BACKGROUND: Various allergens and irritants induced the production of reactive oxygen species (ROS) in the well-established mouse dendritic cell (DC) line XS106 and this production of ROS was inhibited by antioxidants. OBJECTIVE: To investigate the production and functions of ROS in mouse bone marrow-derived DCs (BM-DCs) by various haptens and irritants, we examined the production of ROS, the expression of surface molecules, and the production of interleukin-12 (IL-12) in mouse BM-DCs. METHODS: Six to eight-week-old female C57/BL6 mice were used in this study. Mouse BM-DCs were co-cultured with DNFB, DNCB, TNBS, hydroquinone, NiSO4, CoCl2, MnCl2, thimerosal, SDS, and BKC. The production of ROS and the expression of surface molecules (CD40, CD80, CD86, and MHC-II) were measured by flow cytometry in chemical-treated mouse BM-DCs. In addition, the cells were pretreated with antioxidants to determine whether the production of ROS can be inhibited. The production of IL-12 was also measured in DNCB and SDS-treated mouse BM-DCs using ELISA. Results: The production of ROS in mouse BM-DCs was induced by various allergens, including DNFB, DNCB, TNBS, hydroquinone, MnCl2 and irritants like SDS, BKC. The expression of surface molecules was induced by various chemicals and NiSO4 was the most potent inducer of surface molecules in mouse BM-DCs. The production of ROS in DNCB and SDS-treated mouse BM-DCs was partially inhibited by diphenylene iodonium, but not by rotenone, vitamin E, allopurinol, glutathione. The production of IL-12 was not detected in DNCB and SDS-treated mouse BM-DCs. CONCLUSION: The production of ROS was induced in mouse BM-DCs by various allergens and irritants. The expression of surface molecules was also induced by various chemicals. The production of ROS was partially inhibited by DPI. The production of IL-12 was not detected.
Allergens ; Allopurinol ; Animals ; Antioxidants ; Chlorides ; Dendritic Cells ; Dinitrochlorobenzene ; Dinitrofluorobenzene ; Female ; Flow Cytometry ; Glutathione ; Haptens ; Humans ; Hydroquinones ; Interleukin-12 ; Irritants ; Manganese Compounds ; Mice ; Onium Compounds ; Reactive Oxygen Species ; Rotenone ; Thimerosal ; Vitamin E ; Vitamins

We present a bispecific antibody that recognizes an antigen and a hapten and can be applied to various biological assays, including immunoblotting and immunoprecipitation. In immunoblot analysis of serum, an anti-C5 x anti-cotinine bispecific tandem single-chain variable fragment (scFv)-Fc fusion protein and cotinine-conjugated horseradish peroxidase (HRP) generated a clean signal without the high background that was observed in a parallel experiment using HRP-conjugated goat anti-rabbit immunoglobulin G (Fc-specific) antibody. In immunoprecipitation analysis of serum, use of the bispecific tandem scFv-Fc fusion protein and cotinine-crosslinked magnetic beads significantly reduced the amount of protein contaminants compared with a parallel experiment done with protein A agarose beads. In subsequent immunoblot analysis, use of cotinine-HRP as the secondary probe instead of HRP-conjugated goat anti-rabbit IgG (Fc-specific) antibody successfully eliminated the band corresponding to the bispecific tandem scFv-Fc fusion protein.
Animals ; Antibodies, Bispecific/*immunology ; HEK293 Cells ; Haptens/*immunology ; Humans ; Immunoblotting/*methods ; Immunoprecipitation/methods ; Rabbits ; Single-Chain Antibodies/immunology

In recent years, there has emerged academic tendency towards the neurogenic mechanism of ulcerative colitis (UC). As one of the supports to the hypothesis of UC being a neurogenic inflammation, we have built a colitis model by intrathecal (ith) injection of a haptten 2,4-dinitrochlorobenzene (DNCB) to DNCB-sensitized rats. In order to explore further the neuroimmunal mechanism of this colitis model, we here focused on a pro-inflammatory cytokine, migration inhibitory factor (MIF), to observe its expression in rat colon nervous tissue and spinal cord in the colitis induced by ith injection of DNCB. At the same time we also observed the effect of MIF antibody pretreatment on the disease active index (DAI) score and the colon pathology by HE staining in the colitis rats. The results obtained showed that the immunofluorescence intensity of double staining of MIF protein in colon nervous tissue and spinal cord was increased in 0.8% and 1.6% DNCB-induced colitis groups than that in the control (60% ethanol) group. Both the colon pathology and the DAI score were significantly reduced by MIF antibody ith pretreatment. Ith injection of 0.8% DNCB after MIF antibody (1:10, 1:5) pretreatment could only induce lower DAI score (P<0.01 as compared, respectively, to the IgG pretreatment group). The colon pathological changes in ith 0.8% DNCB rats were mild, even little after MIF antibody (1:10, 1:5) pretreatment. These results suggest that MIF in spinal cord and enteric nervous system is possibly involved in the rat colitis induced by ith injection of DNCB, which reflects a neuroimmunal mechanism underlying this kind of colitis. MIF is possibly one of the important neurochemical factors in this experimental colitis, even in the UC.
Animals ; Antibodies ; pharmacology ; Colitis, Ulcerative ; chemically induced ; metabolism ; Haptens ; adverse effects ; Injections, Spinal ; Macrophage Migration-Inhibitory Factors ; metabolism ; Rats

Hapten antibodies are active components of traditional Chinese medicines, have been widely applied in all of study fields of traditional Chinese medicine. First, hapten monoclonal antibodies could be designed into ELISA kits for quantitative analysis on the content of effective components in plant crude extracts or biological samples, which be applied for quality control and studies on pharmacokenetics of traditional Chinese medicines. Second, hapten monoclonal antibodies could be coupled with solid-phase carriers to generate immunoaffinity chromatography column, which could be used for knock-out extract preparation or pre-treatment of complicated sampless. Finally, a single-chain variable fragment antibody (scFV) gene segment of effective components of hapten monoclonal antibodies could be transformed into relative plant cells to gain new varieties with high-enrichment effective components, and thus achieve the molecular breeding of medicinal plants.
Animals ; Antibodies ; genetics ; immunology ; Chromatography, Affinity ; Enzyme-Linked Immunosorbent Assay ; Gene Knockout Techniques ; Haptens ; immunology ; metabolism ; Humans ; Medicine, Chinese Traditional ; methods

Administration, Topical ; Aged ; Child ; Cyclopropanes ; administration & dosage ; adverse effects ; Female ; Haptens ; administration & dosage ; adverse effects ; Humans ; Male ; Middle Aged ; Vitiligo ; chemically induced

T-cell-mediated drug hypersensitivity represents a significant proportion of immune mediated drug hypersensitivity reactions. In the recent years, there has been an increase in understanding the immune mechanisms behind T-cell-mediated drug hypersensitivity. According to hapten mechanism, drug specific T-cell response is stimulated by drug-protein conjugate presented on major histocompatibility complex (MHC) as it is presented as a new antigenic determinant. On the other hand, p-i concept suggests that a drug can stimulate T cells via noncovalent direct interaction with T-cell receptor and/or peptide-MHC. The drug binding site is quite variable and this leads to several different mechanisms within p-i concept. Altered peptide repertoire can be regarded as an 'atypical' subset of p-i concept since the mode of the drug binding and the binding site are essentially identical to p-i concept. However, the intracellular binding of abacavir to HLA-B*57:01 additionally results in alteration in peptide repertoire. Furthermore the T-cell response to altered peptide repertoire model is only shown for abacavir and HLA-B*57:01 and therefore it may not be generalised to other drug hypersensitivity. Danger hypothesis has been postulated to play an important role in drug hypersensitivity by providing signal 2 but its experimental data is lacking at this point in time. Furthermore, the recently described allo-immune response suggests that danger signal may be unnecessary. Finally, in view of these new understanding, the classification and the definition of type B adverse drug reaction should be revised.
Binding Sites ; Classification ; Drug Hypersensitivity ; Drug-Related Side Effects and Adverse Reactions ; Hand ; Haptens ; HLA Antigens ; Major Histocompatibility Complex ; Receptors, Antigen, T-Cell ; T-Lymphocytes

Many different additives include preservatives, stabilizers, conditioners, thickeners, colorings, flavorings, sweeteners, and antioxidants. Despite the multitude of additives known, only a small number has been associated with hypersensitivity reactions. A number of investigators have suggested that a significant population of patients with allergic diseases has symptoms related to the ingestion of food additives. However, the incidence and mechanism of reactions to additives in patients with chronic urticaria, angioedema, and atopic dermatitis remain unknown. A few studies of monosodium glutamate is reported to be associated with atopic dermatitis, but their relationship remains unknown. The best known dye is tartrazine. The group of azo dyes includes ponceau and sunset yellow. Amaranth (FD&C red no. 5) was banned from use in the US in 1975 because of claims related to carcinogenicity. Most of them are reported to be associated with aggravation of atopic dermatitis. Parabens are aliphatic esters of parahydroxybenzoic acid. Sodium benzoate is a closely related substance usually reported to cross-react with these compounds. These agents, which are widely used as preservatives in both food and drugs, are well recognized as causes of severe contact dermatitis. Additives would have to act as haptens to create a response mediated by IgE. The majority of these reactions are not of the immediate hypersensitivity type. Many cases of additive-provoked urticaria or dermatitis occur as late as 24 hours after challenge, arguing against an IgE-mediated mechanism. In conclusion, the exact relationship between food additives and the allergic diseases still remains to be solved.
Angioedema ; Antioxidants ; Azo Compounds ; Coloring Agents ; Dermatitis ; Dermatitis, Atopic ; Dermatitis, Contact ; Eating ; Esters ; Food Additives ; Food Hypersensitivity ; Haptens ; Humans ; Hypersensitivity ; Hypersensitivity, Immediate ; Immunoglobulin E ; Incidence ; Parabens ; Research Personnel ; Sodium Benzoate ; Sodium Glutamate ; Sweetening Agents ; Tartrazine ; Urticaria

<p>AIMTo synthesize and identify artificial antigen of podophyllotoxin for the production of podophyllotoxin polyclonal antibody.p><p>METHODSThe hapten was synthesized by two different chemical approaches and characterized by TLC, IR, NMR, and MS. Mixed anhydride reaction (MAR) and active ester method (AEM) were used to couple the podophyllotoxin to carrier proteins (BSA and OVA). Characterization of artificial antigens was done by using spectroscopy and electrophoresis. The anti-podophyllotoxin polyclonal antibodies were obtained through immunizing rabbits.p><p>RESULTSThe results from IR, NMR and MS showed that 4-O-succinoyl podophyllotoxin (hapten) was successfully synthesized. The coupling molar ratios of the hapten and carrier proteins were 88.6 for Hapten-BSA1, 40.3 for Hapten-BSA2, 17.8 for Hapten-OVA1, and 54.2 for Hapten-OVA2. Hapten conjugates coupled with BSA yielded two sets of the specific and affinitive polyclonal antibodies. One set of antibodies showed an IC50 value of 2.21 microg.mL(-1) with a detection limit of 0.12 microg.mL(-1).p><p>CONCLUSIONAntigenic conjugates were artificially synthesized, and based on these artificial antigens, polyclonal antibodies against podophyllotoxin were raised from rabbits immunized with two different immunogens and characterized with an indirect ELISA format.p>
Animals ; Antibodies ; analysis ; Antibody Affinity ; Antibody Formation ; Antineoplastic Agents, Phytogenic ; immunology ; Enzyme-Linked Immunosorbent Assay ; Haptens ; chemistry ; immunology ; Immune Sera ; chemistry ; Male ; Ovalbumin ; immunology ; Podophyllotoxin ; immunology ; Proteins ; immunology ; Rabbits ; Serum Albumin, Bovine ; immunology

Many different additives are added to the food that we consume, and the number of additives is estimated to range from 2,000 to 20,000. These substances include preservatives, stabilizers, conditioners, thickeners, colorings, flavorings, sweeteners, and antioxidants. Despite the multitude of additives known, only a surprisingly small proportion of them has been associated with hypersensitivity reactions. A number of investigators have suggested that a significant proportion of patients with chronic urticaria, angioedema, atopic dermatitis, and asthma have symptoms related to the ingestion of food additives. However, the incidence and mechanisms of reactions to additives in patients with chronic urticaria, angioedema, and atopic dermatitis remain unknown. Monosodium glutamate(MSG) produces the flavor-enhancing properties of seaweed, the traditional component of Japanese and Chinese cooking. A few studies on MSG have reported an association between MSG and atopic dermatitis, but the exact nature of the relationship remains unknown. Dyes approved under the Food Dye and Coloring Act are coal tar derivatives, the best known of which is tartrazine(FD&C yellow no. 5). The group of azo dyes includes ponceau(FD&C red no. 4) and sunset yellow (FD&C yellow no. 6). Amaranth(FD&C red no. 5) was banned from use in US in 1975 due to the claims related to carcinogenicity. Most of them were reported to be associated with an aggravation of atopic dermatitis. Parabens are aliphatic esters of parahydroxybenzoic acid. Sodium benzoate is a closely related substance usually reported to cross-react with these compounds. These agents, which are widely used as preservatives in both foods and drugs, are well recognized as the cause of severe contact dermatitis. Additives can serve as haptens to create a response mediated by IgE. Only a few reports have suggested IgE-mediated reactions, notably to sulfites and parabens. Instead, the overwhelming majority of these reactions are not of the immediate hypersensitivity type. Many cases of additive-provoked urticaria or dermatitis occur as late as 24 hours after the challenge,arguing against an IgE-mediated mechanism.
Angioedema ; Antioxidants ; Asian Continental Ancestry Group ; Asthma ; Child ; Coal Tar ; Coloring Agents ; Cooking ; Dermatitis ; Dermatitis, Atopic* ; Dermatitis, Contact ; Eating ; Esters ; Food Additives* ; Food Hypersensitivity ; Haptens ; Humans ; Hypersensitivity ; Hypersensitivity, Immediate ; Immunoglobulin E ; Incidence ; Parabens ; Research Personnel ; Seaweed ; Sodium Benzoate ; Sodium Glutamate ; Sulfites ; Sweetening Agents ; Urticaria

To synthesize salbutamol immunogen and develop an enzyme immunoassay (ELISA), a new salbutamol immunogen was synthesized using 4-aminobenzoic acid as a linker to connect hapten with carrier protein. An enzyme immunoassay based on the antibody prepared was developed and applied to detect salbutamol residue spiked in swine liver. An unusual coating antigen, clenbuterol-ovalbumin (OVA) conjugate instead of salbutamol-OVA conjugate, was used in the immunoassay and the results were discussed based on the structures of related compounds. The antibodies showed high sensitivity in the heterologous assay when using clenbuterol-OVA as a coating antigen, with an IC50 value of 8.97 ng mL(-1) toward salbutamol. The antibodies prepared showed high cross-reactivity with clenbuterol (107%) and were promising for the simultaneous determination of salbutamol and clenbuterol residues in food and food products. Recovery rates from the salbutamol-spiked swine liver samples were in the range of 70%-99%, while the intra-assay and inter-assay coefficients of variation were <13.3% and <14.3%, respectively. In summary, the antibodies of salbutamol have been successfully prepared. Sensitive and stable analysis for the detection of salbutamol residues in swine liver was obtained based on the competitive ELISA methods developed in this study.
4-Aminobenzoic Acid ; chemistry ; Adrenergic beta-Agonists ; analysis ; immunology ; Albuterol ; analysis ; immunology ; Animals ; Antibodies ; immunology ; Antibody Specificity ; Clenbuterol ; analysis ; immunology ; Drug Residues ; analysis ; Enzyme-Linked Immunosorbent Assay ; methods ; Food Contamination ; Haptens ; immunology ; Immunization ; Liver ; chemistry ; Male ; Ovalbumin ; chemistry ; immunology ; Rabbits ; Serum Albumin, Bovine ; chemistry ; immunology ; Swine