Symmetric and asymmetric squarylium dyes as noncovalent protein labels: a study by fluorimetry and capillary electrophoresis

Noncovalent interactions between two squarylium dyes and various model proteins have been explored. NN127 and SQ-3 are symmetric and asymmetric squarylium dyes, respectively, the fluorescence emissions of which have been shown to be enhanced upon complexation with proteins such as bovine serum albumin (BSA), human serum albumin (HSA), beta-lactoglobulin A, and trypsinogen. Although these dyes are poorly soluble in aqueous solution, they can be dissolved first in methanol followed by dilution with aqueous buffer without precipitation, and are then suitable for use as fluorescent labels in protein determination studies. The nature of interactions between these dyes and proteins was studied using a variety of buffer systems, and it was found that electrostatic interactions are involved but not dominant. Dye/protein stoichiometries in the noncovalent complexes were found to be 1:1 for SQ-3, although various possible stoichiometries were found for NN127 depending upon pH and protein. Association constants on the order of 10(5) and 10(7) were found for noncovalent complexes of SQ-3 and NN127, respectively, with HSA, indicating stronger interactions of the symmetric dye with proteins. Finally, HSA complexes with NN127 were determined by capillary electrophoresis with laser-induced fluorescence detection (CE-LIF). In particular, NN127 shows promise as a reagent capable of fluorescently labeling analyte proteins for analysis by CE-LIF without itself being significantly fluorescent under the aqueous solution conditions studied herein.     

Advantages of larval control for African malaria vectors: Low mobility and behavioural responsiveness of immature mosquito stages allow high effective coverage

Background  

Based on sensitivity analysis of the MacDonald-Ross model, it has long been argued that the best way to reduce malaria transmission is to target adult female mosquitoes with insecticides that can reduce the longevity and human-feeding frequency of vectors. However, these analyses have ignored a fundamental biological difference between mosquito adults and the immature stages that precede them: adults are highly mobile flying insects that can readily detect and avoid many intervention measures whereas mosquito eggs, larvae and pupae are confined within relatively small aquatic habitats and cannot readily escape control measures.     

Comparative performance of the Mbita trap, CDC light trap and the human landing catch in the sampling of Anopheles arabiensis, An. funestus and culicine species in a rice irrigation in western Kenya

Background

IL-1β and IL-1RA levels are higher in the serum of cerebral malaria patients than in patients with mild malaria. Recently, the level of IL1B expression was reported to be influenced by a polymorphism in the promoter of IL1, IL1B -31C>T.

Methods

To examine whether polymorphisms in IL1B and IL1RA influence the susceptibility to cerebral malaria, IL1B -31C>T, IL1B 3953C>T, and IL1RA variable number of tandem repeat (VNTR) were analysed in 312 Thai patients with malaria (109 cerebral malaria and 203 mild malaria patients).

Results

In this population, IL1B -31C>T and IL1RA VNTRwere detected, while IL1B 3953C>T (i.e., IL1B 3953T) was not observed in the polymorphism screening for 32 patients. Further analyses for IL1B -31C>T and IL1RA VNTR in 110 cerebral malaria and 206 mild malaria patients showed no significant association of these polymorphisms with cerebral malaria.

Conclusion

The present results suggest that IL1B -31C>T and IL1RA VNTR polymorphisms do not play a crucial role in susceptibility or resistance to cerebral malaria.     

A modified nucleotide in the poly(A) tract of maize RNA

poly(A)+ RNA was isolated from maize by affinity chromatography on columns of oligo(dT)-cellulose. A modified nucleotide ('X') was detected in ribonuclease T2 digests of the RNA as part of a resistant dinucleotide. The dinucleotide was detected by means of the polynucleotide kinase-mediated transfer of a radioactive phosphate atom from adenosine triphosphate to the 5'-OH position of the dinucleotide. Intact poly(A) tracts were released from poly(A)+ RNA by digestion with ribonuclease T1 and A in a high salt buffer and were isolated by oligo(dT)-cellulose chromatography. The poly(A) preparation was found to consist of a series of polyadenylate fragments which varied in chain length from approximately 17 to greater than 70. The modified nucleotide was shown to occupy an internal position in these poly(A) tracts.     

MalariaSphere: A greenhouse-enclosed simulation of a natural Anopheles gambiae (Diptera: Culicidae) ecosystem in western Kenya

Background  

The development and implementation of innovative vector control strategies for malaria control in Africa requires in-depth ecological studies in contained semi-field environments. This particularly applies to the development and release of genetically-engineered vectors that are refractory to Plasmodium infection. Here we describe a modified greenhouse, designed to simulate a natural Anopheles gambiae Giles ecosystem, and the first successful trials to complete the life-cycle of this mosquito vector therein.     

Single-molecule anisotropy imaging

A novel method, single-molecule anisotropy imaging, has been employed to simultaneously study lateral and rotational diffusion of fluorescence-labeled lipids on supported phospholipid membranes. In a fluid membrane composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, in which the rotational diffusion time is on the order of the excited-state lifetime of the fluorophore rhodamine, a rotational diffusion constant, D(rot) = 7 x 10(7) rad(2)/s, was determined. The lateral diffusion constant, measured by direct analysis of single-molecule trajectories, was D(lat) = 3.5 x 10(-8) cm(2)/s. As predicted from the free-volume model for diffusion, the results exhibit a significantly enhanced mobility on the nanosecond time scale. For membranes of DPPC lipids in the L(beta) gel phase, the slow rotational mobility permitted the direct observation of the rotation of individual molecules characterized by D(rot) = 1.2 rad(2)/s. The latter data were evaluated by a mean square angular displacement analysis. The technique developed here should prove itself profitable for imaging of conformational motions of individual proteins on the time scale of milliseconds to seconds.