Epitopos B de plasmodium vivax: un estudio de pacientes infectados en el área endémica venezolana / Epitopes B of

Hemos investigado la respuesta de anticuerpos a la proteína que circunda el esporozoíto (CSP), y a la principal de la superficie de los merozoítos de Plasmodium vivax (MSP-1Pv), en individuos infectados naturalmente en el área malárica meridional de Venezuela. Peptidos sintéticos que representan la subunidad repetida de las variantes VK210 y VK247 de la CSP o fragmentos de la región N y C-terminal de la MSP-1Pv,

How the anti-(metal chelate) antibody CHA255 is specific for the metal ion of its antigen: X-ray structures for two Fab'/hapten complexes with different metals in the chelate.

Antibodies with bound metal-chelate haptens provide new means for exploiting the diverse properties of metallic elements. The murine monoclonal antibody CHA255 (IgG1 lambda) binds the metal-chelate hapten indium (III)-4-[N'-(2-hydroxyethyl)thioureido]-L-benzyl-EDTA (designated In-EOTUBE) with high affinity (K(a) = 1.1 x 10(10) M-1). Antibody binding is highly specific for the indium chelate; the affinity decreases as much as 10(4) with other metals, even those having ionic radii close to indium. To better understand this selectivity, the crystal structure of the antigen-binding fragment (Fab') of CHA255 complexed with its hapten, In(III)-EOTUBE, was determined by molecular replacement and refined at 2.2-A resolution. The structure of CHA255 Fab' complexed with Fe(III)-EOTUBE was also determined and refined at 2.8-A resolution. In both structures, the hapten's EDTA moiety is half-buried near the center of the complementarity-determining regions (CDR's). Five of the six CDR's on the Fab' interact with the hapten through protein side-chain atoms (but not main-chain atoms). A novel feature of the In-EOTUBE/Fab' complex is coordination of the indium by N epsilon of one histidine from the heavy chain's third CDR (distance = 2.4 A). The histidine coordination is not observed in the Fe-EOTUBE/Fab' complex, due mainly to a slightly different hapten conformation that reduces metal accessibility; this may partially explain the 20-fold lower affinity of CHA255 for iron hapten. An unexpected feature of the Fab' overall is an elbow angle of 193 degrees (the angle between the pseudodyad axes of the Fab's constant and variable domains).

Increased antibody expression from Escherichia coli through wobble-base library mutagenesis by enzymatic inverse PCR.

We tested the value of a new library mutagenesis approach, called library enzymatic inverse PCR (LEIPCR), for expression-level enhancement of antibody Fv fragments produced in Escherichia coli. The production level of active, metal chelate-specific antibody from our constructs is limited by a low expression level of the second, heavy-chain cistron. To increase the production level, LEIPCR was applied to the wobble bases of the second cistron leader peptide. In LEIPCR mutagenesis, the entire plasmid is amplified using mutagenic primers with class-IIS restriction endonuclease (ENase) sites at their 5' ends. The PCR product is digested with the class-IIS ENase (here, BsaI; GGTCTCN[symbol: see text]NNNN[symbol: see text]), which removes its own recognition sequence, and the ends are self-ligated. Thus, LEIPCR can be used to make plasmid mutant libraries regardless of the nucleotide sequence, and independent of available ENase sites. The resulting library of 10(7) wobble mutants was screened for active Fv by a colony filter lift. A selected mutant was shown to produce between four- and elevenfold more active Fv than the wild type (wt), and fivefold more heavy chain. Mutations outside of the leader peptide were shown not to be involved. The mutated areas of the mRNAs of two different up-mutants may have less secondary structure than the wt. Thus, the sequence of the mRNA of the second leader peptide was limiting to the expression level of heavy-chain and active Fv.

Expression and characterization of a chimeric bifunctional antibody with therapeutic applications.

A simple method is described for the generation of a biologically produced mouse/human chimeric hetero-bifunctional antibody that has dual specificity for human carcinoembryonic Ag and metal chelate haptens. Two large compound chimeric vectors each containing the genetic information to produce a single antibody specificity were sequentially electroporated into the murine nonsecreting hybridoma SP2/0. This led to the isolation of a clone expressing high levels of total IgG (up to 25 micrograms/ml/10(6) cells), 10 to 20% of which showed simultaneous reactivity with both Ag. Binding studies showed that the immunoreactivities and affinity constants for the individual arms of the bifunctional antibody were equivalent to those seen with the parental antibodies.

Improved immune-specificity in monoclonal radioimmunoimaging using dual radionuclide color functional maps.

Diagnostic radioimmunoimaging is potentially limited by tissue localization of radiolabeled antibody products through mechanisms other than antigen binding. Comparing the distributions of reactive and nonreactive products can distinguish tracer in targeted and nontargeted tissues. To achieve this in a single imaging procedure, dual photopeak scintigraphy was performed using 111In and 67Ga products. Melanoma-bearing athymic mice were coadministered intravenously subtype-matched 111In melanoma-reactive and 67Ga melanoma-nonreactive murine monoclonal antibodies. Paired images from 245 and 93 keV windows were processed with a unique dual parameter color display program. The display algorithm expresses pixel counts from paired photo-peak images in polar coordinates and color-encodes angle as hue and magnitude as intensity. The color functional maps permitted ready distinction of immune from nonimmune uptake. Compared with single tracer imaging methods, this technique better depicts antigen distribution.

Antibodies against metal chelates.

Because monoclonal antibodies can recognize and bind to specific groups of atoms such as tumour antigens, they have promise for use in vivo as carriers of radionuclides, drugs or other appended molecules for diagnosis and treatment of disease. Attachment of metal ions to antibodies by means of bifunctional chelating agents can add the diverse nuclear, physical and chemical properties of the metallic elements to these specific binding proteins (ref. 4 and refs therein). With the ultimate aim of engineering probe-binding properties into the antibodies themselves, we have now prepared monoclonal antibodies against the EDTA chelate of indium. These antibodies show a remarkable preference for indium chelates; changing to another metal such as scandium or gallium can decrease the antibody-binding constant by more than three orders of magnitude. These antibodies also introduce a new degree of control over the biological distributions of chelated radionuclides, markedly altering their uptake in tumours and normal organs.

[Toxic, anaphylactoid, and sensitizing properties of mercaptoquinolinates, metals of the 8th and 3d group]. / Toksicheskie, anafilaktoidnye i sensibiliziruiushchie svoistva merkaptokhinolinatov metallov VIII i III grupp.

A study was made of sensitizing, toxic and anaphylactoid action of eitht compounds of metals belonging to groups VIII and III of the periodic system. It was established that the activity depends on the nature of the central atom in the electronic structure of a complex salt ligand. The mechanism of histamine release was shown to be similar to that of specific antigen and to be related to preservation of the respiratory processes, of the microtubular and cell cyclic nucleotide systems. The sensitizing action of the compounds was shown by the reactions of the immediate and delayed types. Moreover, its realization did not require the presence of high molecular carriers, thus indicating that the latter is formed in the body.