Application of bispecific antibody against antigen and hapten for immunodetection and immunopurification

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.

Imunopatologia da dermatite de contato alérgica / Immunopathology of allergic contact dermatitis

A dermatite de contato alérgica é consequência de uma reação imune mediada por células T contra químicos de baixo peso molecular, denominados haptenos. É uma condição frequente que ocorre em todas as raças e faixas etárias e afeta a qualidade de vida de seus portadores. O mecanismo imunológico desta doença vem sendo revisto nas últimas décadas com significativo avanço no seu entendimento. A metabolização e o caminho dos haptenos, bem como a formação e o mecanismo de ação das células responsáveis tanto pela reação quanto pelo seu término, são discutidos neste artigo.

Allergic contact dermatitis is the consequence of an immune reaction mediated by T cells against low molecular weight chemicals known as haptens. It is a common condition that occurs in all races and age groups and affects the quality of life of those who present it. The immunological mechanism of this disease has been reviewed in recent decades with significant advance in its understanding. The metabolism and pathway of the haptens as well as the activation and mechanism of action of the cells responsible for both the immune reaction and its completion are discussed in this article.

Aging reduces the primary humoral response and the in vitro cytokine production in mice

Aging is accompanied by a decrease in several physiological functions that make older individuals less responsive to environmental challenges. In the present study, we analyzed the immune response of female BALB/c mice (N = 6) of different ages (from 2 to 96 weeks) and identified significant age-related alterations. Immunization with hapten-protein (trinitrophenyl-bovine serum albumin) conjugates resulted in lower antibody levels in the primary and secondary responses of old mice (72 weeks old). Moreover, young mice (2, 16, and 32 weeks old) maintained specific antibodies in their sera for longer periods after primary immunization than did old mice. However, a secondary challenge efficiently induced memory in old mice, as shown by the increased antibody levels in their sera. The number of CD4+ and CD8+ T cells in the spleen increased until 8 weeks of age but there was no change in the CD4+/CD8+ ratio with aging. Splenic T cells from old mice that had or had not been immunized were less responsive to concanavalin-A and showed reduced cytokine production compared to young mice (IL-2: 57-127 vs 367-1104 pg/mL, IFN-g: 2344-12,836 vs 752-23,106 pg/mL and IL-10: 393-2172 vs 105-2869 pg/mL in old and young mice, respectively). These data suggest that there are significant changes in the organization of the immune system throughout life. However, the relevance of these alterations for the functioning of the immune system is unknown.

Antígenos / Antigens

Los antígenos son compuestos de diversa naturaleza química provenientes del medio o generados por el propio organismo que son capaces de inducir una respuesta inmunológica en los vertebrados, propiedad denominada inmunogenicidad. La interacción del antígeno con los productos de la respuesta inmune, y especialmente con los anticuerpos, propiedad denominada antigenicidad, ha permitido conocer la estructura y función de numerosos antígenos, demostrándose que los productos de la respuesta inmune interactúan con regiones específicas del antígeno, denominadas epítopos, los cuales pueden corresponder a una secuencia aminoacídica determinada (epítopos lineales) o a un arreglo espacial de la cadena polipeptídica (epítopos conformacionales). Aunque la capacidad inmunogénica de un antígeno depende de su naturaleza química intrínseca (tamaño, forma, movilidad atómica, presencia de grupos químicos activos y residuos aromáticos) también está relacionada con la capacidad de respuesta del organismo y, en este sentido, son determinantes sus características genéticas y su historial inmunológico

Choque anafiláctico por ingestión de alcohol etílico / Anaphylactic shock after drinking an alcoholic beverage

Se comunica un caso raro de una paciente de 34 años de edad que sufrió un choque anafiláctico casi mortal 15 minutos después de ingerir una bebida alcohólica. La paciente refiere que seis años antes, con diferentes clases de bebidas alcohólicas manifestaba siempre urticaria generalizada, que desaparecía espontáneamente sin medicamantos. Aunque no se pudo demostrar el mecanismo de la reacción de nuesta paciente, la amplia variedad de bebidas alcoholicas que lo provocan sugiere que el etanol o sus metabolitos son el antígeno ofensor que actúa como hepteno

Unusual discrimination against carrier protein antibodies during partial purification of hapten-protein polyclonal antibodies to plant stress hormones.

Polyclonal antibodies (PcAb) were raised against t-zeatin riboside (t-ZR) and abscisic acid (ABA). t-ZR-BSA and ABA-BSA antibodies had high titre and specificity to haptens but also contained BSA specific antibodies as observed in double immuno diffusion and quantitative precipitation tests. Partial purification of antiserum by precipitation, desalting, and ion-exchange chromatography almost completely eliminated interference from BSA specific antibodies. Consequently, very little or no reaction was observed in dot-immunoblot assays even when high concentrations of BSA were probed with partially purified t-ZR-BSA IgG. Further studies with ABA antiserum showed that discrimination against BSA occurred during chromatography and not during salt fractionation. Because antibodies to both hapten and carrier protein were predominantly of IgG class, this unusual discrimination against carrier protein Ab was possibly influenced by two approaches followed for DEAE chromatography, viz. (i) adsorption of IgG at pH 8 followed by elution; or (ii) adsorption of contaminating proteins at neutral pH while specific IgG comes off as unbound fraction.