IVIS visualization of individual first-generation microfilariae from an animal infected with parasites transfected with pBACII-BmGLuc-FLuc and pBmCDH.

Panel A: IVIS visualization of a 384 well plate with each well containing an individual microfilariae. Arrows highlight positive wells. Panel B: GLuc activity in wells identified as negative (n = 374) or positive (n = 10) by IVIS.</p

Quantification of IVIS signals in animals infected with transgenic parasites and treated with flubendazole.

Parasites were transfected with pBACII-BmGLuc-FLuc and pBmCDTH and the animals labeled “Treated” (n = 2) were treated with flubendazole as described in Materials and Methods. The animal labeled “Untreated” (n = 1) was a contemporaneous gerbil infected with parasites transfected with pBACII-BmGLuc-FLuc and pBmCDTH, but not treated with flubendazole. Values represent total photon counts in each animal imaged.</p

Quantification of IVIS signals in animals infected with transgenic parasites: Values represent total photon counts in each animal imaged.

Transfected rep 1 and Trasnfected rep2 = results from two different animals infected with parasites transfected with pBACII-BmGLuc-FLuc and pBmCDTH. Untransfected = results from a contemporaneous control animal infected with untransfected parasites. The graph shown is a representative experiment taken from five separate experiments that were carried out overall. Each experiment consisted of between 2 and 3 animals which were infected with parasites transfected with pBACII-BmGLuc-FLuc and pBmCDTH, together with control animals infected contemporaneously, either with untransfected parasites (shown here) or with parasites transfected with pBACII-BmGLuc-MCS plus pBmCDTH (e.g. Fig 2).</p

IVIS Visualization of transgenic parasites in infected gerbils: Gerbils were infected by IP injection with transgenic parasites and the animals imaged periodically beginning at 7 days through 155 days post-infection, as described in Materials and Methods.

Colors represent the relative level of luciferase activity ranging from low (blue), to medium (green), to high (yellow, red). pBACII-BmGLuc-FLuc = animal infected with parasites transfected with pBACII-BmGLuc-FLuc plus pBmCDTH and pBACII-BmGLuc-MCS = animal infected with parasites transfected with pBACII-BmGLuc-MCS plus pBmCDTH. Arrows point to images of the boxed area on the abdomens of the animals at 155 days post-infection that have been enlarged for clarity. The images shown are representatives taken from five separate experiments that were carried out overall. Each experiment consisted of between 2 and 3 animals which were infected with parasites transfected with pBACII-BmGLuc-FLuc and pBmCDTH, together with control animals infected contemporaneously, either with parasites transfected with pBACII-BmGLuc-MCS plus pBmCDTH (shown here) or untransfected parasites (e.g. Fig 3).</p

Quantification of IVIS signals in animals infected with transgenic parasites cryopreserved with different methods.

Parasites were transfected with pBACII-BmGLuc-FLuc and pBmCDTH and cryopreserved with three different methods as described in Materials and Methods. Values represent total photon counts in each animal imaged. The results shown are an example from two independent experiments.</p

IVIS visualization of animals infected IP or SQ with parasites transfected with pBACII-BmGLuc-FLuc and pBmCDTH.

Panel A: Animal infected IP with parasites transfected with pBACII-BmGLuc-FLuc and pBmCDTH. Panel B: Animal infected SQ with parasites transfected with pBACII-BmGLuc-FLuc and pBmCDTH. Arrows point to the boxed areas on the full animal images that have been enlarged for clarity.</p

Feature map of pBACII-BmGLuc-FLuc: Amp = β lactamase antibiotic selection gene.

ITR = piggyBac inverted terminal repeats GLuc = Gaussia princeps luciferase ORF. FLuc ORF = firefly luciferase ORF. Arrows on the plasmid map indicate direction of transcription.</p

Fabrication of Flexible FCI/PDMS Electromagnetic Shielding Composites Based on Pulsed Magnetic Field-Induced Alignment

Conductive polymer composites have been utilized in the field of electromagnetic interference (EMI) shielding, albeit requiring a high concentration of conductive fillers to achieve desirable EMI performance. To address this issue and enable the creation of superior EMI shielding composites with reduced filler loadings, this study employed a pulsed magnetic field featuring an amplitude of 0.7 T, a pulse width of 10 μs, and a frequency of 100 Hz to align flaky carbonyl iron (FCI) in poly(dimethylsiloxane) (PDMS). This method resulted in an improved EMI shielding performance of the composites. The outcomes revealed that the pulsed magnetic field effectively controlled the orientation of the FCI, forming a conductive network structure, with the average orientation angle of the FCI reaching 69.3°. The aligned composites exhibited a significant improvement in EMI shielding effectiveness, with the enhancement effect reaching 37.53% and the EMI shielding effectiveness reaching 24.87 dB. Moreover, the flexible tensile properties of the aligned composites were superior to those of the unaligned composites, particularly the elongation at break, which reached 197.46%. The concordance between the theoretical analysis and experimental results affirms the efficacy of the microsecond pulsed magnetic field in enhancing the EMI shielding performance of composite materials. Ultimately, the high-performance, flexible electromagnetic shielding composite materials prepared in this study demonstrate potential for use in advanced electronic equipment

Development of a toolkit for <i>piggyBac</i>-mediated integrative transfection of the human filarial parasite <i>Brugia malayi</i>

<div><p>Background</p><p>The human filarial parasites cause diseases that are among the most important causes of morbidity in the developing world. The elimination programs targeting these infections rely on a limited number of drugs, making the identification of new chemotherapeutic agents a high priority. The study of these parasites has lagged due to the lack of reverse genetic methods.</p><p>Methodology/Principal findings</p><p>We report a novel co-culture method that results in developmentally competent infective larvae of one of the human filarial parasites (<i>Brugia malayi</i>) and describe a method to efficiently transfect the larval stages of this parasite. We describe the production of constructs that result in integrative transfection using the <i>piggyBac</i> transposon system, and a selectable marker that can be used to identify transgenic parasites. We describe the production and use of dual reporter plasmids containing both a secreted luciferase selectable marker and fluorescent protein reporters that will be useful to study temporal and spatial patterns of gene expression.</p><p>Conclusions/Significance</p><p>The methods and constructs reported here will permit the efficient production of integrated transgenic filarial parasite lines, allowing reverse genetic technologies to be applied to all life cycle stages of the parasite.</p></div

Primers used in the production of the <i>piggyBac</i> plasmids.

<p>Primers used in the production of the <i>piggyBac</i> plasmids.</p