Fish allergy is a worldwide problem, especially in industrialized countries where fish consumption is high. Fish contains a wide variety of proteins but, only few of them are responsible for triggering an allergic reaction. The major fish allergen, parvalbumin, is a low molecular weight (10-13 kD) heat-stable protein. High homology in amino acid sequences and antibody cross-reactivities have been demonstrated for parvalbumin in different fish species. Although several detection methods based on specific antibodies or DNA amplification are currently employed for detection of allergic components in food products, there is limited number of studies reporting methods for the detection of fish allergens. This study aimed to investigate the antigen binding characteristics of a monoclonal antibody (MAb) 3E1 by comparing its immunoreactivity against various fish and other animal species with a commercially available anti-frog parvalbumin monoclonal antibody (PARV-19) to evaluate the usefulness of MAb 3E1 as a anti-fish parvalbumin reagent. MAb 3E1 was previously developed in our laboratory against heat-treated catfish crude sarcoplasmic protein extract. The antigen binding characteristics of this antibody was investigated by comparing its immunoreactivity against soluble proteins extracts from various cooked fish and other animal meats with MAb PARV-19. Non-competitive indirect ELISA was performed to examine the immunoreactivity of both MAb 3E1 and MAb PARV-19 with sample extracts. Western blot was performed to compare the antigenic protein banding patterns in cooked fish extracts using these two MAbs. Results showed that MAb 3E1 cross reacted with majority of tested fish species and recognized a thermal-stable protein with a molecular weight range of parvalbumin in the extracts. Moreover, ELISA and western blot results revealed that both MAbs 3E1 and PARV-19 had almost identical reaction patterns to the fish species tested. The antigenic protein banding pattern in various fish species blotted by MAb 3E1 corresponds to the molecular weights of parvalbumins recognized by PARV-19. Additionally, both antibodies recognized exactly the same antigenic protein, parvalbumin, but their epitopes (binding sites) overlaped to the extend causing inhibitive binding on the protein. However, screening with non-finfish extracts revealed MAb 3E1 to be strictly finfish specific, while PARV-19 cross-reacted with frog, rat and rabbit extracts. The results obtained in this study clearly indicate that MAb 3E1 is specific to fish parvalbumin. It would, therefore, be a useful probe for investigating the major fish allergen in both raw and processed food. / A Thesis submitted to the Department of Nutrition, Food and Exercise Sciences in partial fulfillment of the requirements for the degree of Master of
Science. / Summer Semester, 2008. / June 23, 2008. / Monoclonal Antibody, Fish Allergen, Parvalbumin, Immunoassay, Heat-Stable Protein / Includes bibliographical references. / Yun-Hwa Peggy Hsieh, Professor Directing Thesis; Kenneth H. Roux, Outside Committee Member; Cathy W. Levenson, Committee Member.
| Identifer | oai:union.ndltd.org:fsu.edu/oai:fsu.digital.flvc.org:fsu_182520 |
| Contributors | Gajewski, Kamil (authoraut), Hsieh, Yun-Hwa Peggy (professor directing thesis), Roux, Kenneth H. (outside committee member), Levenson, Cathy W. (committee member), Department of Nutrition, Food, and Exercise Science (degree granting department), Florida State University (degree granting institution) |
| Publisher | Florida State University, Florida State University |
| Source Sets | Florida State University |
| Language | English, English |
| Detected Language | English |
| Type | Text, text |
| Format | 1 online resource, computer, application/pdf |
| Rights | This Item is protected by copyright and/or related rights. You are free to use this Item in any way that is permitted by the copyright and related rights legislation that applies to your use. For other uses you need to obtain permission from the rights-holder(s). The copyright in theses and dissertations completed at Florida State University is held by the students who author them. |
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