Mouse Anti-Japanese Cedar Pollen (JCP) Antibody Assay Kits

Japanese cedar (Cryptomeria japonica) pollen (JCP) represents a major seasonal type I allergy. Cry j 1 and Cry j 2 are two major allergens present in JCP that can trigger humoral immune responses. Cry j 1 is a pectate lyase, whereas Cry J 2 is a polymethylgalacturonase (1). Sensitization induces plasma cells to secrete allergen-specific immunoglobulin E (IgE), which binds high-affinity FceRI receptors on basophils and mast cells, mediating immediate histamine release upon subsequent allergen exposure (2-4). 

Chondrex, Inc provides various mouse ELISA kits to assay antibodies against JCP extract . The IgG and IgG1 kits employ anti-Cry j 1 antibodies as standards, but can determine antibodies against total JCP allergens (5, 6). Chondrex, Inc. also provides a mouse anti-JCP antibody as well as other antibody ELISA kits against other allergens such as OVA, mites, peanut allergens, gluten allergens, etc.  

For more information, please contact support@chondrex.com.
 

Japanese Cedar Pollen Allergy Protocols

1.      Allergen Extract Preparation

The soluble extract of Japanese cedar (Cryptomeria japonica) pollen, which contains the major allergens Cry j 1 and Cry j 2, is prepared via alkaline bicarbonate extraction. Pollen is mixed with 20 volumes (w/v) of 0.125 M sodium bicarbonate (pH 8.0–8.3) and incubated at 20°C for 30 minutes with gentle agitation. Following centrifugation at 10,000 × g for 20 minutes at 4°C, collect the soluble supernatant, dialyze (MWCO 10 kDa) against distilled water overnight, and lyophilize for storage. Prior to use, reconstitute JCP powder in a pH 8.0 buffer of choice . Protein concentration is determined by bicinchoninic acid (BCA) assay.

2.      Animals

Female BALB/c mice, aged 6–10 weeks, can be used for all experimental models due to their pronounced Th2-biased immune profile and robust production of allergen-specific IgE and IgG1 antibodies.

3.      Protocols

3-1. Allergic Conjunctivitis Model (7, 8)

Systemically sensitize mice via intraperitoneal (i.p.) injection with 0.2 mg JCP extract adsorbed to 2.5 mg aluminum hydroxide (alum) per mouse on days 0 and 7. On days 14 and 16, administer a topical ocular challenge of JCP suspension (1.2 mg in 2 µl per eye). At 15 minutes post-challenge on Day 16, clinical ocular symptoms are scored in a blinded manner across 4 parameters (0–3 scale): lid edema/swelling, tearing/discharge, chemosis, and conjunctival redness. At 24 hours post-challenge (Day 17), globes and conjunctival tissues are harvested for H&E staining to quantify infiltrating eosinophils in the lamina propria mucosa and blood is collected for JCP-specific IgE and IgG antibody ELISAs (Figure 1).

Figure 1: Timeline of Systemic Sensitization and Ocular Challenge Protocol.

3-2. Allergic Rhinitis Models (9-12)

Systemically sensitize the mice via i.p. injection of 10–50 µg JCP extract adsorbed to alum per mouse on days 0, 7, and 14. Administer intranasal challenges (10–50 µg JCP per mouse) daily from days 21 to 25.  On Day 26, nasal cavities are collected and fixed and decalcified for coronal sectioning and mucosal eosinophil counts, and blood is collected for plasma/serum JCP–specific IgE and IgG antibody ELISAs (Figure 2).

Figure 2: Timeline of Systemic Sensitization and Intranasal Provocation Protocol.

Alternatively, utilize an adjuvant-free, direct airway model by exposing the external nares to native dry pollen for 1 minute daily across three 5-day cycles (days 0–19), followed by intranasal provocation on days 26–30. Quantify sneezing and nasal rubbing post-challenge, and collect nasal lavage fluid (NALF) for differential cell counts and mucosal secretory IgA and blood for JCP-specific IgE/IgG antibody ELISAs (Figure 3).

Figure 3: Timeline of Adjuvant-Free Direct Airway Sensitization and Provocation Protocol.

3-3. Allergen-Specific Tolerance Model (13, 14)

To evaluate oral immunotherapy, administer a therapeutic agent (0.1–0.3 mg potential vaccine per mouse per day) to the mice continuously from day 0 to 40. Systemically sensitize the mice via i.p. injection with 5 µg JCP adsorbed to alum per mouse and subcutaneous boosting with 5 µg JCP per mouse on days 14 and 28, respectively. Administer intranasal challenges from days 36 to 40 and conduct serial blood sampling to evaluate JCP-specific IgE and IgG antibody levels (Figure 4).

Figure 4: Timeline of Oral Administration, Systemic Immunization, and Nasal Challenge Protocol.

References

1. T. Ohtsuki, Y. Taniguchi, K. Kohno, S. Fukuda, M. Usui, M. Kurimoto, Cry j 2, a major allergen of Japanese cedar pollen, shows polymethylgalacturonase activity. Allergy 50(6), 483-8 (1995).
   
2. T. Osada, M. Okano, Japanese cedar and cypress pollinosis updated: New allergens, cross-reactivity, and treatment. Allergol. Int. 70, 281–290 (2021).
   
3. M. Sakaguchi, M. Hashimoto, H. Nigi, H. Yasueda, Y. Takahashi, M. Watanabe, T. Nagoya, Y. Taniguchi, M. Kurimoto, S. Inouye, Epitope specificity of IgE antibodies to a major allergen (Cry j 1) of Japanese cedar pollen in sera of humans and monkeys with pollinosis. Immunology 91, 161–166 (1997).
   
4. D. Utsumi, K. Suzuki, H. Matsuhara, E. Kuroda, K. J. Ishii, K. Ohashi-Doi, Enhancement of immune modulation by toll-like receptor 9 adjuvant combination subcutaneous immunotherapy for Japanese cedar pollinosis. Biol. Pharm. Bull. 48, 1191–1198 (2025).
   
5. K. Fukuda, W. Ishida, Y. Harada, Y. Wakasa, H. Takagi, F. Takaiwa, A. Fukushima, Prevention of allergic conjunctivitis in mice by a rice-based edible vaccine containing modified Japanese cedar pollen allergens. Br. J. Ophthalmol. 99, 705–709 (2015).
   
6. Y. Su, E. Romeu-Bonilla, T. Heiland, Next generation immunotherapy for tree pollen allergies. Hum. Vaccin. Immunother. 13, 2402–2415 (2017).
   
7. K. Fukuda, W. Ishida, Y. Harada, Y. Wakasa, H. Takagi, F. Takaiwa, A. Fukushima, Efficacy of oral immunotherapy with a rice-based edible vaccine containing hypoallergenic Japanese cedar pollen allergens for treatment of established allergic conjunctivitis in mice. Allergol. Int. 67, 119–123 (2018).
   
8. W. Ishida, T. Kishimoto, F. Takaiwa, K. Fukuda, Prophylactic and therapeutic effects of oral immunotherapy on birch pollen-induced allergic conjunctivitis in mice with a rice-based edible vaccine expressing a hypoallergenic birch pollen allergen. Cells 10, 3361 (2021).
   
9. S. Awane, K. Nishi, M. Ishida, M. Nagano, K. Hashiguchi, A. Fujii, T. Sugahara, Inhibitory effect of Japanese black vinegar on IgE-mediated degranulation of RBL-2H3 cells and a murine model of Japanese cedar pollinosis. Cytotechnology 70, 961–974 (2018).
   
10. T. Hamasaki, K. Teruya, Y. Katakura, Effect of Hita Tenryo WaterTM, a natural mineral water, on allergic symptoms induced by cedar in mice. Heliyon 10, e26915 (2024).
    
11.  I. Kingetsu, N. Ohno, N. Hayashi, M. Sakaguchi, S. Inouye, S. Saito, Common antigenicity between Japanese cedar (Cryptomeria japonica) pollen and Japanese cypress (Chamaecyparis obtusa) pollen, I. H-2 complex affects cross responsiveness to Cry j 1 and Cha o 1 at the T- and B-cell level in mice. Immunology 99, 625–629 (2000).
    
12. N. Ohno, T. Ide, M. Sakaguchi, S. Inouye, S. Saito, Common antigenicity between Japanese cedar (Cryptomeria japonica) pollen and Japanese cypress (Chamaecyparis obtusa) pollen, II. Determination of the cross-reacting TT-cell epitope of cry j 1 and cha o 1 in mice. Immunology 99, 630–634 (2000).
    
13. Y. Kawabe, Y. Hayashida, K. Numata, S. Harada, Y. Hayashida, A. Ito, M. Kamihira, Oral immunotherapy for pollen allergy using T-cell epitope-containing egg white derived from genetically manipulated chickens. PLoS One 7, e48512 (2012).
    
14. F. Takaiwa, Next-generation allergen-specific immunotherapy for Japanese cedar pollinosis using molecular approaches. ImmunoTargets Ther. 10, 213–224 (2021).