The type I hypersensitivity reaction mediated by IgE antibodies is a typical clinical feature of allergic diseases, such as asthma, eczema, hay fever, and urticaria. Ovalbumin (OVA) is widely used as an antigen for studying the IgE-mediated allergic reaction in mice. Chondrex, Inc. provides allergenic and non-allergenic mouse anti-OVA IgE and IgG monoclonal antibodies, as well as low endotoxin OVA for studying in vivo hyper-sensitivity reactions. Additionally, Chondrex, Inc. manufactures Mouse Anti-OVA Antibody Assay Kits, Cytokine Detection ELISAs, and Chemokine Detection ELISAs for evaluating immune responses in OVA-Induced and Anti-OVA IgE-Induced Asthma models.
To learn more about the OVA-Induced Asthma Model and the Anti-OVA IgE-Induced Asthma Model, please proceed to the Table fo Contents below.
If you have any questions about Chondrex, Inc.’s Mouse Allergic Disease Models, please contact us at firstname.lastname@example.org.
Note: These mouse asthma models are highly reproducible due to OVA’s high purity and antigenicity. However, different allergens, such as house dust mites (HDM), may also be involved in the pathogenesis of human allergic asthma. Chondrex, Inc, also provides HDM extracts and Mouse Anti-HDM Antibody Assay Kits for studies using the House Dust Mite-Induced Asthma Model.
Mouse Anti-OVA IgE and IgG Monoclonal Antibodies
|Product||In Vitro Application||In Vivo Application||Quantity||Catalog #||Price (USD)|
|Mouse Anti OVA Monoclonal IgG1 Antibody L71||No||1 mg, lyophilized||3008||406.00|
|Mouse Anti OVA Monoclonal IgE Antibody E-C1||Mast Cell Activation||Hyper-Sensitivity Reaction||1 mg, lyophilized||3006||812.00|
|Mouse Anti OVA Monoclonal IgE Antibody E-G5||No||No||1 mg, lyophilized||3007||406.00|
Ovalbumin From Chick Egg White
|Product||Catalog #||Price (USD)|
|Low Endotoxin Ovalbumin From Chick Egg White, 10 mg||3022||129.00|
|Ovalbumin From Chick Egg White, 50 mg||30211||42.00|
1. Allergenic Antibodies for Mast Cell Degranulation
Mast cells are degranulated (activated) by cross-linking the two IgE receptors of which two adjacent IgE antibodies bound to the IgE receptors on mast cells capture a polyvalent allergen (Figure 1a). Alternatively, immune-complexes of allergens with IgG antibodies can also degranulate mast cells by bridging the allergens bound to IgE antibodies on mast cells (Figure 3b). Therefore, not only IgE, but also IgG against allergens play roles in development of allergic reactions.
Figure 1. Two mechanisms of mast cell-bound IgE cross-linking. a) Cross-linkage by a single allergen and b) Cross-linkage by a single IgG antibody to two allergen molecules.
2. Asthma models
In 2013, approximately 22.6 million people in the United States (7.3% of the population) had asthma, including 6.1 million children and 16.5 million adults (1). In order to develop new therapeutics for allergic asthma that will improve patient quality of life, reliable animal models of human asthma are required. The following mouse asthma models demonstrate comparable human asthma symptoms, such as allergen-induced early and late asthmatic responses, airway hyper-responsiveness (AHR), and airway inflammation.
a) OVA-Induced Asthma model
Ovalbumin (OVA) has traditionally been used to induce mouse allergic asthma models (2-5). Mice are intraperitoneally immunized with OVA adsorbed to an aluminum hydroxide adjuvant (Alum) on days 0 and 14. During this sensitization phase, the mice produce anti-OVA IgE antibodies which bind IgE receptors on mast cells. After this sensitization, the mice are intratracheally challenged with OVA, resulting in OVA cross-linked IgE on mast cells, leading to degranulating mast cells. Mice then develop clinical features of asthma (Figure 2 top).
Unfortunately, it is difficult to evaluate therapeutics efficiently using this model since asthma induction takes several weeks. In addition, a highly purified OVA is required for sensitization because endotoxin contamination in OVA may reduce the Th2 immune reaction eliciting IgE antibodies. Therefore, we recommend using low endotoxin OVA for high reproducibility.
Many protocols for developing acute and chronic OVA-induced asthma models are published (6). We recommend establishing and optimizing a protocol according to your study needs.
b) Anti-OVA IgE Asthma Model
In order to improve the efficiency of asthma research, Chondrex, Inc. introduces the Anti-Ovalbumin IgE-Induced Asthma model. Because directly injecting anti-OVA IgE antibodies bypasses the anti-OVA IgE development step, mice can develop allergic reactions in one week rather than in several weeks required by the traditional OVA-induced asthma model (Figure 2 bottom) (1, 2). Mice are administrated anti-OVA IgE monoclonal antibodies intraperitoneally and challenged intratracheally with OVA several times. The clinical features of asthma in mice should appear after the fourth sensitization/challenge cycle (Figure 2 bottom). We offer a detailed protocol for the Anti-OVA IgE Asthma model (refer here).
Note: In recent references, the mouse anti-OVA monoclonal IgE antibody, E-C1 (catalog# 3006), has been referred to by the clone name "OE-1" and mouse anti OVA monoclonal IgG antibody, L71 (catalog# 3008), has been referred to by the clone name "O1-10".
Figure 2. Comparing a typical protocol for an acute OVA-induced asthma model and a protocol for the anti-OVA IgE Asthma Model.
c) Evaluating Ovablumin Asthma Models
Both the OVA-Induced Asthma model and the Anti-OVA IgE Asthma model develop similar disease symptoms, therefore similar disease evaluation methods can be used to determine the effect of any experimental therapeutics.
1) Inflammatory Cell Counts in BALF and Specific airway resistance (sRaw) (7, 8)
Successful asthma induction in the both the OVA-Induced Asthma and Anti-OVA IgE model will increase immune cell counts in bronchoalveolar lavage fluid (BALF) (Figure 3 Left). Interestingly, the Anti-OVA IgE model does not induce eosinophilia as seen in the OVA-Induced Asthma model and clinical asthma. However, further anti-OVA IgE administration and OVA challenges can induce eosinophilia. In addition, effects of the Inhibitor in Anti-OVA IgE model using the E-C1 IgE antibody can be evaluated with cell counts (Figure 3 Right).
Specific airway resistance (sRaw) assayed by a two-chambered, double flow plethysmograph system (Pulmos-I; M.I.P.S) increased during disease progression; the increase in sRAW was prevented with the C3a receptor antagonist (inhibitor) treatment.
Figure 3. Comparison of OVA/Alum and Anti-OVA IgE (E-C1)-induced Asthma Model.
2) Cytokine and Chemokine Analysis
Successful asthma induction leads to higher levels of Th2 response related cytokines (IL-4, IL-5, IL-13, TNF-α) and lower levels of Th1 response related cytokines (IFN-γ). Not only quantifying these cytokines in samples work to evaluate the effect of experimental therapeutics (9), but also quantifying chemokine levels work for the purpose (10). For this purpose, Chondrex Inc. provides Mouse Cytokine Detection Assay Kits and Mouse Chemokine Detection Assay Kits.
3) Anti-OVA IgE and IgG antibody levels
High levels of serum anti-OVA IgE antibodies are required to develop the OVA-Induced Asthma model. Therefore, measuring the antibody levels of anti-OVA IgE and IgG can be markers for determining effective sensitization and treatments. Regarding anti-OVA IgG levels, anti-OVA IgG1 antibodies are related with Th2 response, and anti-OVA IgG2a or IgG2b antibodies are related with Th1 response. Chondrex, Inc. provides Anti-Ovalbumin Antibody Assay Kits to quantify specific subtype and subclass immunoglobulins against OVA in mouse samples.
3. A Mouse Nasal Hypersensitivity Model (11)
Dr. Saka et al. introduced a new protocol for inducing nasal hypersensitivity in C57BL/6 mice using Chondrex, Inc.’s anti-OVA IgE monoclonal antibody, E-C1, along with an OVA induced whole-body sensitization model (Figure 4). For more information on this protocol, please see our report here.
Figure 4. Protocols of the the whole-body sensitization model (A) and nasal hypersensitivity model (B)
4. A Mouse Footpad Delayed-type Hypersensitivity Model
The anti-OVA IgE monoclonal antibody, clone EC-1, can induce severe hypersensitivity reactions in vivo. Please refer to our protocol. Balb/c mice received 10 μg of monoclonal antibody, E-C1 or E-G5 by IV injection, then were challenged by OVA (50 μg) or aggregated OVA (Agg-OVA, 50 μg) through intradermal injections at footpads 24 hours later. Food pad thickness (mm) was determined by a Loop Handle Dial Thickness Gauge.
E-C1 antibody solely induced swelling of footpad in vivo (Figure 5), suggesting that E-C1 antibody might recognize the repetitive epitopes of OVA. Furthermore, a combination of E-C1 and aggregated OVA carrying multiple epitopes increased footpad swelling, likely due to the higher levels of cross-linkage formation of IgE molecules on mast cell surfaces (12). On the other hand, anti-OVA IgE antibody, E-G5, failed to induce these allergic reactions because E-G5 recognizes a single epitope on OVA (Figures 5). This suggests that the E-G5 antibody can be used as a control.
Figure 5. Foot pad hypersensitivity reaction induced by E-C1 in Balb/c mice.
5. Mast Cell Degranulation Assay
The anti-OVA IgE monoclonal antibody, clone EC-1, can also induce mast cell degranulation in vitro(Please see our protocol). Rat Basophil Leukemia Cells cultured in 24-well plates were sensitized by anti-OVA monoclonal antibodies, E-C1 (1 μg/ml), E-G5 (1 μg/ml) and L71 (50 μg/ml), respectively, and then activated by OVA (5 μg/ml). As a positive control, a combination of anti-DNP IgE antibodies (1:200 dilution of culture media) and DNP-HSA (0.05 μg/ml) was used. E-C1 alone can degranulate mast cells in an antigen-specific manner, whereas E-G5 (IgE) and L-71 (IgG1) failed to degranulate mast cells under the same conditions. In addition, Chondrex’s IgE subtype control can be used as a negative control for these assays.
Figure 6. Degranulation of mast cells by monoclonal IgE antibodies: a) Comparison of E-C1 and E-G5 antibodies b) Dose response of E-C1