to their ingestion, skin contact, and inhalation. Recent advancement in the food processing sector implies a great focus on the identification of food allergens by developing a rapid and user-friendly method to assess risk through proper labeling of foods and proper management. According to recent European Union regulations, food producers are advised to declare the presence of foods that are potentially allergic ingredients and highlight them by proper labeling. These efforts enable consumers' health to be guaranteed. There are various techniques for detection such as DNA-based, Aptamer based, and protein-based. These are more efficient and reliable detection methods for seafood allergen and will ensure the food safety and health of the consumer.

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“…[107] In order to reduce droplet rain generation, gradient temperature annealing and changes in PCR cycling were performed. [108] In order to obtain lower LODs and LOQs, Mayer…”

Section: J O U R N a L P R E -P R O O Fmentioning

confidence: 99%