Journal: bioRxiv

Article Title: A mRNA-LNP vaccine against Dengue Virus elicits robust, serotype-specific immunity

doi: 10.1101/2021.01.05.425517

Figure Lengend Snippet: A) Schematic of the DENV genome and engineered mRNA construct. An mRNA encoding for the prM and ENV viral proteins was engineered with N-terminal signal peptide sequence, 5’ and 3’ untranslated regions (UTR) flanking the coding sequence, a 3’ poly-A tail, and a 5’ cap-1 structure. In vitro synthesized mRNA is encapsulated in a lipid nanoparticle for use in in vitro and in vivo experiments. B) 293T cells were transfected with the in vitro transcribed mRNA encoding for the wild-type sequence (WT), or a mutant version with amino acid substitutions in the fusion-loop epitope (ΔFL). Lysate was analyzed by western blot with the domain III specific 1A1D-2 monoclonal antibody and the fusion-loop specific 4G2 monoclonal antibody. C) Supernatant from transfected cells was purified and concentrated through ultracentrifugation and analyzed for VLPs by western blots with the 1A1D-2 monoclonal antibody or anti-GAPDH. Unpurified cell lysate from WT mRNA transfected cells is included as a control. Shown are representative blots. D) Electron microscopy image of VLPs from purified supernatant of transfected 293T cells showing homogenous shape and size of approximately 30nm.

Article Snippet: Membranes were blotted with envelope domain III specific 1A1D-2 (1:600) monoclonal antibody (CDC Arbovirus Reference Collection) or envelope fusion-loop specific 4G2 (3.33 mg/ml) (BEI Cat# NR-50327, Novus Biologicals Cat# NBP2-52709FR).

Techniques: Construct, Sequencing, In Vitro, Synthesized, In Vivo, Transfection, Mutagenesis, Western Blot, Purification, Control, Electron Microscopy

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Tissue Responses to Shiga Toxin in Human Intestinal Organoids

doi: 10.1016/j.jcmgh.2020.02.006

Figure Lengend Snippet: HIOs express the Stx receptor Gb3 and are susceptible to Stx by luminal or basolateral delivery. ( A ) HIOs express the Stx receptor Gb3. Cryosections were stained for DNA (4′,6-diamidino-2-phenylindole, blue), and antibody to Gb3 (red). Intense staining was observed on epithelial cells. Scale bar : 50 μm. ( B ) Stx causes loss of epithelial barrier function. Representative images of intact HIOs loaded with the fluorescent dye FITC, and exposed to Stx2a (30 ng) introduced by microinjection (luminal exposure) or added to the medium (interstitial exposure) over time. Scale bar : 100 μm. ( C and D ) Time- and dose-dependent responses to Stx. Toxin was delivered by microinjection (luminal exposure) or added to the medium (interstitial exposure). Epithelial barrier function was assessed by fluorescence retention, quantified by ImageJ, and plotted as the percentage mean at time zero ± SD, n = 3. Values at 72 hours compared with time zero were assessed using 1-way analysis of variance, Dunnett’s posttest. ( C ) HIO sensitivity to Stx2a. ( D ) HIO sensitivity to Stx1.

Article Snippet: Toxins and antibodies were obtained from the following sources: Stx2a (cat. NR-4478, lot 62905537; BEI Resources, Manassas, VA), Stx1 (cat. NR-857, lot 58338341; BEI Resources), TcdA (cat. 152C, lot 15215A5; List Biological Laboratories, Inc, Campbell, CA), TcdB (cat. 155L, lot 15521A2; List Biological Laboratories Inc), microcystin-LR (cat. ALK-350-012-C100, lot L30360; Enzo Life Sciences), LPS (cat. tlrl-3pelps, lot L3P-32-02; InvivoGen, San Diego, CA); monoclonal Stx2, subunit A (cat. NR-845, lot 4469047; BEI Resources), monoclonal Stx2, subunit B (cat. NR-9352, lot 57845852; BEI Resources).

Techniques: Staining, Fluorescence

Journal: Genes

Article Title: Improved DNA Extraction and Illumina Sequencing of DNA Recovered from Aged Rootless Hair Shafts Found in Relics Associated with the Romanov Family

doi: 10.3390/genes13020202

Figure Lengend Snippet: Description of the hair shaft samples that were used in this study and purification methods that were used during DNA extraction.

Article Snippet: The MinElute columns were, therefore, replaced first with the Ultraclean Production (UCP) MinElute columns (cat # 56204, Qiagen).

Techniques: Purification, Extraction

Journal: Genes

Article Title: Improved DNA Extraction and Illumina Sequencing of DNA Recovered from Aged Rootless Hair Shafts Found in Relics Associated with the Romanov Family

doi: 10.3390/genes13020202

Figure Lengend Snippet: Bioanalyzer electropherograms of reagent blank libraries when DNA purification was performed using ( A ) MinElute columns, or ( B ) magnetic beads from G-Biosciences.

Article Snippet: The MinElute columns were, therefore, replaced first with the Ultraclean Production (UCP) MinElute columns (cat # 56204, Qiagen).

Techniques: DNA Purification, Magnetic Beads