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Background: Sweat aggravates atopic dermatitis (AD). In patients with AD, type-I hypersensitivity to sweat may be shown by histamine release from patients' basophils in response to the semi-purified sweat antigen (QR), and the presence of specific immunoglobulin E (IgE) binding to MGL_1304, the component of QR. However, there has been no information on the immunological changes of type-I hypersensitivity to the sweat antigen in patients with well-controlled AD using topical corticosteroids (TCSs) and/or biologics as treatments., Method: Histamine-releasing tests using patients' basophils and QR and the detection of serum IgE against MGL_1304 and mite allergen Der f 1 were performed in patients with AD who were well controlled by topical TCS with/without dupilumab for 53-96 weeks., Results: In total, 14 patients were enrolled. Seven patients received TCS therapy alone (TCS group), and seven patients received TCS with dupilumab therapy (dupilumab group). In all participants, the level of specific IgE against MGL_1304 decreased after treatments, but histamine release from basophils in response to QR did not show a statistically significant reduction; rather, it increased. In the dupilumab group, all changes in histamine release induced by QR (increase), the IgE level against MGL_1304 (decrease), and that against Der f 1 (decrease) were statistically significant, whereas the TCS group showed no significant change in any of them., Conclusion: The well-controlled condition for 53-96 weeks resulted in no reduction of the hyperreactivity of basophils against in patients with AD, even with the treatment with dupilumab. This study suggests persistent basophil hyperreactivity to sweat antigen over a year or longer., Competing Interests: TN has received a research grant from Sun pharma and Kyowa-Kirin. ST has received a research grant from Sanofi, Maruho, and Taiho Pharma and an honorarium from Novartis. MH has received a research grant from GlaxoSmithKline, Kaken Pharmaceutical, Mitsubishi-Tanabe, Novartis, Sanofi, and Taiho Pharma, honorarium from Ely-Lilly, Kaken-Pharmaceutical, Kyowa-Kirin, Mitsubishi-Tanabe, MSD, Novartis, Sanofi, Taiho Pharma, Teikoku seiyaku, and Uriach. AT has received a research grant from Maruho, Sanofi, Eisai, and Mitsubishi Tanabe Pharma and an honorarium from Maruho, Torii Pharma, Mitsubishi Tanabe Pharma, Sanofi, and Taiho Pharma. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Numata, Takahagi, Ishii, Morioke, Kan, Mizuno, Yanase, Kawaguchi, Tanaka and Hide.)

Background: Many patients with atopic dermatitis and cholinergic urticaria display an immediate-type allergy to autologous sweat. Although the histamine release test (HRT) using semi-purified sweat antigen (QR) was available for the detection of immediate sweat allergy, the existence of HRT low responders could not be disregarded. Furthermore, it has not been established whether the results of the HRT are consistent with the autologous sweat skin test (ASwST). We aimed to compare the HRT and basophil activation test (BAT) for the diagnosis of immediate sweat allergy., Methods: The HRT and BAT were performed on 47 subjects (35 ASwST positive, 12 negative) whose symptoms had worsened on sweating. For the BAT, blood was incubated with QR or crude sweat and CD203c upregulation was assessed. A commercial HRT was performed and histamine release induced by QR was quantified., Results: When excluding non-responders for anti-IgE antibody, the BAT using QR and the HRT had a sensitivity of 100% and 44% and specificity of 75% and 100%, respectively. The BAT and HRT had a positive predictive value of 91.3% and 100% and negative predictive value of 100% and 30%, respectively. The BAT detected 0% non-responders, whereas the HRT identified 22.5%. When using crude sweat for the BAT, the false-positives observed when using QR were not detected., Conclusions: The BAT using QR displayed a higher sensitivity and negative predictive value and a lower number of non-responders compared with the HRT. Furthermore, the BAT using crude sweat can also be an alternative tool for the ASwST., (Copyright © 2019 Japanese Society of Allergology. Production and hosting by Elsevier B.V. All rights reserved.)

Background: Histamine is a crucial mediator in the development of anaphylaxis. Although histamine is promptly degraded because of its short half-life in plasma, basophils, which release histamine, remain in the blood for days. To explore basophils as a potential marker and their involvement in the pathogenesis of anaphylaxis, we evaluated the intracellular histamine concentration and the degree of basophil activation in anaphylaxis patients., Methods: We conducted a case-control study enrolling anaphylaxis patients and healthy controls. Basophil activation was evaluated by flow cytometry using up-regulation of CD203c expression., Results: We enrolled 23 patients and measured their blood histamine concentration. Basophil activation was analyzed in seven of 23 patients. The median intracellular histamine concentrations at admission were significantly lower in patients compared with controls (16.4 ng/mL [interquartile range {IQR}, 2.70 to 34.0] vs. 62.3 ng/mL [IQR, 46.0 to 85.1]; p < 0.0001). The median basophil number at admission was also significantly lower in patients compared with controls (2.21 cell/μL [IQR, 0.75 to 12.3] vs. 21.0 cell/μL [IQR, 19.5 to 28.9]; p = 0.027). CD203c expression was not up-regulated in any of the seven patients in vitro, but it was up-regulated in response to anti-IgE stimulation in vitro in two patients at admission and four patients at follow-up., Conclusions: Anaphylaxis is associated with a decrease in intracellular histamine, and a reduced number and reactivity of peripheral basophils. Impaired basophil function and a decrease in their number and intracellular histamine levels in the circulation may reflect the underlying mechanism, suggesting that basophils may be a marker of anaphylaxis., (Copyright © 2019 Japanese Society of Allergology. Production and hosting by Elsevier B.V. All rights reserved.)

Basophils and mast cells have high affinity IgE receptors (FcεRI) on their plasma membrane and play important roles in FcεRI-associated allergic diseases, such as pollen allergy, food allergy, chronic spontaneous urticarial (CSU), and atopic dermatitis (AD). To date, several reports have revealed that high IgE antibody concentrations activate mast cells-which reside in tissue-in the absence of any antigens (allergens). However, IgE antibody-induced activation of basophils-which circulate in blood-has not been reported. Here, we investigated whether IgE antibodies may regulate functions of human peripheral basophils without antigens in vitro. We successfully removed IgE antibodies bound to FcεRI on the surface of human peripheral basophils by treating with 0.1% lactic acid. We also demonstrated that high IgE antibody concentrations (>1 μM) induced histamine release, polarization, and CD203c upregulation of IgE antibody-stripped basophils. Thus, high IgE antibody concentrations directly activate basophils, which express IgE-free FcεRI on the cell surface. This mechanism may contribute to the pathogenesis of patients with AD and CSU who have higher serum IgE concentrations compared to healthy donors.

Background: Atopic dermatitis (AD) is exacerbated by sweating, and the skin of most patients with AD are resided by Malassezia (M.) fungi. Recently, MGL&#95;1304 produced by Malassezia globosa was identified as the major histamine releasing antigen in human sweat., Methods: The full length cDNA of the counterpart of MGL&#95;1304 in Malassezia restricta (Mala r 8), was cloned by degenerate PCR and rapid identification of cDNA ends (RACE). Recombinant MGL&#95;1304, and its counterparts, Mala s 8 (produced by Malassezia sympodialis) and Mala r 8 were prepared, and compared in their allergenicities by dot blot analysis and histamine release tests with sera and basophils of patients with AD., Results: The identities between MGL&#95;1304 and Mala s 8, MGL&#95;1304 and Mala r 8, and Mala s 8 and Mala r 8 were 68%, 78%, and 76%, respectively, in protein sequences. Dot blot analysis revealed that the level of IgE binding to Mala s 8 was higher than that of MGL&#95;1304. However, histamine release tests revealed that MGL&#95;1304 and Mala r 8 possessed higher activity than Mala s 8. In addition, the crude lysate of M. globosa showed higher histamine release ability than that of M. sympodialis., Conclusions: Patients with AD showed hypersensitivities against MGL&#95;1304 and its homologs. However, the allergenicities of the homologs are variable and the histamine release activities may be different from the solid-phase binding activities for IgE. Sweat allergy should be carefully evaluated with biological activities of MGL&#95;1304 and its homologs of other Malassezia fungi residing on the skin., (Copyright © 2017 Japanese Society of Allergology. Production and hosting by Elsevier B.V. All rights reserved.)

MGL&#95;1304, a major allergen in human sweat for patients with atopic dermatitis and/or cholinergic urticaria, is secreted from Malassezia globosa on human skin. The amounts of MGL&#95;1304 and IgE against MGL&#95;1304 are evaluated by the histamine release test using basophils or mast cells sensitized with serum containing IgE against MGL&#95;1304, and enzyme linked sorbent assay (ELISA) using MGL&#95;1304 and anti-MGL&#95;1304 antibodies. Here, we identified a human monoclonal IgE (ABS-IgE) that binds to the high affinity IgE receptor (FcεRI) and MGL&#95;1304 with high affinity (KD = 1.99 nM) but does not release histamine from basophils and mast cells. An ELISA using ABS-IgE as a standard IgE revealed that the amount of IgE against MGL&#95;1304 (1000 U/ml) in the standard sera of patients with AD, employed in our previous report, is 32 ng/ml. A sandwich ELISA using ABS-IgE as a detection antibody showed approximately 10 times lower detection limit for MGL&#95;1304 than ELISA in which MGL&#95;1304 is directly bound to an ELISA plate. Moreover, ABS-IgE prevented histamine release from mast cells and basophils by neutralizing MGL&#95;1304 not only in a free form in solution, but also on FcεRI expressed on the cell surface without cell activation. ABS-IgE may be used both to quantify the amount of MGL&#95;1304 and anti-MGL&#95;1304 IgE, and possibly for the treatment of diseases caused/aggravated by type I allergy to MGL&#95;1304., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background: Non-steroidal anti-inflammatory drugs (NSAIDs), especially aspirin, and food additives (FAs) may exacerbate allergic symptoms in patients with chronic idiopathic urticaria and food-dependent exercise-induced anaphylaxis (FDEIA). Augmentation of histamine release from human mast cells and basophils by those substances is speculated to be the cause of exacerbated allergic symptoms. We sought to investigate the mechanism of action of aspirin on IgE-mediated histamine release., Methods: The effects of NSAIDs, FAs or cyclooxygenase (COX) inhibitors on histamine release from human basophils concentrated by gravity separation were evaluated., Results: Benzoate and tartrazine, which have no COX inhibitory activity, augmented histamine release from basophils similar to aspirin. In contrast, ibuprofen, meloxicam, FR122047 and NS-398, which have COX inhibitory activity, did not affect histamine release. These results indicate that the augmentation of histamine release by aspirin is not due to COX inhibition. It was observed that aspirin augmented histamine release from human basophils only when specifically activated by anti-IgE antibodies, but not by A23187 or formyl-methionyl-leucyl-phenylalanine. When the IgE receptor signaling pathway was activated, aspirin increased the phosphorylation of Syk. Moreover, patients with chronic urticaria and FDEIA tended to be more sensitive to aspirin as regards the augmentation of histamine release, compared with healthy controls., Conclusions: Aspirin enhanced histamine release from basophils via increased Syk kinase activation, and that the augmentation of histamine release by NSAIDs or FAs may be one possible cause of worsening symptoms in patients with chronic urticaria and FDEIA.

A technique to visualize living cell activation in a real time manner without any labeling is required in the fields of life sciences and medicine. We have reported that surface plasmon resonance (SPR) sensors detect large changes of refractive index (RI) with living cells, such as mast cells, human basophils and lymphocytes. However conventional SPR sensors detect only an average change of RI with thousands of cells at detectable area on a sensor chip. Therefore, we developed an SPR imaging (SPRI) sensor with a CMOS camera and an objective lens in order to visualize RI of individual living cells and their changes upon stimuli. The SPRI sensor we developed could detect reactions of individual rat basophilic leukemia (RBL-2H3) cells and mouse keratinocyte cells in response to specific or nonspecific stimuli. Moreover, the sensor could detect the reactions of individual human basophils isolated from patients in response to antigens (allergens). Thus the technique can visualize the effect of various stimuli, inhibitors and/or conditions on cell reactions as change of intracellular RI distribution at single cell levels. Establishment of the technique to rapidly isolate cells from patient blood should enable us to utilize SPRI system as a high throughput screening system in clinical diagnosis, such as type I hypersensitivity and drug hypersensitivity, and as a tool to reveal novel phenomena in evanescent fields around plasma membranes.

Basophil activation in response to antigen may represent specificities of type I allergy of individuals and their reactions in the body. We previously demonstrated that surface plasmon resonance (SPR) sensor could detect the activation of human basophils in response to antigens. In this study, we further developed a technique based on SPR imaging (SPRI) system to detect reactions of individual basophils isolated from human blood, and investigated the potential of this sensor as a tool for diagnosis of type I allergy. To detect the change of refractive index (RI) in individual basophils, human basophils were isolated by negative selection with antibodies conjugated with magnetic beads, fixed on a gold film with anti-basophil antibody and stimulated with various antigens under the measurement of SPRI. The sensor could detect the reactions of individual basophils in response to specific antigens as well as non-specific activators. Moreover, the sensor well allocated two spots of basophils on a sensor chip and detected individual reactions to antigen. Thus, the technique developed in this study can visualize the effect of various stimuli or inhibitors on basophils as change of intracellular RI distribution at the single cell level. In combination with a device to rapidly isolate basophils from peripheral blood, this technique may be a useful tool as a high throughput screening system in clinical diagnosis for type I allergy., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Background: The histamine releasing test which detects histamine released from basophils in vitro is safe, sensitive and widely used for clinical examination in the field of allergy. However, basophils of certain individuals do not release histamine, because of dysfunctions in their intracellular signal transduction (non-responder). To overcome potential shortcomings of the histamine releasing test, we applied surface plasmon resonance (SPR) to detect the activation of basophils., Methods: Basophils of patients with allergy, and those of non-allergic volunteers were isolated from peripheral blood. A batch of basophils obtained from a healthy volunteer was treated with lactic acid and IgE of a patient with atopic dermatitis in order to replace their endogenous IgE. They were fixed on the sensor chip of the SPR apparatus, pretreated with or without various inhibitors for intracellular signal transduction, and exposed to the antigens or anti-IgE antibody., Results: When basophils were sensitized with antigen specific IgE, they immediately caused the increase of resonance angle (AR) in response to either anti-IgE antibody or corresponding antigens, even when they did not release histamine. Moreover, the dose dependent reactions of basophils were reflected by the increase of AR as well as the release of histamine. The increase of AR in response to anti-IgE antibody was reduced by pre-treatment of basophils with inhibitors for intracellular signal transduction, but not more than the level for histamine release., Conclusions: SPR biosensors may be superior to the histamine release test for studying functions of human basophils including those not releasing histamine.

Chronic spontaneous urticaria (CSU) is a common skin disorder characterized by daily or almost daily recurring skin edema and flare with itch and pruritus anywhere on the body for more than 6 weeks. Although basophil- and mast cell-released inflammatory mediators, such as histamine, play important roles in the pathogenesis of CSU, the detailed underlying mechanism is not clear. Since several auto-antibodies, IgGs which recognize IgE or the high-affinity IgE receptor (FcεRI) and IgEs against other self-antigens, are detected in CSU patients, they are considered to activate both mast cells in the skin and basophils circulating in the blood. In addition, we and other groups demonstrated that the coagulation and complement system also contribute to the development of urticaria. Here, we summarized the behaviors, markers and targets of basophils in relation to the coagulation–complement system, and for the treatment of CSU. [ABSTRACT FROM AUTHOR]

In patients with wheat-dependent exercise-induced anaphylaxis (WDEIA), anaphylactic shock occurs frequently, therefore avoidance of wheat products is recommended. We aimed to evaluate efficacy and safety of long-term omalizumab treatment for adult patients with WDEIA. In this phase 2, multicentre single-arm trial, 20 adult patients with WDEIA were enrolled (UMIN 000019250). All patients were administered 150–600 mg of omalizumab subcutaneously and evaluations (basophil activation and blood examination) were performed at regular intervals during administration period (0–48 weeks) and observation period (48–68 weeks). Primary endpoint was proportion of the patients who achieved a basophil activation rate below 10% with fractionated wheat preparations, and secondary endpoint was proportion of the patients with no allergic reactions after wheat products ingestion. During the omalizumab treatment, more than 80% of the patients achieved the basophil activation rate less than 10% against all fractionated wheat preparations, and 68.8% of the patients who achieved the primary endpoint experienced no allergic reaction. During the observation period, the proportion of the patients who achieved a basophil activation rate below 10% decreased gradually, and the proportion of patients with positive allergic reactions increased gradually thereafter and reached maximum of 46.7%. Severe adverse events were not observed during the study. Long-term omalizumab treatment is safe and effective for adult patients with WDEIA when assessed by basophil activation rate with wheat allergens as well as allergic reactions after lifting of restrictions on wheat intake. However, this is not enough to achieve desensitization. [Display omitted] [ABSTRACT FROM AUTHOR]

We previously reported that about 80 % of patients with atopic dermatitis and 60 % with cholinergic urticaria revealed type I allergy against sweat, by means of skin test against autologous sweat and/or histamine-release test for peripheral blood basophils with semi-purified sweat antigen. In this study, we developed an assay for sera to neutralize histamine-releasing activity of semi-purified sweat antigen. The semi-purified sweat antigen was pre-incubated with serially diluted sera for 30 min at 37 °C and was subjected to histamine-release activity. Histamine release-neutralization (HRN) activities were calculated by measuring the amount of histamine release from basophils in the presence or absence of semi-purified sweat antigen. Of 62 subjects, 39 showed positive histamine release (≥5 %) from their basophils in response to semi-purified sweat antigen, and sera of 34 out of 39 subjects (87.2 %) were also positive in HRN activity (≥10 %). The specificity of the HRN assay was 0.522. Moreover, HRN activities in sera were largely correlated with degrees of histamine release from peripheral blood basophils of the same donors in response to sweat antigen. To identify the substance that neutralizes histamine-release activity, we removed IgE and IgG from the sera of HRN (+) subjects by column chromatography. The HRN activities in 30 out of 42 sera were largely reduced by the removal of IgG. On the other hand, sera of four subjects lost HRN activity by the removal of IgE, suggesting that the majority of HRN (+) subjects have serum IgG against the sweat antigen as well as IgE bound to peripheral basophils. Thus, the HRN assay maybe useful for the screening of type I allergy against sweat antigen. [ABSTRACT FROM AUTHOR]

Abstract: IL-4 plays a crucial role in the pathogenesis of allergic diseases, such as the induction of IgE synthesis and the development of mast cells. To further understand the effect of IL-4 on mast cells in skin, we utilized a mast cell/fibroblast co-culture system as an in vitro model of dermal mast cells. IL-4 induced mast cell growth in the culture with fibroblasts. Immunoblot analysis revealed that IL-4 activated Stat6 in both mast cells and fibroblasts. The over-expression of dominant-negative Stat6 in fibroblasts in the presence of IL-4 decreased the histamine content per mast cell, but not the number of mast cells. In contrast, the over-expression of constitutively-active Stat6 in fibroblasts increased the histamine content per mast cell, indicating that the activation of Stat6 in fibroblasts supports the maturation of mast cells co-cultured with fibroblasts. [Copyright &y& Elsevier]

Sweating aggravates itch in atopic dermatitis, but the mechanism is unclear. In this study, we examined the involvement of type I hypersensitivity in the aggravation of atopic dermatitis by sweating. Skin tests with autologous sweat were positive in 56 of 66 patients (84.4%) with atopic dermatitis, but only in 3 of 27 healthy volunteers (11.1%). Sweat samples from both patients and healthy volunteers induced varying degrees of histamine release from basophils of patients with atopic dermatitis. However, the histamine release was impaired by removal of IgE on the basophils. Incubation of basophils with myeloma IgE before sensitization with serum of patients blocked the ability to release histamine-induced sweat. IgE antibody against antigen(s) in sweat may be present in serum of patients with atopic dermatitis. [ABSTRACT FROM AUTHOR]

Label-free evaluation and monitoring of living cell conditions or functions by means of chemical and/or physical sensors in a real-time manner are increasingly desired in the field of basic research of cells and clinical diagnosis. In order to perform multi-parametric analysis of living cells on a chip, we here developed a surface plasmon resonance (SPR) imaging (SPRI)-impedance sensor that can detect both refractive index (RI) and impedance changes on a sensor chip with comb-shaped electrodes. We then investigated the potential of the sensor for label-free and real-time analysis of living cell reactions in response to stimuli. We cultured rat basophilic leukemia (RBL)-2H3 cells on the sensor chip, which was a glass slide coated with comb-shaped electrodes, and detected activation of RBL-2H3 cells, such as degranulation and morphological changes, in response to a dinitro-phenol-conjugated human serum albumin (DNP-HSA) antigen. Moreover, impedance analysis revealed that the changes of impedance derived from RBL-2H3 cell activation appeared in the range of 1 kHz–1 MHz. Furthermore, we monitored living cell-derived RI and impedance changes simultaneously on a sensor chip using the SPRI-impedance sensor. Thus, we developed a new technique to monitor both impedance and RI derived from living cells by using a comb-shaped electrode sensor chip. This technique may enable us to clarify complex living cell functions which affect the RI and impedance and apply this to medical applications, such as accurate clinical diagnosis of type I allergy. [ABSTRACT FROM AUTHOR]

Basophils and mast cells have high affinity IgE receptors (FcεRI) on their plasma membrane and play important roles in FcεRI-associated allergic diseases, such as pollen allergy, food allergy, chronic spontaneous urticarial (CSU), and atopic dermatitis (AD). To date, several reports have revealed that high IgE antibody concentrations activate mast cells—which reside in tissue—in the absence of any antigens (allergens). However, IgE antibody-induced activation of basophils—which circulate in blood—has not been reported. Here, we investigated whether IgE antibodies may regulate functions of human peripheral basophils without antigens in vitro. We successfully removed IgE antibodies bound to FcεRI on the surface of human peripheral basophils by treating with 0.1% lactic acid. We also demonstrated that high IgE antibody concentrations (>1 μM) induced histamine release, polarization, and CD203c upregulation of IgE antibody-stripped basophils. Thus, high IgE antibody concentrations directly activate basophils, which express IgE-free FcεRI on the cell surface. This mechanism may contribute to the pathogenesis of patients with AD and CSU who have higher serum IgE concentrations compared to healthy donors. [ABSTRACT FROM AUTHOR]

Background: Most urticarias are induced by vasoactive mediators such as histamine released from mast cells. Although mast cells are activated by allergens through cross-linking of cell-surface-bound IgE, this mechanism does not appear to explain most cases of chronic urticaria, which, when allergic, infectious, drug-induced, or physical causes cannot be identified, are classified as idiopathic. Methods: We recruited 26 patients with chronic idiopathic urticaria, in whom intradermal injection of autologous serum caused a wheal-and-flare response. Serum from four patients that induced marked histamine release from basophils from a donor with very low serum IgE levels was studied with respect to the IgE dependence of the histamine release, the activity of the IgG fractions, and the neutralizing effect of a recombinant preparation of the soluble extracellular domain of the α subunit of the high-affinity IgE receptor (sFcεRIα). Results: The histamine-releasing activity of the serum was abolished by passive sensitization of basophils with myeloma IgE, enhanced after dissociation of IgE by treatment with lactic acid, and induced by IgG fractions from the serum of all four patients. Preincubation of the serum and isolated IgG with sFcεRIα resulted in almost complete neutralization. Conclusions: Histamine-releasing IgG autoantibodies against the α subunit of the high-affinity IgE receptor are present in the circulation of some patients with chronic urticaria. Autoantibody-induced cross-linking of IgE receptors may be an important mechanism in the pathogenesis of chronic urticaria and other diseases mediated by mast cells. (N Engl J Med 1993;328:1599-604.) [ABSTRACT FROM AUTHOR]

Abstract: Surface plasmon resonance (SPR) biosensors detect large changes of angle of resonance (AR) when RBL-2H3 mast cells are cultured on a sensor chip and stimulated with antigen. However, the detail of molecular events that are responsible for such large changes of AR remained unknown. In this study, we investigated the relationship between intracellular signaling events induced by antigen and the change of AR, by genetic manipulation of intracellular signaling molecules; spleen tyrosine kinase (Syk), src-like adaptor protein (SLAP), linker for activation of T cells (LAT), growth-factor-receptor-bound protein 2 (Grb2), Grb2-related adaptor protein (Gads), and isotypes of protein kinase C (PKC). RBL-2H3 mast cells overexpressing dominant-negative Syk or SLAP, which both interfere with active Syk, exhibited only minimal increase of AR in response to antigen stimulation. Likewise, the interference of the activation of LAT and Gads, by expressing dominant-negative LAT and Gads, respectively, resulted in nearly complete suppression of the antigen-induced increase of AR. The cells overexpressing PKCs, apart from PKCβ, showed a reduced extent of increase of AR in response to antigen stimulation. Moreover, the introduction of the small interfering RNA targeted against PKCβ suppressed the antigen-induced increase of AR. These results indicate that the activation of Syk, LAT, Gads, and subsequent PKCβ is indispensable for the antigen-induced increase of AR of mast cells detected by SPR biosensors. [Copyright &y& Elsevier]