Food Allergen & Detection Methods

Transcription

1 Food Allergen & Detection Methods Vipa Surojanametakul Institute of Food Research and Product Development Kasetsart University October 22 th, 2015

2 Food : What come the first importance? Taste Enjoyment Full Safety! Economy Nutrition Health Savor Shoji,2012

3 Understanding Food Allergy Food allergy is any abnormal reactions, occurs when the immune system (IgE-mediated system) mistakenly attacks a food protein (which is called food allergen ) when ingested by allergic individual. Ingestion of the offending food may trigger the sudden release of chemicals, including histamine, resulting in symptoms of an allergic reaction, ranging from relatively mild to severe (as anaphylaxis) or sometimes potential life threatening. The symptoms typically develop in less than 10 minutes up to 2 hours after consumption of the offending food

4 How come Food allergy is a serious problem? Significant impacts to our society The prevalence is marked and moreover increasing in both developed & developing countries, especially in baby & children Incredible health hazard Severity and complexity of symptom is increasing and more frequency cause of fatal consequence, i.e. in USA 150 persons dried /year due to food allergy Significant socioeconomic expense As top ten chronic disease in USA and is estimated the medication cost for food allergy is ca, $ 25 Billion Currently, there is no known medication for curing food allergy

5 How can food allergy be managed? Presently, the most effective way of preventing allergic reactions is the strict avoidance of the offending allergenic food ( as self protection ). In order to help food allergic individuals, the regulators of International and National levels have introduced the regulation for allergen labeling of prepackaged foods.

6 Food Ingredients for Food Allergen Labeling a: Shrimp and crab are the only crustacean listed. Grain included wheat and buckwheat Others includes food for which labeling is recommended but not required:abalone, squid, salmon roe, oranges, cashew nut, kiwifruit, beef, walnuts, sesame, salmon, mackerel, soybeans, chicken, bananas, pork, matsutake mushrooms, peaches, yams, apples and gelatin b: Shrimp and crab are the only crustacean listed. Grain included wheat and buckwheat. Mackerel is the only fish. Other includes pork, peaches and tomato.

7 Thai government action related to Food allergen Ministry of Public Health: Thai Food and Drug Administration is already announced to enforce Food Labeling regulation including Food Allergen labeling (following the Codex ) since June 6 th, Ministry of Agriculture and Cooperative: National Bureau of Agricultural Commodity and Food Standards prepared the instruction, Food Allergens, for Thai food exporter facilitating to adjust themselves for North America, EU, Japan and Australia markets etc.

8 What is Food Allergens? Food allergens Protein or glycoproteins with the MW Kdal (Not all proteins are allergens) Naturally occurring (plants & animals) or newly introduced in GM foods Comparatively stable to processing, cooking, proteolysis, and the digestive processes Have a specific surface features that antibodies recognize Causing abnormal immunological reactions (immediate)

9 Structure Protein with a globular compact structure Multi IgE binding sites (Contain more than 1 Epitope) Processing impact conformational change cause a change in solubility and allergenicity properties Source: Shoji, M and Obata,T.2010

10 Allergenic Foods More than 160 foods have been associated with allergic reaction Eight types of allergenic food responsible for over 90% of all allergic reaction, named Big 8 Cow s milk Egg Fish Shellfish Wheat Peanut Tree nut Soya The amount of allergen required to elicit an allergic reaction differs from person to person No threshold level has been identified for most allergens

11 Major allergenic protein in foods Milk Food items Major allergenic protein casein / beta-lactoglobulin/ alpha-lactalbumin/bovine serum albumin, Immunoglobulins, Lactoferrin Egg Fish Shellfish Wheat Peanut Soybean Tree nut and Seeds Fresh fruits and Vegetables Egg white : ovalbumin /ovomucoid / lysozyme /ovaltransferin Egg yolk : α livetin parvalbumin tropomyosin (Cross-reactivity in various seafood : shrimp, crab, squid, abalone) albumin, globulin, prolamin (gluten / gliadine) Seed storage protein (globulin, albumin)/ conarachin/ arachin1-8 beta-conglycinin, gylcinin Seed storage protein (albumins, globulin) Bet v1, latex

12 Allergen presence in food Food Allergen Intentionally added Unintentionally added Hidden allergen Appropriate labeling Allergen testing

13 Food allergen detection Why Test for Allergens? Consumer safety Ensure accuracy of food labeling Cleaning effectiveness Consumer complaints Uses of Allergen Tests Raw material/ingredients Effectiveness of allergen control program (cleaning/ environment) Finish products for labeling compliance Investigate/confirm consumer complaints Regulatory agency effort (Lauren S. Jackson,2010)

14 Allergen Detection Immunochemical methods (protein based) ELISA Lateral flow devices (dipsticks) DNA-based methods PCR Generic/non-specific (cleaning) Protein ATP Visual inspection Mass spectrometry

15 Enzyme-Linked Immunoassay (ELISA) Antibody-based detection of allergenic protein or other protein in food Available in well (micro plate) and lateral flow formats( dip stick) Sandwich format most common The most commonly used as screening test ELISA analysis take between 1 2 h to complete Quantitative & qualitative Kits available for most of the major allergens Used for ingredients, finished products, cleaning fluids, swabs, environmental samples (Lauren S. Jackson, JIFSAN Advisory Council Spring Symposium, 2010 )

16 Sandwich ELISA

17 Lateral Flow Devices (LFD) - Dipstick Tests Rapid qualitative test of allergen (can be semiquantitative with reader) Suitable for industrial purpose Available for many allergenic foods Typically used for environmental sampling, cleaning verification, screening of foods True measurement of presence of allergenic food Satisfactory sensitive (DL~ 5 ppm)

18 Enzyme-Linked Immunosorbent Assay (ELISA) Advantages High selective to allergen residue High sensitive (in low ppm range) Powerful - qualitative /quantitative tool Crude sample use Many samples possible for one analysis (not for LFD) Disadvantages Long developmental time Cross-reactivity possible Matrix effect (polyphenol, ph, oil etc.) Trained staff requirement ( but not for LFD) Lack of reference material, need do in house validation

19 ELISA Points of Consideration Antibody Specificity total protein vs. allergen Polyclonal vs. Monoclonal Antibodies Calibrators Effects of Processing on Detection Efficiency of Extraction method Sensitivity Limits (Joe Baumert, 2011)

20 PCR Method Confirmation test for allergen detection by ELISA Specific to the source (DNA) but not to the allergenic proteins Sensitive (high) Qualitative /Semi-quantitative Depends on specific primers Available for many allergenic food sources

21 Polymerase chain reaction (PCR) Advantages Detected origin of allergic food (species specific) DNA stable than protein Suitable for confirmation Short developmental time Disadvantages Not detected allergenic protein False positive result Not practical for in-plant use Required trained staff and isolated lab DNA is low in some high protein food (milk, egg white, protein isolated) Some foods cannot be differentiated (beef/milk; egg/chicken) Food matrix & processing can negatively impact on DNA extraction and detection

22 General Protein Tests Chemical reaction (Biuret method) Detects protein regardless of source but not specific for allergenic protein Can be used for routine sanitation verification but should first validate the cleaning methods with specific ELISAs ATP/Bioluminescence Tests Does not prove presence of protein and None are specific for allergens

23 Method selection The specificity of the test method : protein, allergenic protein, DNA, ATP The sensitivity of the test method : detection limit of target Recommended to validate removal of allergenic residue using specific ELISA Testing for total protein or ATP can be helpful in some cases. They can provide a good quick check on sanitation effectiveness during routine cleaning Allergen-specific ELISAs will allow very specific and sensitive detection for qualitative and quantitative of allergenic residue (Joe Baumert, 2011)

24 Conclusion Food allergy is regarded as a serious health hazard, globally To protect the health hazard of food allergy, food allergen labeling regulation has been enforced in many countries, including Thailand Undeclared food allergen is become an emerging issue related to food safety hazard Food allergen detection for accurate and reliable result is needed to meet the requirement of food allergen labeling regulation ELISA is a very powerful tool for allergenic residue testing

25 Food Allergen Detection Service (IFRPD, Kasetsart U.) FQA (ศ นย บร การประก นค ณภาพอาหาร) Food Allergen test : ELISA technique Wheat Gluten Peanut Soy bean Milk Egg Web site: http//www. Ifrpd-foodallergy.com ข อม ลองค ความร เก ยวก บภ ม แพ อาหาร สารก อภ ม แพ กฎระเบ ยบ ว าด วยการแสดงฉลากสารก อภ ม แพ เทคน คการตรวจว เคราะห... ฯลฯ

26 Thank you for your attention