The binding of lanthanides to non-immune rabbit immunoglobulin G and its fragments.

The binding of Gd(III) to rabbit IgG (immunoglobulin G) and the Fab (N-terminal half of heavy and light chain), (Bab')2 (N-terminal half of heavy and light chains joined by inter-chain disulphide bond), Fc (C-terminal half of heavy-chain dimer)and pFc' (C-terminal quarter of heavy-chain dimer) fragments was demonstrated by measurements of the enhancement of the solvent-water proton relaxation rates in the appropriate Gd(III) solutions. At pH 5.5 there are six specific Gd(III)-binding sites on the IgG. These six sites can be divided into two classes; two very 'tight' sites on the Fc fragment (Kd approx. 5 muM) and two weaker sites on each Fab region (Kd approx. 140 muM). Ca(II) does not apparently compete for these metal-binding sites. The metal-binding parameters for IgG can be explained as the sum of the metal binding to the isolated Fab and Fc fragments, suggesting that there is no apparent interaction between the Fab and Fc regions in the IgG molecule. The binding of Gd(III) to Fab and Fc fragments was also monitored by measuring changes in the electron-spin-resonance spectrum of Gd(III) in the presence of each fragment and also by monitoring the effects of Gd(III) on the protein fluorescence at 340 nm (excitation 295 nm). The fluorescence of Tb(III) solutions of 545 nm (excitation 295 nm) is enhanced slightly on addition of Fab or Fc.     

Selective inhibition of prostaglandin biosynthesis by gold salts and phenylbutazone

Gold salts and phenylbutazone selectively inhibit the synthesis of PGF_2α and PGE₂ respectively. Lowered production of one prostaglandin species is accompanied by an increased production of the other. Selective inhibition by these drugs was observed in the presence of adrenaline, reduced glutathione and copper sulphate under conditions when most anti-inflammatory compounds inhibited PGE₂ and PGF_2α syntheses equally. It is postulated that selective inhibitors may have a different mode of action and beneficial effects may be related to the endogenous ratio of PGE to PGF required for normal function.     

Gas exchange characteristics and nitrogen relations of two Mediterranean root hemiparasites:Bartsia trixago andParentucellia viscosa

Plant height, light-saturated rates of photosynthesis (A _max) and foliar nitrogen concentration (N ₁) were measured forBartsia trixago under field conditions in Mallorca. All three variables were postively correlated, and were also positively related to the abundance of nitrogen-fixing legumes in the associated vegetation (putative host species).A _max forB. trixago ranged from 7.7 to 18.8 mol m^-2 s^-1; similar rates were measured for a second hemiparasiteParentucellia viscosa, and both species were within the range of rates measured for six putative hosts (10.6–19.2 mol m^-2 s^-1). Fertilization of unattachedB. trixago plants with inorganic nitrogen (ammonium nitrate) elicited neither the growth nor the photosynthetic responses observed in plants considered to be parasitic on legumes and in receipt of an enriched organic nitrogen supply. Both hemiparasites had high diurnal leaf conductances (g _s) (469–2291 mmol m^-2 s^-1) and were at the upper end of the range of those measured in putative hosts (409–879 mmol m^-2 s^-1). In contrast with the latter, high nocturnal rates ofg _s were also recorded for the two hemiparasites (517–1862 mmol m^-2 s^-1). There was no clear relationship between eitherA _max orN ₁ and eitherg _s, transpiration (E) or water use efficiency (A _max/E) inB. trixago plants. The economics of water loss appear to be independent of both the supply of nitrogen from the host and autotrophic carbon fixation.     

A streptavidin-biotin binding system that minimizes blocking by endogenous biotin

Pretargeted radioimmunotherapy specifically targets radiation to tumors using antibody-streptavidin conjugates followed by radiolabeled biotin. A potential barrier to this cancer therapy is the presence of endogenous biotin in serum, which can block the biotin-binding sites of the antibody-streptavidin conjugate before the administration of radiolabeled biotin. Serum-derived biotin can also be problematic in clinical diagnostic applications. Due to the extremely slow dissociation of the biotin-streptavidin complex, this endogenous biotin can irreversibly block the biotin-binding sites of streptavidin and reduce therapeutic efficacy, as well as reduce sensitivity in diagnostic assays. We tested a streptavidin mutant (SAv-Y43A), which has a 67-fold lower affinity for biotin than wild type streptavidin, and three bivalent bis-biotin constructs as replacements for wild-type streptavidin and biotin used in pretargeting and clinical diagnostics. Biotin dimers were engineered with certain parameters including water solubility, biotinidase resistance, and linker lengths long enough to span the distance between two biotin-binding sites of streptavidin. The bivalent biotins were compared to biotin in exchange, retention, and off-rate assays. The faster off-rate of SAv-Y43A allowed efficient exchange of prebound biotin by the biotin dimers. In fluorescent competition experiments, the biotin dimer ligands displayed high avidity binding and essentially irreversible retention with SAv-Y43A. The off-rate of a biotinidase-stabilized biotin dimer from SAv-Y43A was 4.36 x 10(-)(6) s(-)(1), over 640 times slower compared to biotin. These findings strongly suggest that employing a mutant streptavidin in concert with a bivalent biotin can mitigate the deleterious impact of endogenous biotin, by allowing exchange of bound biotin and retention of the biotin dimer carriers.     

Immune- and enzyme histochemical characterisation of leukocyte populations within lymphoid and mucosal tissues of Atlantic halibut (Hippoglossus hippoglossus)

Leukocyte populations within the kidney, spleen, posterior intestine and gills of Atlantic halibut were investigated using a panel of histological, enzyme- and immunohistochemical methods. In the kidney and spleen, a diverse population of leukocytes was associated with the extensive network of sinusoids and larger blood vessels present in these tissues. IgM+ cells (B-cells, plasma cells and IgM-bearing macrophages) and large mononuclear cells showing reactivity for non-specific esterase (NSE) and acid phosphatase (ACP), representing macrophage populations, were often associated with vessel walls that were also the site of trapping of fluorescent microspheres. In the kidney, trapping of 0.1 and 0.5 microm diameter microspheres occurred at these sites but in the spleen, the 0.1 microm diameter microspheres were retained in ellipsoids. The lymphoid tissues of the kidney and spleen possessed a spread population of 5'-nucleotidase+ (5'N+) cells but compartmentalisation of the splenic white pulp was suggested by an absence of these 5'N+ reticular cells in areas associated with melanomacrophage accumulations and in areas rich in IgM+ cells. A striking feature of the mucosal tissues was the diversity of leukocyte populations within the epithelium particularly of the posterior intestine, including IgM+ cells and NSE+, ACP+ and 5'N+ mononuclear cells. Although limited in numbers in the posterior intestine, IgM+ cells were more common in the epithelium than in the lamina propria. In the gills, leukocytes as detected by enzymatic reactivity were scarce, but IgM+ cells were very abundant in the stratified epithelium of the gill arch and filaments. The difference in distribution of these leukocyte populations between the intestines and gills suggested a compartmentalisation of the mucosal immune system and the need to assess the immunological competence of mucosal tissues in Atlantic halibut.     

A Genome-Wide Scan for Breast Cancer Risk Haplotypes among African American Women

Genome-wide association studies (GWAS) simultaneously investigating hundreds of thousands of single nucleotide polymorphisms (SNP) have become a powerful tool in the investigation of new disease susceptibility loci. Haplotypes are sometimes thought to be superior to SNPs and are promising in genetic association analyses. The application of genome-wide haplotype analysis, however, is hindered by the complexity of haplotypes themselves and sophistication in computation. We systematically analyzed the haplotype effects for breast cancer risk among 5,761 African American women (3,016 cases and 2,745 controls) using a sliding window approach on the genome-wide scale. Three regions on chromosomes 1, 4 and 18 exhibited moderate haplotype effects. Furthermore, among 21 breast cancer susceptibility loci previously established in European populations, 10p15 and 14q24 are likely to harbor novel haplotype effects. We also proposed a heuristic of determining the significance level and the effective number of independent tests by the permutation analysis on chromosome 22 data. It suggests that the effective number was approximately half of the total (7,794 out of 15,645), thus the half number could serve as a quick reference to evaluating genome-wide significance if a similar sliding window approach of haplotype analysis is adopted in similar populations using similar genotype density.