Anti-idiotypic monoclonal antibody AB3, reacting with the primary antigen (CEA), can localize in human colon-carcinoma xenografts as efficiently as AB1.

BALB/c mice were immunized with anti-idiotypic monoclonal (MAb) antibody (anti-Id or Ab2) directed against an AB1 MAb anti-carcinoembryonic (CEA) in order to obtain AB3 MAbs (anti-anti-Id). AB3 MAbs were shown to recognise the primary antigen (CEA) and one of them was tested extensively in vitro and in vivo. This AB3 MAb was shown to bind specifically to CEA on frozen sections of a human colon carcinoma by immunoperoxidase. Scatchard plot analyses showed that the affinity of this AB3 was of the same order of magnitude as the AB1. In vivo experiments, in nude mice bearing CEA-producing human colon-carcinoma xenografts showed that up to 30% of the intravenously injected dose of 125I-labelled AB3 were localized per gram of tumour tissue. Furthermore, calculation of the ratios of AB3 concentration in the tumour over those in normal organs such as lung, liver, kidney, spleen and bone gave relatively high values similar to results obtained with AB1. All together our results show that AB3 can localize as efficiently and specifically in the tumour as AB1, despite the fact that the mice from which it was derived were immunized with a mouse MAb (AB2) and had never been exposed to CEA.

A 24-month randomized clinical trial of a two- and three-step etch-and-rinse technique

Purpose This 24-month randomized paired tooth clinical study evaluated the performance of All Bond 3 used in the simplified (2-step) and full (3-step) versions Methods 33 patients, with at least two similar sized non-carious cervical lesions participated in this study A total of 66 restorations were placed, half using the 2-step All Bond 3 (AB3-2) and the other half using 3-step All Bond 3 (AB3-3) The restorations were placed incrementally using the composite resin Aelite The restorations were evaluated at baseline and after 6, 12 and 24 months following the modified USPHS criteria Statistical differences between the adhesive were tested using with McNemar`s test and clinical performance over time for each material with the Fisher`s exact test (alpha= 0 05) Results After 24 months, six AB3-2 and four AB3-3 were rated as bravo for marginal discoloration but did not differ from each other significantly (P> 0 05) The retention rates at 24 months of AB3-2 and AB3-3 were 90 9% and 97 0%, respectively (P> 0 05) (Am J Dent 2010,23 231-236)

Induction of an immune response through the idiotypic network with monoclonal anti-idiotype antibodies in the carcinoembryonic antigen system.

Anti-idiotype antibodies can mimic the conformational epitopes of the original antigen and act as antigen substitutes for vaccination and/or serological purposes. To investigate this possibility concerning the tumor marker carcinoembryonic antigen (CEA), BALB/c mice were immunized with the previously described anti-CEA monoclonal antibody (MAb) 5.D11 (AB1). After cell fusion, 15 stable cloned cell lines secreting anti-Ids (AB2) were obtained. Selected MAbs gave various degrees of inhibition (up to 100%) of the binding of 125I-labeled CEA to MAb 5.D11. Absence of reactivity of anti-Id MAbs with normal mouse IgG was first demonstrated by the fact that anti-Id MAbs were not absorbed by passage through a mouse IgG column, and second because they bound specifically to non-reduced MAb 5.D11 on Western blots. Anti-5.D11 MAbs did not inhibit binding to CEA of MAb 10.B9, another anti-CEA antibody obtained in the same fusion as 5.D11, or that of several anti-CEA MAbs reported in an international workshop, with the exception of two other anti-CEA MAbs, both directed against the GOLD IV epitope. When applied to an Id-anti-Id competitive radioimmunoassay, a sensitivity of 2 ng/ml of CEA was obtained, which is sufficient for monitoring circulating CEA in carcinoma patients. To verify that the anti-Id MAbs have the potential to be used as CEA vaccines, syngeneic BALB/c mice were immunized with these MAbs (AB2). Sera from immunized mice were demonstrated to contain AB3 antibodies recognizing the original antigen, CEA, both in enzyme immunoassay and by immunoperoxidase staining of human colon carcinoma. These results open the perspective of vaccination against colorectal carcinoma through the use of anti-idiotype antibodies as antigen substitutes.

Cytotoxic study of organometallic compounds of palladium (II) in mice peritoneal macrophages

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Caracterização de anticorpos anti-idiotípicos e anti-anti-idiotípicos de BEVACIZUMAB

Propomos avaliar o potencial de uma abordagem imunoterápica que tem o fator de crescimento do endotélio vascular (VEGF) como alvo. A angiogênese é necessária para nutrir o tumor e propicia sua metastatização. Sabe-se que VEGF é produzido pela maioria dos tumores sólidos metastatizantes. A eficiência do anticorpo anti-VEGF, BEVACIZUMAB, já foi demonstrada. Entretanto, efeitos adversos limitam sua utilização em alguns casos e, com base na teoria da rede idiotípica e na experiência do grupo com anticorpos anti-idiotípicos (anti-Id ou AB2), anticorpos AB2 capazes de mimetizar VEGF foram obtidos. Assim, com o objetivo de explorar o rationale, dois anticorpos monoclonais (MAb) anti-Id de Bevacizumab, 10.D7 e 3.E3, foram produzidos em nosso laboratório. O controle da purificação de anticorpos foi feito através da avaliação do material purificado em géis de poliacrilamida. Immunoblots comprovaram a especificidade do reconhecimento do idiotipo de Bevacizumab pelos MAbs anti-Id. Foi mostrado que os AB2 obtidos se ligam a moléculas da superfície de células endoteliais, que expressam receptores de VEGF, sugerindo que esses anticorpos mimetizam VEGF. Entretanto, não foi possível verificar inibição do crescimento dessas células quando cultivadas em presença desses MAbs. Será necessária a otimização dos protocolos utilizados para uma avaliação mais definitiva. Uma vez demonstrada a atividade anti-idiotípica dos AB2 in vitro, 10.D7 e 3.E3 foram utilizados como imunógenos em camundongos Balb/c, com o objetivo de avaliar a resposta AB3 ou anti-anti-Id. Por meio de ensaio imunoenzimático (ELISA) indireto foi verificada a presença de anticorpos anti-anti-Id ligantes de VEGF nos soros dos camundongos imunizados. Os animais imunizados com o MAb 10.D7 responderam melhor do que os imunizados com 3.E3. Anticorpos IgG purificados dos soros desses animais reconheceram VEGF em ensaios imunoenzimáticos. Foram então selecionados os ...

Characterization of anti-anti-idiotypic antibodies that bind antigen and an anti-idiotype

Two mouse monoclonal anti-anti-idiotopic antibodies (anti-anti-Id, Ab3), AF14 and AF52, were prepared by immunizing BALB/c mice with rabbit polyclonal anti-idiotypic antibodies (anti-Id, Ab2) raised against antibody D1.3 (Ab1) specific for the antigen hen egg lysozyme. AF14 and AF52 react with an “internal image” monoclonal mouse anti-Id antibody E5.2 (Ab2), previously raised against D1.3, with affinity constants (1.0 × 109 M−1 and 2.4 × 107 M−1, respectively) usually observed in secondary responses against protein antigens. They also react with the antigen but with lower affinity (1.8 × 106 M−1 and 3.8 × 106 M−1). This pattern of affinities for the anti-Id and for the antigen also was displayed by the sera of the immunized mice. The amino acid sequences of AF14 and AF52 are very close to that of D1.3. In particular, the amino acid side chains that contribute to contacts with both antigen and anti-Id are largely conserved in AF14 and AF52 compared with D1.3. Therapeutic immunizations against different pathogenic antigens using anti-Id antibodies have been proposed. Our experiments show that a response to an anti-Id immunogen elicits anti-anti-Id antibodies that are optimized for binding the anti-Id antibodies rather than the antigen.

Inositol 1,4,5-trisphosphate receptor subtype 3 in pancreatic islet cell secretory granules revisited.

It has been reported that the inositol 1,4,5-trisphosphate receptor subtype 3 is expressed in islet cells and is localized to both insulin and somatostatin granules [Blondel, O., Moody, M. M., Depaoli, A. M., Sharp, A. H., Ross, C. A., Swift, H. & Bell, G. I. (1994) Proc. Natl. Acad. Sci. USA 91, 7777-7781]. This subcellular localization was based on electron microscope immunocytochemistry using antibodies (affinity-purified polyclonal antiserum AB3) directed to a 15-residue peptide of rat inositol trisphosphate receptor subtype 3. We now show that these antibodies cross-react with rat, but not human, insulin. Accordingly, the anti-inositol trisphosphate receptor subtype 3 (AB3) antibodies label electron dense cores of mature (insulin-rich) granules of rat pancreatic beta cells, and rat granule labeling was blocked by preabsorption of the AB3 antibodies with rat insulin. The immunostaining of immature, Golgi-associated proinsulin-rich granules with AB3 antibodies was very weak, indicating that cross-reactivity is limited to the hormone and not its precursor. Also, the AB3 antibodies labeled pure rat insulin crystals grown in vitro but failed to stain crystals grown from pure human insulin. By immunoprecipitation, the antibodies similarly displayed a higher affinity for rat than for human insulin. We could not confirm the labeling of somatostatin granules using AB3 antibodies.

Long-term biogenic particle fluxes in the Bering Sea and the central subarctic Pacific Ocean

Time-series sediment traps were deployed for five consecutive years in two distinctively different subarctic marine environments. The centrally located subarctic pelagic Station SA (49°N, 174°W; water depth 5406 m) was simultaneously studied along with the marginal sea Station AB (53.5°N, 177°W; water depth 3788 m) in the Aleutian Basin of the Bering Sea. A mooring system was tethered to the sea-floor with a PARFLUX type trap with 13 sample bottles, which was placed at 600 m above the sea-floor at each of the two stations. Sampling intervals were synchronized at the stations, and they were generally set for 20 days during highly productive seasons, spring through fall, and 56 days during winter months of low productivity. Total mass fluxes, which consisted of mainly biogenic phases, were significantly greater at the marginal sea Station AB than at the pelagic Station SA for the first four years and moderately greater for the last year of the observations. This reflects the generally recognized higher productivity in the Bering Sea. Temporal excursion patterns of the mass fluxes at the two stations generally were in parallel, implying that temporal changes in their biological productivity are strongly governed by a large-scale seasonal climatic variability over the region rather than local phenomena. The primary reason for the difference in total mass flux at the two stations stems mainly from varying contributions of siliceous and calcareous planktonic assemblages. A significantly higher opal contribution at Station AB than at Station SA was mainly due to diatoms. Diatom fluxes at the marginal sea station were about twice those observed at the pelagic station, resulting in a very high opal contribution at Station AB. In contrast to the opal fluxes, CaCO3 fluxes at Station AB were slightly lower than at Station SA. The ratios of Corg/Cinorg were usually significantly greater than one in both regions, suggesting that preferentially greater organic carbon from cytoplasm than skeletal inorganic carbon was exported from the surface layers. Such a process, known as the biological pump, leads to a carbon sink which effectively lowers p CO2 in the surface layers and then allows a net flux of atmospheric CO2 into the surface layer. The efficiency of the biological pump is greater in the Bering Sea than at the open-ocean station.