Allergen Tropomyosin Research Articles

Combination of Structural Analysis and Proteomics Strategy Revealed the Mechanism of Ultrasound-Assisted Cold Plasma Regulating Shrimp Allergy.

Allergic incidents of crustacean aquatic products occur frequently, and tropomyosin (TM) is the main allergen. Therefore, it is worthwhile to develop technologies to efficiently reduce the allergenicity of TM. In this study, ultrasound-assisted cold plasma (UCP) treatment was used to regulate shrimp allergy. The remarkable changes in TM structure were substantiated by alteration in secondary structure, reduction in sulfhydryl content, change in surface hydrophobicity, and disparity in surface morphology. The IgE and IgG binding ability of TM significantly decreased by 52.40% and 46.51% due to UCP treatment. In the Balb/c mouse model, mice in the UCP group showed most prominent mitigation of allergic symptoms, proved by lower allergy score, changes in levels of TM-specific antibodies, and restoration of Th1/Th2 cytokine imbalance. Using a proteomics approach, 439 differentially expressed proteins (DEPs) in the TM group (vs phosphate-buffered saline group) and 170 DEPs in the UCP group (vs TM group) were determined. Subsequent analysis demonstrated that Col6a5, Col6a6, and Epx were potential biomarkers of TM allergy. Moreover, Col6a5, Col6a6, Dcn, and Kng1 might be the target proteins of UCP treatment, while PI3K/Akt/mTOR might be the regulated signaling pathway. These findings proved that UCP treatment has great potential in reducing TM allergenicity and provide new insights into the development of hypoallergenic shrimp products.

Modulating allergenicity of prawn tropomyosin (penaeus chinensis) via pulsed electric field-induced conformational changes

The effect of electric field intensities (EFIs, 5–20 kV/cm) and treatment times (0.5–2 h) on allergenicity and spatial conformation of prawn tropomyosin was evaluated. The results demonstrated that the IgG and IgE binding capacity of tropomyosin maximally increased by 24.34 % and 29.16 % respectively, followed by a subsequent decrease after 20 kV/cm treatment for 1 h. Interestingly, 5–10 kV/cm treatments significantly decreased the α-helix content (P < 0.05) and fluorescence intensity, while 20 kV/cm treatment promoted extensive spiralization, resulting in a tightly packed structure. The increased flexibility further exposed the hydrolysis sites and strengthened the gastrointestinal digestibility of tropomyosin. Additionally, molecular dynamic simulation indicated that extended EFIs increased structural flexibility and depolymerized the tropomyosin dimers through destroying intermolecular hydrogen bonds (formed within arginine and glutamate), which allowed tropomyosin to be easily recognized by IgG/IgE. Whereas, decrease of solvent-accessibility surface area (SASA), hydrophobic surface area induced conformation folded and caused epitopes masked.

Assessment of a crustacean allergen in fishery products using a nanomodified label-free voltammetric immunosensing approach

In this work, a label-free immunosensor for the rapid analysis of the crustacean allergen tropomyosin (TPM) is presented. A screen-printed carbon electrode (SPCE) was (bio)modified with oxidized carbon nanotube/manganese(II) ferrite nanomaterial (CNT_O@MnFe2O4) and monoclonal antibodies. The immune interaction with the allergenic protein was detected using a redox probe ([Fe(CN)6]3−/4−) by differential pulse voltammetry. The CNT_O@MnFe2O4 led to a 4-fold increase of the analytical signal compared to the unmodified SPCE, indicating the usefulness of this material in TPM analysis. Moreover, the detection potential was reduced (i.e., closer to 0 V), which increased the selectivity of the sensor even further. The immunosensor enabled a fast analysis time (1 h), providing a limit of detection of 1.6 ng mL−1, and precise results (coefficient of variation = 6.5 %). The analysis of ten commercial foodstuffs was accomplished, and the results were validated with an ELISA assay. The highest TPM concentrations were quantified in shrimp species (M. affinis > P. latisulcatus > P. indicus) and low allergen levels were found in surimi lobster, clam, fresh calamari, and mayonnaise. The simplicity of the sensor makes it a good choice for quality control and food safety assessment in industrial settings.

Effects of Glycosylation Combined with Phosphate Treatment on the Allergenicity and Structure of Tropomyosin in Litopenaeus vannamei.

Tropomyosin (TM) is the main allergen in shrimp (Litopenaeus vannamei). In this study, the effects of allergenicity and structure of TM by glycosylation (GOS-TM), phosphate treatment (SP-TM), and glycosylation combined with phosphate treatment (GOS-SP-TM) were investigated. Compared to GOS-TM and SP-TM, the IgG/IgE binding capacity of GOS-SP-TM was significantly decreased with 63.9 ± 2.0 and 49.7 ± 2.7%, respectively. Meanwhile, the α-helix content reduced, surface hydrophobicity increased, and 10 specific amino acids (K30, K38, S39, K48, K66, K74, K128, K161, S210, and K251) were modified by glycosylation on six IgE linear epitopes of GOS-SP-TM. In the BALB/c mice allergy model, GOS-SP-TM could significantly reduce the levels of specific IgE, IgG1, and CD4+IL-4+, while the levels of IgG2a, CD4+CD25+Foxp3+, and CD4+IFN-γ+ were increased, which equilibrated Th1 and Th2 cells, thus alleviating allergic symptoms. These results indicated that glycosylation combined with phosphate treatment can provide a new insight into developing hypoallergenic shrimp food.

Insight into the allergenicity of digestion-resistant epitopes in Antarctic krill (Euphausia superba) tropomyosin

The allergenic potential of four digestion-resistant linear epitopes (EADRKYDE, ERAEERAEAG, RSVQKLQKEVDR, and VNEKEKYKGI) in Antarctic krill tropomyosin was investigated through in vivo and in vitro experiments. Mice induced by the four epitopes exhibited obvious clinical allergic symptoms, with significant increases in serum specific antibody levels and blood lymphocytes, neutrophils, and eosinophils (p < 0.05). Additionally, mast cells exhibited pronounced degranulation, as well as increased release of histamine and mMCP-1, and enhanced vascular permeability. Mice challenged with the four epitopes further displayed severe pathological damage. In contrast to the antagonistic effect of interleukin-4 and interferon-γ in the plasma of mice in previous studies, stimulation of the epitopes resulted in immune system dysregulation in mice, with significant increases in the levels of interleukin-4 and interferon-γ to 110.03 pg/mL and 103.59 pg/mL, respectively. The enhanced binding ability of the four epitopes to IgE was also indicated in vitro. The four digestion-resistant epitopes are the major factors to the allergenicity of tropomyosin and have strong sensitizing abilities. This study provides a theoretical basis for using site directed mutagenesis to reduce the allergenicity of tropomyosin in Antarctic krill.

Identification of the pan-allergen tropomyosin from the common bed bug (Cimex lectularius)

Tropomyosin is a muscle contraction protein documented across all animal life. Despite its ubiquity, its unique structure in invertebrates leads to allergic responses in humans that vertebrate tropomyosin does not. High degrees of homology can explain cross-reactivity between tropomyosin derived from distantly-related arthropod species and establishes tropomyosin as a panallergen. Given this cross-reactivity and that they are commonly found in high numbers indoors, research on the potential of the common bed bug (Cimex lectularius L.) to contribute tropomyosin to the indoor environment is needed. Therefore, we investigated tropomyosin homology between bed bugs and known tropomyosin allergens from other taxa, tropomyosin in bed bug bodies, feces, and exuviae (cast skins), tropomyosin persistence over time, and impacts of common bed bug treatment strategies on detectable tropomyosin. Tropomyosin was detected in mechanically fractured bed bug cadavers and was detectable in bed bugs cadavers aged for 18 months. Additionally, a survey of pest management professionals showed dead bed bugs are not cleaned up following treatment. As such, dead bed bugs could act as tropomyosin reservoirs following bed bug treatment and exposure to tropomyosin from bed bugs could sensitize individuals and lead to increased responses to other arthropod tropomyosin.

Unlocking the potential of waxworm (Galleria mellonella) proteins: Extraction, fractionation, and protein quality assessment

Edible insects are increasingly recognized as an alternative and sustainable protein source to meet future protein demands. Owing to its significant protein content (33.43%), the edible insect Waxworm (Galleria mellonella) was characterized in this study for its protein extractability, structural properties, and protein quality. Protein extractability under various pH levels and ionic strengths was evaluated. Subsequently, Osborne fractionation was employed to characterize protein fractions based on their preferential solubilities. The extracted proteins and their respective fractions were visualized by SDS-PAGE, and the most intense bands were excised for proteomics data analysis. Secondary structure, pI, molecular mass, and denaturation temperature were also examined. The quality of waxworm protein was assessed by determining the IVPD, PDCAAS, and amino acid profile. Waxworm proteins exhibited greater stability in alkaline conditions compared to acidic environments. These might also be more sensitive to ionic strength than other alternative proteins, such as those derived from plants. The allergen tropomyosin was identified by SDS-PAGE with an approximate molecular weight of ∼32.5 kDa; however, it can be selectively removed by salt extraction. The secondary structures of waxworm proteins, extracted under various conditions, appeared stable, with α-helix and β-conformation being the predominant conformation. Among the protein fractions, prolamins displayed an α-helix conformation, while albumins, globulins, and glutelins predominantly adopted a β-sheet structure. It was observed that structural features could influence the overall quality of the protein. As for the protein quality, globulins are the protein fraction recommended as these showed the highest EAAI% (924.02), PDCAAS (0.84) and IVPD (76.33%) values.

Effects of different high temperature-pressure processing times on the sensory quality, nutrition and allergenicity of ready-to-eat clam meat

Investigating technologies to control the allergenicity of seafood is particularly important to safeguard consumer health, but there is currently a dearth of research focused on reducing the allergenicity of clam meat. This study aimed to investigate the effects of high temperature–pressure (HTP) processing times (121 °C, 0.14 MPa; 5, 10, 15, 20 min) on the sensory quality, nutrition, and allergenicity of ready-to-eat clam meat. With the extension of HTP time, the hardness of clam meat gradually decreased, the chewiness decreased initially and then increased, and the meat became tender. HTP processing endowed clam meat with abundant esters and aldehydes. Among all the processing groups, the umami and saltiness were better at 15 min, correlating with the highest overall acceptability. Ready-to-eat clam meat contained high-protein nutritional value. Compared with raw clam meat, the tropomyosin allergenicity of clam meat treated with HTP for 15 and 20 min was significantly reduced by 51.9 % and 56.5 %, respectively (P < 0.05). However, there was no significant difference between these two groups. Appropriate HTP processing time might be an efficient condition to reduce the tropomyosin allergenicity of ready-to-eat clam meat and improve its quality, particularly for the time of 15 min. The results of this study could provide a reliable theoretical basis for the development of hypoallergenic clam foods.

A case study on Norwegian commercial harvesting and production of Saccharina latissima (Part 1): Effect of processing seaweed on chemical hazards, allergens, and microbiological hazards

When producing seaweed for human consumption, incorporating a “Food Safety-by-Design approach” can help protect human health and ensure the safety of future consumers. Post-harvest processing of seaweed is one strategy to reduce undesirable compounds. The study aimed to determine the effects of a commercial processing strategy (blanching and fermentation) of Norwegian-harvested Saccharina latissima (sugar kelp) on the presence of prioritized food safety hazards. We compiled and prioritized food safety hazards to be analyzed based on the current impact of the hazard, the analytical feasibility, and the fit of the hazard to the case study site. Twelve prioritized hazards were included in the case study analyses: arsenic (total and inorganic), iodine, cadmium, lead, mercury, copper, zinc, the allergen tropomyosin, Salmonella spp., Bacillus spp., norovirus, and Vibrio spp. We found that blanching was very effective in reducing iodine concentrations but mildly increased the zinc concentration. Fermentation of blanched seaweed further decreased iodine concentrations and had similar effects on inorganic arsenic, cadmium, and zinc. An unintentional increase in copper was observed. Bacillus cereus was detected in a blanched sample and fermented samples. Vibrio alginolyticus seemed to be introduced during blanching, yet fermentation greatly reduced the detection frequency. Our study illustrates the importance of clearly defining the goals of seaweed processing and optimizing the process for food safety hazards. Meanwhile, it is advised to monitor for unintended effects that may occur during processing, such as, in our case, the introduction of copper and pathogens like B. cereus and Vibrio spp. This study serves as an example for seaweed stakeholders on how to consider a Food Safety-by-Design concept and a risk-based approach during seaweed harvest and post-harvest processing.

Colorimetric and surface-enhanced Raman scattering dual-mode lateral flow immunosensor using phage-displayed shark nanobody for the detection of crustacean allergen tropomyosin

Tropomyosin (TM) is the primary allergenic protein responsible for crustacean food allergies, and thus sensitive and rapid methods are required for the screening of crustacean TM in food. In this study, using the phage-displayed shark nanobody (PSN) as a multifunctional biomaterial, we developed a colorimetric and surface-enhanced Raman scattering dual-mode lateral flow immunosensor (CM/SERS-LFI) for competitive detection of crustacean TM. The SERS tag AuMBA@AgNPs with the Raman signal molecule 4-mercaptobenzoic acid (4-MBA) was prepared and immobilized on the PSN to construct the immunoprobe AuMBA@Ag-PSN. The probe can identify free TM that competes with TM on the T-line, and the optimized CM/SERS-LFI enables quantitative analysis of TM using the probe with a limit of detection (LOD) of 0.0026 μg/mL (SERS mode) and 0.0057 μg/mL (colorimetric mode), respectively. Additionally, it can implement a qualitative analysis by the naked eye with a visual LOD of 0.01 μg/mL. The CM/SERS-LFI exhibited excellent performance in the tests of selectivity, accuracy, precision, and stability. Moreover, the method’s effectiveness in the analysis of real samples was confirmed by a commercial ELISA kit. Therefore, the developed CM/SERS-LFI was demonstrated to be a powerful and reliable tool for the rapid and sensitive detection of crustacean TM in food.

Reducing the allergenicity of tropomyosin in shrimp by covalent conjugation with quercetin and chlorogenic acid

The study aimed to assay the allergenicity of shrimp tropomyosin (TM) following covalent conjugation with quercetin (QR) and chlorogenic acid (CA). The structure of the TM-polyphenol covalent conjugates was examined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), circular dichroism (CD), fluorescence, differential scanning calorimetry (DSC), and Fourier Transform infrared spectroscopy (FTIR). Potential allergenicity was evaluated using in vitro and in vivo methods. The results showed that QR and CA induced structural changes in TM through aggregation. RBL-2H3 cell results showed that TM-QR and TM-CA covalent conjugates reduced the release of β-hexosaminidase and histamine, respectively. In the mice model, TM-QR and TM-CA covalent conjugates reduced the level of IgE, IgG, IgG1, histamine, and mMCP-1 in sera. Furthermore, the allergenicity was reduced by suppressing Th2-related cytokines (IL-4, IL-5, IL-13) and promoting Th1-related cytokines (IFN-γ). These research findings demonstrate that the covalent binding of TM with QR and CA, modifies the allergenic epitopes of shrimp TM, thereby reducing its potential allergenicity. This approach holds practical applications in the production of low-allergenicity food within the food industry.

Lateral Flow Immunochromatographic Assay for Competitive Detection of Crustacean Allergen Tropomyosin Using Phage-Displayed Shark Single-Domain Antibody.

The common food allergy crustacean tropomyosin (TM) poses a significant food safety challenge, which requires rapid and sensitive methods for screening TM in food. Herein, the variable new antigen receptor (VNAR) single-domain antibodies specific for the crustacean TM were isolated from a naïve phage-displayed shark VNAR library. Subsequently, a lateral flow immunochromatographic assay (LFIA) based on the gold nanoparticle-labeled phage-displayed shark VNAR (AuNPs@PSV) probe was developed for the detection of TM in food. The AuNPs@PSV-LFIA took 15 min for one test and had a visual limit of detection (vLOD) of 0.1 μg/mL and an instrumental LOD of 0.02 μg/mL. Good selectivity, accuracy, precision, and stability were confirmed for the AuNPs@PSV-LFIA. Moreover, the test results of 21 commercially available food products consisted of the allergen labels and were validated by a commercial ELISA kit. Therefore, this work demonstrated the great potential of VNAR for detecting TM in food by LFIA.

Effect of transglutaminase-catalyzed glycosylation on the allergenicity of tropomyosin in the Perna viridis food matrix.

Food allergy is one of the hot issues in the field of food safety, and there have been a lot of concerns on how to reduce the allergenicity of food allergens. Food processing can change the allergenicity of allergens in the food matrix. In this study, ten IgE linear epitopes of the major allergen tropomyosin (TM) in Perna viridis were identified by bioinformatics prediction and serological experiments. The transglutaminase-catalyzed glycosylation modification sites glutamine, lysine and arginine were highly represented in the IgE linear epitopes of TM. The Perna viridis food matrix was treated with transglutaminase-catalyzed glycosylation. This reaction changed the secondary structure of protein in the food matrix, increased the content of β-sheets and decreased the content of β-turns. The intensity of intrinsic fluorescence and surface hydrophobicity were reduced. The IgE-binding activity of TM in the food matrix was reduced by modifying seven amino acid residues on six IgE linear epitopes. Transglutaminase-catalyzed glycosylation products decreased allergic symptoms in allergic mice, reduced the proportion of CD4+IL-4+ Th2 cells, and increased the proportion of CD4+IFN-γ+ Th1 cells and Treg cells. Mouse serum levels of IgE and IgG1 antibodies in the food matrix and TM were reduced. Therefore, this study provided a theoretical basis for the development of hypoallergenic Perna viridis products.

Comparative analysis of tropomyosin allergenicity in three different species of molluscs: insights into the role of amino acid composition in IgE epitopes.

Limited research has been conducted on the differences in allergenicity among Alectryonella plicatula tropomyosin (ATM), Haliotis discus hannai tropomyosin (HTM), and Mimachlamys nobilis tropomyosin (MTM) in molluscs. Our study aimed to comprehensively analyze and compare their immunoreactivity, sensitization, and allergenicity while simultaneously elucidating the underlying molecular mechanisms involved. We assessed the immune binding activity of TM utilizing 86 sera from allergic patients and evaluated sensitization and allergenicity through two different types of mouse models. The dot-blot and basophil activation test assays revealed strong immunoreactivity for HTM, ATM, and MTM, with HTM exhibiting significantly lower levels compared to ATM. In the BALB/c mouse sensitization model, all TM groups stimulated the production of specific antibodies, elicited IgE-mediated immediate hypersensitivity responses, and caused an imbalance in the IL-4/IFN-γ ratio. Similarly, in the BALB/c mouse model of food allergy, all TM variants induced IgE-mediated type I hypersensitivity responses, leading to the development of food allergies characterized by clinical symptoms and an imbalance in the IL-4/IFN-γ ratio. The stimulation ability of sensitization and the severity of food allergies consistently ranked as ATM > MTM > HTM. Through an in-depth analysis of non-polar amino acid frequency and polar hydrogen bonds, HTM exhibited higher frequencies of non-polar amino acids in its amino acid sequence and IgE epitopes, in comparison with ATM and MTM. Furthermore, HTM demonstrated a lower number of polar hydrogen bonds in IgE epitopes. Overall, HTM exhibited the lowest allergenic potential in both allergic patients and mouse models, likely due to its lower polarity in the amino acid sequence and IgE epitopes.

Development of a fluorescent multiplexed lateral flow immunoassay for the simultaneous detection of crustacean allergen tropomyosin, sarcoplasmic calcium binding protein and egg allergen ovalbumin in different matrices and commercial foods

A quantum dot (QD) based multiplexed lateral flow immunoassay (xLFIA) for the simultaneous detection of egg allergen ovalbumin, crustacean allergen tropomyosin (TM) and sarcoplasmic calcium binding protein (SCP) was developed in this study. QD-labeled rabbit anti-ovalbumin, SCP and TM antibodies were applied as fluorescent detection probes. The chromatography system was optimized to reduce the mutual interference of different test lines. Visual and instrumental detection limits of the xLFIA were 0.1 and 0.05 μg/mL for SCP, both 0.05 μg/mL for ovalbumin and both 0.5 μg/mL for TM. As low as 0.10 % crab powder, 0.01 % egg white powder and 0.05 % shrimp powder could be detected in all three model foods using xLFIA. Besides, the xLFIA detection results of 23 of 28 commercial foods were consistent with ingredient labels. These findings indicate that the developed xLFIA is a practical tool for point-of-care detection of egg and crustacean allergens in processed and commercial foods.

A SILAC-based accurate quantification of shrimp allergen tropomyosin in complex food matrices using UPLC-MS/MS

The carryover of trace allergens in complex food matrices poses challenges for detection techniques. Here, we demonstrate an accurate UPLC-MS/MS quantification assay for the shrimp allergen tropomyosin with a full-length isotope-labelled recombinant tropomyosin (TM-I) internal standard in complex food matrices. The TM-I, expressed based on the SILAC technique, exhibited a high isotope labelling ratio (>99%), purity, and alignment with the natural sequence. This method determined the tropomyosin ranging from 0.2 to 100 ng/mL. Mean recoveries ranged from 89 to 116%, with intra- and inter-day RSDs below 12%, for three signature peptides across three types of commercially processed food matrices. The limits of quantitation were 1 μg/g in pop food and sauce, and 10 μg/g in surimi product, respectively. This study supports the use of recombinant full-length isotope-labelled proteins rather than stable-isotope labelling peptides as internal standards to achieve more accurate quantitation of food allergens as the digestion error is corrected.

The effect of papain hydrolysis on tropomyosin levels in shrimp

Shrimp allergy is a serious public health problem. Epidemiological studies indicated the main cause of food allergy is the ingestion of shrimp and crab. Consequently, reducing the risk of shrimp and crab allergy is a major research focus. Few studies have demonstrated the effect of enzyme hydrolysis on reduction of shrimp allergens in food. In this study, we used papain, a commonly used enzyme in the food industry, as an ingredient to decrease levels of allergen tropomyosin in shrimp. 0–50U of papain was used to treat the shrimp meat, and then heated to measure the levels of tropomyosin, and determined the change of the protein secondary structure. The results showed that 20U of papain decrease the tropomyosin levels up to 80 %, which was further enhanced when combined with 3 min of heating. Furthermore, the result of Fourier-transform infrared spectroscopy (FTIR) showed alteration of secondary protein structure after the processing. The processing developed in this study may be an effective method used to change the levels and structure of tropomyosin.

Digestion-Resistant Linear Epitopes as Dominant Contributors to Strong Allergenicity of Tropomyosin in Antarctic Krill (Euphausia superba).

Although tropomyosin has been identified as a major allergen in Antarctic krill, the digestive fate of Antarctic krill tropomyosin and its relationship with allergenicity are unknown. In this study, Antarctic krill tropomyosin was administered to BALB/c mice via both gavage and intraperitoneal injection to explore its sensitizing and eliciting capacity, and its digestion products were analyzed for structural changes and digestion-resistant linear epitopes. Mice gavaged with tropomyosin exhibited lower levels of specific IgE and IgG1, mast cell degranulation, vascular permeability, and anaphylaxis symptoms than those in the intraperitoneal injection group. This may be due to the destruction of macromolecular aggregates, loose expansion of the tertiary structure, complete disappearance of α-helix, and significant changes in molecular force upon the digestion of tropomyosin. Nevertheless, the intragastric administration of Antarctic krill tropomyosin still triggered strong allergic reactions, which was attributed to the existence of seven digestion-resistant linear epitopes (Glu26-His44, Thr111-Arg125, Glu157-Glu164, Glu177-Gly186, Val209-Ile225, Arg244-Arg255, and Val261-Ile270).

Effects of AAPH on structure, allergenicity and physicochemical properties of shrimp myofibrillar protein

Lipid peroxidation has been shown to associate with meat quality by affecting physicochemical properties of myofibrillar proteins. In this work, shrimp myofibrillar protein (MP) was oxidized using 2,2′-azobis (2-amidinopropane) dihydrochloride (AAPH), which could generate peroxyl radicals in lipid peroxidation. Results showed that increasing AAPH concentration led to an increase in carbonyl groups and a decrease in free amino and sulfhydryl thiol groups in MP. Secondary structural analysis of MP revealed a conversion of α-helix to β-sheet content after AAPH treatment. SDS-PAGE showed the band intensity of tropomyosin was reduced after AAPH treatment, indicating a reduced allergenicity of tropomyosin. Moderate oxidation (1–10 mmol/L AAPH) promoted the formation of a dense network of MP gel, resulting in enhanced textural and rheological properties. However, excessive oxidation (10–20 mmol/L AAPH) weakened the MP gel strength and decreased its water-holding capacity. These findings provide theoretical guidance for developing shrimp surimi products with desirable textures.

In vivo biological analysis of cold plasma on allergenicity reduction of tropomyosin in shrimp

In vivo biological regulations of the allergenicity of tropomyosin (TM) treated by cold plasma (CP) were investigated by in vivo mouse model. The sensitization models of Balb/c mice were successfully established. CP treatment reduced the allergic symptoms of mice and regulated the Th1/Th2 balance to prevent allergy by activating Treg cells, which was deduced by serum and cytokines analysis. For intestinal flora analysis, allergy occurrence was accompanied by the decreased species abundance and the increased species diversity of intestinal flora. The significant species composition difference between the TM group and the PBS group showed a possible connection between bacterial diversity and allergy. Furthermore, Firmicutes, Bacteroidetes, Parabacteroides, Alloprevotella, Bacteroides, and Lachnospiraceae could relate to allergy occurrence. Intestinal section analysis suggested that allergy occurrence was accompanied by the damaged intestinal structure, and CP treatment could relieve the damage caused by an allergy.