ABSTRACT
BACKGROUND: Total glucosides of peony (TGP), extracted from the root and rhizome of Paeonia lactiflora Pall, has well-confirmed immunomodulatory efficacy in the clinic. However, the mechanism and active ingredients remain largely unclear. HYPOTHESIS/PURPOSE: Our previous study revealed a low systemic exposure but predominant gut distribution of TGP components. The aim of this study was to investigate involvement of the gut microbiota in the immunoregulatory effects and identify the active component. METHODS: Mice received 3% DSS to establish a model of colitis. The treatment group received TGP or single paeoniflorin (PF) or albiflorin (AF). Body weight, colon length, inflammatory and histological changes were assessed. Gut microbiota structure was profiled by 16s rRNA sequencing. Antibiotic treatment and fecal transplantation were used to explore the involvement of gut microbiota. Metabolomic assay of host and microbial metabolites in colon was performed. RESULTS: TGP improved colonic injury and gut microbial dysbiosis in colitis mice, and PF was responsible for the protective effects. Fecal microbiota transfer from TGP-treated mice conferred resilience to colitis, while antibiotic treatment abrogated the protective effects. Both TGP and PF decreased colonic indole-3-lactate (ILA), a microbial tryptophan metabolite. ILA was further identified as an inhibitor of epithelial autophagy and ILA supplementation compromised the benefits of TGP. CONCLUSION: Our findings suggest that TGP acts in part through a gut microbiota-ILA-epithelial autophagy axis to alleviate colitis.
Subject(s)
Colitis/drug therapy , Colitis/microbiology , Gastrointestinal Microbiome/drug effects , Glucosides/pharmacology , Indoles/metabolism , Monoterpenes/pharmacology , Animals , Autophagy/drug effects , Bridged-Ring Compounds/pharmacology , Colitis/chemically induced , Drugs, Chinese Herbal/pharmacology , Dysbiosis/drug therapy , Dysbiosis/microbiology , Feces/microbiology , Gastrointestinal Microbiome/physiology , Glucosides/immunology , HCT116 Cells , Humans , Immunologic Factors/pharmacology , Male , Mice, Inbred BALB C , Paeonia/chemistry , RNA, Ribosomal, 16S/geneticsABSTRACT
Gastrodin (GAS) is a Chinese medicine with wide application for the treatment of nervous system disease. Previous studies reported that GAS exhibited non-specific immunomodulatory activities. To explore the effects of GAS as a vaccine adjuvant, the expression levels of CD80, CD86, MHCI and MHCII activated markers were detected after GAS treatment in vitro and in vivo, and the expression levels of IL-2 and TNF-α in splenocytes were detected after GAS treatment in vivo. Besides, the expression levels of IL-2 and IFN-γ in CD4+T cells and perforin, TNF-α and IFN-γ in CD8+T cells were detected. The effects of GAS on the survival rate and tumor size of tumor-challenged mice and the effect of cytotoxicity on CD8+T cells were also investigated. Our data showed that GAS ameliorated CD8+T cell mediated immune response and significantly improved protection of tumor-challenged animals. The results demonstrated that GAS is a potential adjuvant contributing to anticancer immunomodulation.
Subject(s)
Benzyl Alcohols/immunology , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , Cancer Vaccines/immunology , Glucosides/immunology , Melanoma/immunology , Adjuvants, Immunologic , Animals , B7-1 Antigen/metabolism , B7-2 Antigen/metabolism , Cytokines/metabolism , Female , Humans , Immunomodulation , Lymphocyte Activation , Medicine, Chinese Traditional , Mice , Mice, Inbred C57BL , Tumor BurdenABSTRACT
The involvement of innate receptors that recognize pathogen- and danger-associated molecular patterns is critical to programming an effective adaptive immune response to vaccination. The synthetic TLR4 agonist glucopyranosyl lipid adjuvant (GLA) synergizes with the squalene oil-in-water emulsion (SE) formulation to induce strong adaptive responses. Although TLR4 signaling through MyD88 and TIR domain-containing adapter inducing IFN-ß are essential for GLA-SE activity, the mechanisms underlying the synergistic activity of GLA and SE are not fully understood. In this article, we demonstrate that the inflammasome activation and the subsequent release of IL-1ß are central effectors of the action of GLA-SE, as infiltration of innate cells into the draining lymph nodes and production of IFN-γ are reduced in ASC-/- animals. Importantly, the early proliferation of Ag-specific CD4+ T cells was completely ablated after immunization in ASC-/- animals. Moreover, numbers of Ag-specific CD4+ T and B cells as well as production of IFN-γ, TNF-α, and IL-2 and Ab titers were considerably reduced in ASC-/-, NLRP3-/-, and IL-1R-/- mice compared with wild-type mice and were completely ablated in TLR4-/- animals. Also, extracellular ATP, a known trigger of the inflammasome, augments Ag-specific CD4+ T cell responses, as hydrolyzing it with apyrase diminished adaptive responses induced by GLA-SE. These data thus demonstrate that GLA-SE adjuvanticity acts through TLR4 signaling and NLRP3 inflammasome activation to promote robust Th1 and B cell responses to vaccine Ags. The findings suggest that engagement of both TLR and inflammasome activators may be a general paradigm for induction of robust CD4 T cell immunity with combination adjuvants such as GLA-SE.
Subject(s)
Adjuvants, Immunologic/pharmacology , Antigens/immunology , B-Lymphocytes/immunology , Inflammasomes/immunology , Th1 Cells/immunology , Toll-Like Receptor 4/immunology , Vaccines/immunology , Adaptor Proteins, Vesicular Transport/metabolism , Adenosine Triphosphate/metabolism , Animals , CARD Signaling Adaptor Proteins/genetics , Female , Glucosides/immunology , Immunity, Humoral , Interferon-beta/immunology , Interferon-gamma/immunology , Interleukin-1beta/metabolism , Interleukin-2/immunology , Lipid A/immunology , Mice , Mice, Inbred C57BL , Mice, Knockout , Myeloid Differentiation Factor 88/metabolism , NLR Family, Pyrin Domain-Containing 3 Protein/genetics , Receptors, Interleukin-1 Type I/genetics , Squalene/immunology , Toll-Like Receptor 4/agonists , Toll-Like Receptor 4/genetics , Tumor Necrosis Factor-alpha/immunology , VaccinationABSTRACT
Immunoassay systems using monoclonal antibodies (mAbs) are one of the most useful techniques in the analytical, biochemical, and clinical fields. In this study, a combination enzyme-linked immunosorbent assay (ELISA) using both anti-glycyrrhizin and anti-liquiritin mAbs (anti-GL/Liq mixture mAbs) was developed for quality control of licorice and its products. The combination ELISA demonstrated high sensitivity, reproducibility, and specificity for the total content of GL and Liq by a single assay. The developed ELISA was effective and useful as the first screening method in the selection of high-quality licorice from the Glycyrrhiza species and in confirming the quality of licorice-containing Kampo medicines.
Subject(s)
Antibodies, Monoclonal/immunology , Enzyme-Linked Immunosorbent Assay , Flavanones/analysis , Glucosides/analysis , Glycyrrhiza/chemistry , Glycyrrhizic Acid/analysis , Medicine, Kampo , Plant Roots/chemistry , Antigen-Antibody Reactions , Flavanones/immunology , Glucosides/immunology , Glycyrrhizic Acid/immunologyABSTRACT
Flos Lonicerae Japonicae (FLJ), the flower bud of Lonicera japonica Thunb. (Caprifoliaceae), is a widely used traditional Chinese medicine with various pharmacological activities. Luteoloside is a major active compound and a quality control marker of FLJ. Luteolin-7-O-glucuronide (LG), an analog of luteoloside, was conjugated with bovine serum albumin (BSA) and ovalbumin (OVA) to create the immunogen and coating antigen, respectively. A sensitive and specific monoclonal antibody (mAb), designated as mAb3A4, was generated with LG-BSA. To screen the authenticity and quality of FLJ, an indirect competitive enzyme-linked immunosorbent assay (icELISA) was established. The concentration of luteoloside producing 50 % inhibition and the working range of the icELISA were 42.3 and 9.1-258.1 µg L(-1), respectively. The icELISA showed cross-reactivity values of 2414, 402, 230, and <1 % for LG, baicalin, scutellarin, and other analogs of luteoloside, respectively. The average recovery of luteoloside in the FLJ samples as determined by icELISA ranged from 83.0 to 112.5 %. The luteoloside content was determined for different Lonicera herbal samples with icELISA, and the results were confirmed by high-performance liquid chromatography analysis. Thus, this icELISA is suitable for the quality assurance of FLJ samples. Graphical abstract Specific monoclonal antibody-based enzyme-linked immunosorbent assay for luteoloside.
Subject(s)
Antibodies, Monoclonal/immunology , Antibody Formation , Enzyme-Linked Immunosorbent Assay/methods , Glucosides/analysis , Glucosides/immunology , Lonicera/chemistry , Luteolin/analysis , Luteolin/immunology , Plant Extracts/chemistry , Animals , Antibodies, Monoclonal/chemistry , Cell Line , Female , Haptens/chemistry , Haptens/immunology , Lonicera/immunology , Luteolin/chemistry , Mice, Inbred BALB C , Plant Extracts/immunologyABSTRACT
In this study, we developed a fluorescent immunoassay approach to detect paeoniflorin (PF) using a fluorescently labelled monoclonal antibody. The PF-specific antibody was purified by the caprylic acid-ammonium sulfate method and protein G Sepharose 4 Fast Flow column and then labelled with fluorescein isothiocyanate (FITC). The FITC-labelled monoclonal antibody was highly specific for PF, with less than 0.076 % cross-reactivity to seven structurally related compounds. The FITC-labelled monoclonal antibody was then used to develop an indirect competitive enzyme-linked immunosorbent assay (icELISA) and indirect competitive fluorescence-linked immunosorbent assay (icFLISA), respectively. FLISA is simple, rapid and sensitive, with a 500-fold lower limit of detection (LOD) compared with conventional ELISA. Finally, using a variety of standards, FLISA was validated. We observed a strong correlation between the results determined by either FLISA or conventional HPLC for the quantification of PF levels (R(2) = 0.9927). Collectively, this study shows that the icFLISA method can be successfully applied for the detection and quantification of PF in medicines and biological samples.
Subject(s)
Antibodies, Monoclonal/immunology , Drugs, Chinese Herbal/analysis , Enzyme-Linked Immunosorbent Assay/methods , Glucosides/analysis , Immunosorbents/chemistry , Monoterpenes/analysis , Fluorescence , Fluorescent Antibody Technique , Glucosides/immunology , Limit of Detection , Monoterpenes/immunologyABSTRACT
This study aimed to develop an indirect competitive enzyme-linked immunosorbent assay based on monoclonal antibodies against paeoniflorin to study the effects of different doses of glycyrrhizinic acid on the pharmacokinetics of paeoniflorin in mice. An anti-paeoniflorin monoclonal antibody was produced from a hybridoma created through a fusion of splenocytes immunized with paeoniflorin-bovine serum albumin and conjugated with the hypoxanthine-aminopterin-thymidine-sensitive mouse myeloma cell line SP2/0. The resultant antibody was used to develop and validate a rapid, specific and sensitive, indirect competitive enzyme-linked immunosorbent assay for the measurement of paeoniflorin (linear range 4.8-312.5 ngâ·âmL(-1)). The intraday and interday precision values of the indirect competitive ELISA method were well within the recommended range (≤ 10â%), and the recovery rate was, on average, 101.13â%. Pharmacokinetic parameters obtained from mouse blood samples at various intervals following the oral administration of paeoniflorin and glycyrrhizic acid at three doses (1â:â0.3, 1â:â1, 1â:â3, respectively) demonstrated that the highest dose of glycyrrhizic acid inhibits the absorption of paeoniflorin.
Subject(s)
Enzyme-Linked Immunosorbent Assay/methods , Glucosides , Glycyrrhizic Acid/pharmacokinetics , Monoterpenes , Animals , Antibodies, Monoclonal , Drug Interactions , Female , Glucosides/immunology , Glucosides/isolation & purification , Glucosides/pharmacokinetics , Hybridomas/immunology , Mice , Mice, Inbred BALB C , Monoterpenes/immunology , Monoterpenes/isolation & purification , Monoterpenes/pharmacokineticsABSTRACT
Oxidation method with sodium iodide was used to synthesize immunogenic antigen (PF-BSA) and coating antigen (PF-OVA) of paeoniflorin. UV spectroscopy showed that paeoniflorin was successfully conjugated with BSA and OVA. After immunized by PF-BSA, the mice can produce anti-paeoniflorin antibodies specifically. The ELISA test results showed the high titer (1:12 800) and specificity (IC50 = 0.791 mg x L(-1)) of the antiserum from mice injected with PF-BSA. Also, the antiserum showed low cross activities against nine traditional Chinese medicine (TCM) of small molecules. These artificial antigens were successfully synthesized and the anti-paeoniflorin antibody well prepared, which provides the experimental basis for the further study of ELISA and its kit.
Subject(s)
Antigens/chemistry , Drugs, Chinese Herbal/chemistry , Glucosides/chemistry , Monoterpenes/chemistry , Animals , Antibodies/analysis , Antigens/immunology , Enzyme-Linked Immunosorbent Assay , Glucosides/immunology , Male , Mice , Mice, Inbred BALB C , Monoterpenes/immunology , Serum Albumin, Bovine/chemistry , Serum Albumin, Bovine/immunologyABSTRACT
Arabinogalactan-proteins are glycoproteins that occur in higher plants and are involved in important processes like cell differentiation and plant growth. In the medicinal plant Echinacea purpurea L., they belong to the putative immunomodulating compounds and are structurally well characterized. For microscopic localization of arabinogalactan-proteins, synthetic (ß-D-Glc)3 Yariv phenylglycoside that specifically binds to most plant arabinogalactan-proteins was used to label arabinogalactan-proteins in fresh cut sections of stems and petioles of Echinacea purpurea. Polyclonal antibodies against (ß-D-Glc)3 Yariv phenylglycoside were used to detect the arabinogalactan-protein-(ß-D-Glc)3 Yariv phenylglycoside complex. After addition of fluorescein isothiocyanate-conjugated secondary antibodies, the sections were analyzed by confocal laser scanning microscopy. Arabinogalactan-proteins are localized mainly in the central cylinder in the collateral vascular bundles, especially in the area of the xylem. In cell walls of fully differentiated vessels and tracheids, arabinogalactan-proteins have been detected mainly at the inner area of the wall close to the cell lumina. Intense labeling occurs around pit canals connecting adjacent vessels. Furthermore, arabinogalactan-proteins are present in the lumina of cells of the sclerenchyma caps and in companion cells of the phloem.
Subject(s)
Antibodies , Echinacea/chemistry , Glucosides/immunology , Mucoproteins/immunology , Phloroglucinol/analogs & derivatives , Antibodies/immunology , Cell Wall/chemistry , Cell Wall/metabolism , Cell Wall/ultrastructure , Echinacea/metabolism , Echinacea/ultrastructure , Glycoproteins/immunology , Glycoproteins/metabolism , Indicators and Reagents , Microscopy, Confocal , Mucoproteins/metabolism , Phloroglucinol/immunology , Plant Proteins/immunology , Plant Proteins/metabolism , Plant Stems/chemistry , Plant Stems/metabolism , Plant Stems/ultrastructure , Plant Vascular Bundle/chemistry , Plant Vascular Bundle/metabolism , Plant Vascular Bundle/ultrastructure , Plants, Medicinal , Sensitivity and Specificity , Staining and LabelingABSTRACT
INTRODUCTION: Paeoniae radix is one of the most important crude drugs used in Kampo medicines (KMs). A part of its pharmaceutical properties is due to the presence of paeoniflorin (PF) and albiflorin (AF). OBJECTIVE: For the specific and easy identification of PF and AF, an immunostaining technique was developed using anti-PF monoclonal antibody (MAb). METHODOLOGY: PF and AF were treated with a NaIO4 solution and reacted with bovine serum albumin (BSA) preparing PF- and AF-BSA conjugates on the polyethersulphone (PES) membrane. Anti-PF MAb was bound and then antibody labelled with peroxidase directed against anti-PF MAb. Finally, a substrate was added and then PF and AF were detected. RESULTS: Anti-PF MAb recognised not only PF but also AF when 10 µg was present on the PES membrane. As little as 0.5 µg of PF and AF were still detected under immunostaining. Various Paeoniae radix samples and KMs were qualitatively analysed, and total amounts of PF and AF were visually detected by immunostaining technique. This method was applied to investigate the distribution of PF and AF in fresh peony root using immunoblotting by transferred from peony root to the PES membrane. CONCLUSION: The technique permitted the visualisation of PF and AF on PES membrane using immunostaining. The immunostaining technique established would be a powerful tool for probing the sources of PF and AF in plant extracts.
Subject(s)
Benzoates/analysis , Bridged-Ring Compounds/analysis , Glucosides/analysis , Immunoblotting/methods , Animals , Antibodies, Monoclonal/immunology , Benzoates/chemistry , Benzoates/immunology , Bridged-Ring Compounds/chemistry , Bridged-Ring Compounds/immunology , Cattle , Glucosides/chemistry , Glucosides/immunology , Membranes, Artificial , Molecular Structure , Monoterpenes , Paeonia/chemistry , Perchlorates , Plant Extracts/analysis , Plant Extracts/chemistry , Polymers/chemistry , Reproducibility of Results , Serum Albumin, Bovine/chemistry , Sodium Compounds , Staining and Labeling/methods , Sulfones/chemistryABSTRACT
The seed of Plantago asiatica L. is one of the most popular folk herbal medicines used in China and other Asian countries. In this study, phenylethanoid glycosides and polysaccharides were isolated from the seed of P.asiatica L. by using phytochemical investigation methods. A screening model of immunological activity by using dendritic cells as target cells was established to investigate the effects of these compounds on the phenotypic and functional maturation of dendritic cells. Compared with untreated cells, dendritic cells treated with acteoside, isoacteoside or polysaccharides expressed higher level of class II MHC and costimulatory molecule CD86 (B7-2). Functional maturation was confirmed by decreased endocytosis and increased naïve T cell stimulatory activity of dendritic cells. These results showed that acteoside, isoacteoside and polysaccharides from the seed of P.asiatica L. had significant immunoenhancing activity by inducing the maturation of dendritic cells.
Subject(s)
Bone Marrow , Dendritic Cells/cytology , Glucosides/immunology , Phenols/immunology , Plantago/chemistry , Polysaccharides/immunology , Seeds/chemistry , Animals , Cell Proliferation , Dendritic Cells/immunology , Endocytosis , Female , Glucosides/isolation & purification , Lectins, C-Type/metabolism , Mannose Receptor , Mannose-Binding Lectins/metabolism , Mice , Phenols/isolation & purification , Phenotype , Polysaccharides/isolation & purification , Receptors, Cell Surface/metabolism , T-Lymphocytes/cytology , T-Lymphocytes/immunologyABSTRACT
A recombinant single chain variable-fragment (scFv) antibody against paeoniflorin (PF) was produced using the hybridoma cell line C31B9. Variable regions of heavy (V (H)) and light (V (L)) chain antibody genes were directly cloned from cDNA resources of hybridoma C31B9 and assembled using splicing by overlap extension (SOE)-PCR using a (Gly (4)Ser) (3) linker DNA. The constructed scFv genes were cloned into pET28a vectors for the generation of recombinant proteins in Escherichia coli. Most of the recombinant proteins were expressed in inclusion bodies. The yield of refolded and purified scFv was 1.89 mg per 100 mL of cell culture. The recombinant scFv displayed cross-reactivity as its mother monoclonal antibody (MAb) C31B9. Therefore, the newly expressed scFv protein was applied to quantitative ELISA to determine the total paeoniflorin (PF) and albiflorin (Alb) concentrations in peony root samples. Using PF as a standard compound, the full linear range of the assay was extended from 0.78 to 25 microg/mL. The results obtained by ELISA employing both the recombinant scFv and the original MAbC31B9 showed a reasonably good agreement with each other.
Subject(s)
Antibodies, Monoclonal/immunology , Benzoates/immunology , Bridged-Ring Compounds/immunology , Enzyme-Linked Immunosorbent Assay/methods , Glucosides/immunology , Immunoglobulin Fragments/immunology , Immunoglobulin Variable Region/immunology , Antibodies, Monoclonal/biosynthesis , Antibody Affinity , Benzoates/analysis , Benzoates/chemistry , Bridged-Ring Compounds/analysis , Bridged-Ring Compounds/chemistry , Cell Line , Cross Reactions , Drugs, Chinese Herbal/chemistry , Escherichia coli/metabolism , Glucosides/analysis , Glucosides/chemistry , Immunoglobulin Fragments/biosynthesis , Immunoglobulin Fragments/genetics , Immunoglobulin Variable Region/genetics , Monoterpenes , Paeonia/chemistry , Recombinant Proteins/biosynthesis , Recombinant Proteins/immunologyABSTRACT
This work describes an immunochemical approach for the quality control of Paeoniae Radix by an enzyme-linked immunosorbent assay (ELISA) based on determination of the total concentration of paeoniflorin (PF) and albiflorin (Alb), which are major bioactive constituents in Paeoniae Radix. Four hybridromas secreting monoclonal antibodies against PF and Alb were produced by fusing splenocytes from a mouse immunized by a PF-bovine serum albumin (BSA) conjugate with the hypoxanthine-aminopterin-thymidine (HAT)-sensitive mouse myeloma cell line, P3-X63-Ag8-653. A relatively higher reactivity of monoclonal antibodies (MAbs) with PF and Alb than oxypaeoniflorin (OP) and benzoylpaeoniflorin (BP) was observed, while other monoterpenes and benzoic acid did not cross-react. When PF was used as a standard, the assay can cover a measuring range of 20-600 ng/ml for PF and Alb. A series of Paeoniae Radix samples have been determined, and the results showed good agreement with that determined by traditional high-performance liquid chromatography (HPLC). The developed competitive ELISA was 100 times more sensitive than the HPLC method. Meanwhile, fifteen Chinese traditional prescriptions were determined by the competitive ELISA.
Subject(s)
Antibodies, Monoclonal/immunology , Benzoates/analysis , Bridged-Ring Compounds/analysis , Glucosides/analysis , Medicine, Chinese Traditional/standards , Animals , Antibodies, Monoclonal/biosynthesis , Benzoates/immunology , Bridged-Ring Compounds/immunology , Cell Fusion , Chromatography, High Pressure Liquid , Cross Reactions , Enzyme-Linked Immunosorbent Assay/methods , Glucosides/immunology , Hybridomas/immunology , Mice , Monoterpenes , Tumor Cells, CulturedABSTRACT
Syringin was found to possess immunomodulatory activity by which it inhibited the in-vitro immunohaemolysis of antibody-coated sheep erythrocytes by guinea-pig serum through suppression of C3-convertase of the classical complement. In this study, we examined its in-vitro and in-vivo activity on tumour necrosis factor (TNF)-alpha and nitric oxide (NO) production, CD4+ T cell and CD8+ cytotoxic T cell (CTLL-2) proliferation, and croton oil-, arachidonic acid- and fluorescein-isothiocynate (FITC)-induced mouse ear oedema model. Syringin significantly inhibited both TNF-alpha production from lipopolysaccharide (LPS)-stimulated RAW264.7 cells and CD8+ T cell (CTLL-2) proliferation in a dose-dependent manner, whereas neither NO production nor CD4+ T cell proliferation were blocked even by high concentrations of syringin. In the invivo experiments, syringin also significantly suppressed FITC-induced ear oedema in mice but not the ear oedema induced by croton or arachidonic acid. These results suggest that syringin may be implicated as an immunomodulator having an anti-allergic effect rather than an anti-inflammatory effect. The anti-allergic effect of syringin seems to be due, in part, to inhibition of TNF-alpha production and cytotoxic T cell proliferation.
Subject(s)
Adjuvants, Immunologic/pharmacology , Glucosides/immunology , Glucosides/pharmacology , Phenylpropionates/immunology , Phenylpropionates/pharmacology , T-Lymphocytes, Cytotoxic/drug effects , Tumor Necrosis Factor-alpha/drug effects , Animals , Cell Culture Techniques , Cell Division/drug effects , Guinea Pigs , Immune System/drug effects , Male , Mice , Mice, Inbred BALB C , Mice, Inbred ICR , Sheep , Tumor Necrosis Factor-alpha/biosynthesisABSTRACT
Cellular immune responses are initiated when T lymphocytes expressing alphabeta TCR recognize peptide antigens bound to MHC molecules or, less frequently, double-stranded glycolipid antigens bound to CD1 molecules. In the allergy to Parietaria judaica, human alphabeta CD8+ Th2 lymphocytes react to a non-peptide pollinic antigen presented by B cells. The environmental allergen was purified and identified as a new flavonoid pigment: 2'-O-sulfate, 6-O-betaD-glucuronopyranosyl, 2',5,6-trihydroxy-isoflavone. Its specific recognition by alphabeta CD8+ Th2 T cells (1) depends upon an MHC- and CD1-independent presentation mediated by B cells, (2) is determined by the flavonoid carbohydrate and sulfate groups and (3) leads to positive skin prick test in allergic patients. Hence, an unusual mode of aromatic sulfated antigen recognition by alphabeta CD8+ Th2 T lymphocytes might underlie the cellular mediation of human allergy to plant allergens.