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detection and quantification of almond (Prunus dulcis) in food with ELISAOrebrand, Ulrika January 2006 (has links)
<p>Reliable methods to analyze food for the presence of almond are important – not only for those allergic to almond, but also for monitoring the compliance with labelling regulations (EG directive 2003/89). Until now the Swedish National Food Administration has used methods like rocket immunoelectrophoresis and real-time PCR to detect almond in food. These methods are, however, not sensitive enough for protecting the most sensitive individuals. Therefore, the performance of a commercial ELISA kit was tested with regard to specificity/cross reactivity and limit of detection for almond both in solution and in different matrixes.</p><p>The limit of quantitation was at least 3,1 ppm (mg/kg) in solution and similar concentrations were measured in bisquits and chocolate. The ELISA method was about 100-fold more sensitive than rocket immunoelectrophoresis and PCR.</p><p>The specificity of the test kit was evaluated against a number of different nuts and seeds. No important cross reactivity was found. The antibodies against almond used in the kit can not differentiate between almond and apricot kernel. For such purposes the PCR method could be used.</p>
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detection and quantification of almond (Prunus dulcis) in food with ELISAOrebrand, Ulrika January 2006 (has links)
Reliable methods to analyze food for the presence of almond are important – not only for those allergic to almond, but also for monitoring the compliance with labelling regulations (EG directive 2003/89). Until now the Swedish National Food Administration has used methods like rocket immunoelectrophoresis and real-time PCR to detect almond in food. These methods are, however, not sensitive enough for protecting the most sensitive individuals. Therefore, the performance of a commercial ELISA kit was tested with regard to specificity/cross reactivity and limit of detection for almond both in solution and in different matrixes. The limit of quantitation was at least 3,1 ppm (mg/kg) in solution and similar concentrations were measured in bisquits and chocolate. The ELISA method was about 100-fold more sensitive than rocket immunoelectrophoresis and PCR. The specificity of the test kit was evaluated against a number of different nuts and seeds. No important cross reactivity was found. The antibodies against almond used in the kit can not differentiate between almond and apricot kernel. For such purposes the PCR method could be used.
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Characterization of a rabbit-antiserum for detection of pea protein in foodsLundholm, Linnéa January 2008 (has links)
Food allergy is an IgE-mediated immunological disease, which affects almost 4% of the adult population and up to 6% of children. Proteins from milk, egg, peanuts, soybean, wheat, fish and nuts are the main cause of food allergies. A less common allergen is pea protein. The National Food Administration analyses undeclared pea protein and contaminations of pea protein in foods using rocket immunoelectrophoresis and immunodiffusion. For both methods an antiserum against pea protein is needed. The aim of this study has been to characterize a newly developed rabbit-antiserum against pea protein. It is important to know if the antiserum is specific against peas, the detection as well as the quantification limits before it can be taken into use. The results of the study show that the antiserum was not absolutely specific, since it cross-reacted with chickpeas, fenugreek and lenses. However there is an "in-house" established PCR-method that can distinguish between chickpeas, fenugreek and peas and that method can be used as a complement to the rocket immunoelectrophoresis. The PCR-method cannot be used alone because it is not quantitative. Rocket immunoelectro¬phoresis detects 0,003% pea protein with purified IgG-antibodies from the antiserum.
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Development of immunological methods and Real-Time PCR for detection of Macadamia nut (Macadamia spp.)Eliasson, Hanna January 2005 (has links)
A new European labeling directive (2003/89/EC) states that certain foods and products derived thereof must always be declared. Among the tree nuts specified is Macadamia nut (Macadamia spp.). During the last few years, cases of IgE-allergic reactions, even severe anaphylaxes, have been reported. Reliable methods for the detection of this nut are needed. Protein from Macadamia nuts was isolated. Polyacrylamide gel electrophoresis in SDS revealed two main protein bands of about 20 and 50kDa. These protein bands were cut and extracted from the gel and rabbits were immunized with each protein. Immunoblotting showed dominant reactivity with the respective antigens. The antisera were further tested for specificity in immunodiffusion and in rocket immunoelectrophoresis. In addition, a specific DNA-method was developed, based on Real-Time PCR using Macadamia vicilin as target sequence. Two different primer pairs were tested. Specificity was tested against potentially related nuts. Optimisation of primer and probe concentrations was performed. The limit of detection was 2-4 pg DNA, corresponding to a macadamia nut concentration of 50 to 100 μg per g. In a background of soybean DNA, down to 0,01 % macadamia DNA could be detected.
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Comparison of isoelectric focusing and immunofixation electrophoresis to distinguish oligoclonal from monoclonal immunoglobulin bands.January 1998 (has links)
submitted by Liu Dan. / Thesis (M.Sc.)--Chinese University of Hong Kong, 1998. / Includes bibliographical references (leaves 66-80). / Abstract also in Chinese. / CONTENTS --- p.i / LIST OF TABLES --- p.iii / LIST OF FIGURES --- p.iv / LIST OF ABBREVIATIONS --- p.v / ACKNOWLEDGEMENTS --- p.vi / ABSTRACT --- p.vii / Chapter Chapter 1 --- INTRODUCTION --- p.1 / Chapter 1.1 --- History --- p.1 / Chapter 1.2 --- Immunoglobulins --- p.3 / Chapter 1.2.1 --- Structure --- p.3 / Chapter 1.2.2 --- Properties of immunoglobulins --- p.7 / Chapter 1.3 --- Monoclonal proteins and monoclonal gammopathies --- p.12 / Chapter 1.3.1 --- Monoclonal proteins --- p.12 / Chapter 1.3.2 --- Monoclonal gammopathies --- p.14 / Chapter 1.4 --- Laboratory investigation of monoclonal immunoglobulin --- p.17 / Chapter 1.4.1 --- The current procedure of investigation in laboratory --- p.17 / Chapter 1.4.2 --- Problems in identifying monoclonal immunolgobuin --- p.19 / Chapter 1.5 --- Comparison of different techniques --- p.20 / Chapter 1.5.1 --- Immunoelectrophoresis --- p.20 / Chapter 1.5.2 --- Immunofixation electrophoresis --- p.22 / Chapter 1.5.3 --- Isoelectric focusing and immunoisoelectric focusing --- p.24 / Chapter 1.6 --- Aim of the present study --- p.27 / Chapter 1.7 --- Design of experiment --- p.27 / Chapter Chapter 2 --- MATERIALS AND METHODS --- p.30 / Chapter 2.1 --- Study subjects --- p.30 / Chapter 2.2 --- Apparatus --- p.30 / Chapter 2.2 --- Apparatus --- p.30 / Chapter 2.3 --- Reagents and materials --- p.32 / Chapter 2.4 --- Preparation of gels --- p.35 / Chapter 2.5 --- Isoelectric focusing procedure --- p.36 / Chapter 2.6 --- Acid fixation and staining --- p.37 / Chapter 2.7 --- Technical factors affecting results --- p.38 / Chapter Chapter 3 --- RESULTS --- p.40 / Chapter 3.1 --- Interpretation of results in isoelectric focusing --- p.40 / Chapter 3.2 --- Affecting factors --- p.47 / Chapter 3.3 --- Comparison of the results between IEF and IFE --- p.53 / Chapter Chapter 4 --- DISCUSSION --- p.59 / Chapter Chapter 5 --- CONCLUSION --- p.65 / References --- p.66
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