Journal: bioRxiv

Article Title: Sticky Situations: Bacterial Attachment Deciphered by Interferometry of Silicon Microstructures

doi: 10.1101/793125

Figure Lengend Snippet: Different microtopologies promote varying degrees of wetting, in turn promoting differences in bacterial adhesion amongst strains. (A) Contact angle values (left panel) of MB measured after 200 s on oxidized (OX) and amine-functionalized (AMINE) pores, pillars, and planar Si substrates suggest that the buffer does not completely infiltrate the pore topologies, as illustrated (right panel). MB positions partially within the microtopologies, forming small vapor pockets in pore structures, characteristic of a Cassie-Baxter wetting regime. (B) PRISM assays of E. coli WT, E. coli K-12, and S. epidermidis adhesion to OX Si pores and Si pillars over time (n = 3). Measurements were collected every 2 minutes, but error bars are placed every 10 minutes for visual clarity. (C) False-colored SEM images of bacterial cells within the microstructured pores (left) and pillars (right) reveal attachment behavior to the oxidized surfaces for each of the different strains. Scale bars represents 2 μm. (D) Summary of the Δ2nL (%) values for each E. coli WT, E. coli K-12, and S. epidermidis (abbreviated SE) after 120 min of accumulation onto oxidized pores and pillars (n = 3).

Article Snippet: As preliminary tests, the PRISM responses of three non-pathogenic bacteria strains, namely the laboratory mutant Escherichia coli ( E. coli ) K-12, clinically isolated E. coli (ATCC 25922, termed as WT), and Staphylococcus epidermidis ( S. epidermidis ATCC 14990), were observed over time, with Δ2nL (%) corresponding to the accumulation of cells in the topologies ( ).

Techniques:

Journal: bioRxiv

Article Title: Sticky Situations: Bacterial Attachment Deciphered by Interferometry of Silicon Microstructures

doi: 10.1101/793125

Figure Lengend Snippet: Motility is an underlying factor leading to bacterial adhesion within the microstructures. (A) PRISM of E. coli with a cheZ deletion causing excessive tumbling, E. coli without flagella, and E. coli with deleted chemotaxis receptors exhibit differences in adhesion behavior to oxidized (OX) pore and pillar substrates. Ryu flagellar staining reveals the presence or absence of flagella on the strains in optical microscope images. Scale bars represents 1 μm. (B) Summary of final Δ2nL (%) values reached after 120 min for the genetically modified E. coli mutants, E. coli WT, E. coli K-12, and S. epidermidis. (C) Semi-soft agar motility assays of E. coli WT, E. coli K-12, and S. epidermidis reveal that E. coli WT is significantly more motile than the K-12 strain (n = 4), while S. epidermidis is immotile. Optical microscope images after Ryu staining of E. coli WT and E. coli K-12 (right) reveal that the K-12 cells tend to have fewer flagella present. Scale bar represents 2 μm.

Article Snippet: As preliminary tests, the PRISM responses of three non-pathogenic bacteria strains, namely the laboratory mutant Escherichia coli ( E. coli ) K-12, clinically isolated E. coli (ATCC 25922, termed as WT), and Staphylococcus epidermidis ( S. epidermidis ATCC 14990), were observed over time, with Δ2nL (%) corresponding to the accumulation of cells in the topologies ( ).

Techniques: Chemotaxis Assay, Staining, Microscopy, Genetically Modified

Journal: bioRxiv

Article Title: Sticky Situations: Bacterial Attachment Deciphered by Interferometry of Silicon Microstructures

doi: 10.1101/793125

Figure Lengend Snippet: Substrate surface charge plays a role in bacterial adhesion. (A) AMINE Si surfaces yield a positive surface charge, while OX surfaces yield a negatively charged surface in motility buffer. (B) Zeta potential measurements of bacteria suspended in MB (n = 3) measure the surface charge of the cells. (C) Comparison bar graph of Δ2nL (%) PRISM values for OX and AMINE after 120 min of accumulation time. (D) Individual real-time PRISM accumulation curves for E. coli WT, E. coli K-12, and S. epidermidis on AMINE (positively charged) pore and pillar substrates. Note that the scale of the E. coli WT attachment curves is 5-times greater than that of E. coli K-12 and S. epidermidis. (E) False-colored SEM images of (i) E. coli WT on AMINE pores. (ii) E. coli WT on AMINE pillars reveal bridging between pillars and elongation of E. coli cells, in contrast to (iii) E. coli K-12 on AMINE pillars and (iv) S. epidermidis on AMINE pillars.

Article Snippet: As preliminary tests, the PRISM responses of three non-pathogenic bacteria strains, namely the laboratory mutant Escherichia coli ( E. coli ) K-12, clinically isolated E. coli (ATCC 25922, termed as WT), and Staphylococcus epidermidis ( S. epidermidis ATCC 14990), were observed over time, with Δ2nL (%) corresponding to the accumulation of cells in the topologies ( ).

Techniques: Zeta Potential Analyzer, Bacteria, Comparison

Journal: PLoS ONE

Article Title: FLT3 Mutations in Early T-Cell Precursor ALL Characterize a Stem Cell Like Leukemia and Imply the Clinical Use of Tyrosine Kinase Inhibitors

doi: 10.1371/journal.pone.0053190

Figure Lengend Snippet: Mutational events in ETP-ALL compared to non-ETP T-ALL.

Article Snippet: FLT3mut (ITD/TKD) were analyzed using a commercially available FLT3 mutation assay (InVivoScribe Technologies, San Diego, USA).

Techniques:

Journal: PLoS ONE

Article Title: FLT3 Mutations in Early T-Cell Precursor ALL Characterize a Stem Cell Like Leukemia and Imply the Clinical Use of Tyrosine Kinase Inhibitors

doi: 10.1371/journal.pone.0053190

Figure Lengend Snippet: Combinations of antigens as a surrogate marker for FLT3 mutations in ETP-ALL.

Article Snippet: FLT3mut (ITD/TKD) were analyzed using a commercially available FLT3 mutation assay (InVivoScribe Technologies, San Diego, USA).

Techniques: Marker

Journal: PLoS ONE

Article Title: FLT3 Mutations in Early T-Cell Precursor ALL Characterize a Stem Cell Like Leukemia and Imply the Clinical Use of Tyrosine Kinase Inhibitors

doi: 10.1371/journal.pone.0053190

Figure Lengend Snippet: Molecular characteristics of FLT3 mut ETP-ALL versus FLT3 wt ETP-ALL patients.

Article Snippet: FLT3mut (ITD/TKD) were analyzed using a commercially available FLT3 mutation assay (InVivoScribe Technologies, San Diego, USA).

Techniques: Mutagenesis

Journal: PLoS ONE

Article Title: FLT3 Mutations in Early T-Cell Precursor ALL Characterize a Stem Cell Like Leukemia and Imply the Clinical Use of Tyrosine Kinase Inhibitors

doi: 10.1371/journal.pone.0053190

Figure Lengend Snippet: Fourty-eight hours (hrs) after transfection, cells were seeded and cultured for additionally 48 hrs with tyrosine kinase inhibitors (PKC412, TKI258, and Sorafenib) and chemotherapy (AraC). Cell proliferation was measured using the WST-1 proliferation reagent. The mean optical density (OD) values corresponding to non-treated FLT3-ITD transfected cells were taken as 100%. The results were expressed in percentages of the OD of treated versus untreated control cells. Two experiments were performed in duplicates. For each drug two different doses were used. All results were expressed as means ±S.D. A : Jurkat cells. B : MOLT4 cells. C : BE13 cells.

Article Snippet: FLT3mut (ITD/TKD) were analyzed using a commercially available FLT3 mutation assay (InVivoScribe Technologies, San Diego, USA).

Techniques: Transfection, Cell Culture, Control

Journal: Molecular Biology of the Cell

Article Title: Direct interaction of the Golgi V-ATPase a-subunit isoform with PI(4)P drives localization of Golgi V-ATPases in yeast

doi: 10.1091/mbc.E17-05-0316

Figure Lengend Snippet: Down-regulation of intracellular PI(4)P level induces mislocalization and impaired function of V-ATPases containing Stv1. (A, B) Localization of Stv1-GFP and Vph1-mCherry in pik1-39 mutant. (A) Localization at the permissive temperature of 25°C (representative of n = 511 yeast cells) or (B) after a 2-h incubation at the restrictive temperature of 34°C (representative of n = 1203 yeast cells). Vacuoles are marked by Vph1-mCherry. (C) Mean ± SEM of Pearson correlation coefficient for colocalization of GFP and mCherry from three independent experiments is represented in a histogram ( p < 0.00005). (D) Growth assay of congenic WT, vph1Δ , pik1-139 , and vph1Δ pik1-139 cells in media buffered to pH 5.0 (left) and pH 7.5 (right). Cells from each strain were diluted to the same density, then 10-fold serial dilutions (left to right) were made and pinned to YEPD plates buffered to the indicated pH. The growth assay shown is representative of two independent experiments.

Article Snippet: Functional effects of STV1 mutations have been analyzed in vph1∆ mutant background ( Finnigan et al. , 2012 ).

Techniques: Mutagenesis, Incubation, Growth Assay

Journal: Molecular Biology of the Cell

Article Title: Direct interaction of the Golgi V-ATPase a-subunit isoform with PI(4)P drives localization of Golgi V-ATPases in yeast

doi: 10.1091/mbc.E17-05-0316

Figure Lengend Snippet: Localization of Stv1NT-mCherry to puncta. Stv1NT-mCherry (in which the transmembrane C-terminal domain of Stv1 is replaced by mCherry) was expressed from the genomic STV1 locus in (A) S. cerevisiae wild-type strain BY4741; (B) BY4741 vph1∆ cells, which lack the second V o a-subunit isoform; and (C) BY4741 vac14∆ cells, which lack PI(3,5)P 2 . For each set of panels, differential interference contrast images are shown on the left and mCherry fluorescence is shown on the right.

Article Snippet: Functional effects of STV1 mutations have been analyzed in vph1∆ mutant background ( Finnigan et al. , 2012 ).

Techniques: Fluorescence

Journal: Molecular Biology of the Cell

Article Title: Direct interaction of the Golgi V-ATPase a-subunit isoform with PI(4)P drives localization of Golgi V-ATPases in yeast

doi: 10.1091/mbc.E17-05-0316

Figure Lengend Snippet: K84A mutation in Stv1FL causes partial mislocalization of Stv1FL-GFP and impairs function of V-ATPases bearing Stv1. (A) Localization of wild-type Stv1FL-GFP in yeast cells stained with vacuolar dye FM 4-64 (representative of n = 410). (B) Localization of Stv1FL(K84A)-GFP in yeast cells stained with FM 4-64 ( n = 204). (C) Histogram representing mean percentage ± SEM of cells with Stv1FL-GFP (WT or K84A) localized to the vacuole ( p < 0.0005). (D) Serial-dilution growth assay of congenic WT, vma2∆ (representing a total loss of V-ATPase function), vph1Δ , vph1 ∆ stv1 (K84A), and vph1Δ 2µ-STV1 (overexpressing Stv1) cells on YEPD, pH 5 (left), YEPD + 4 mM Zn 2+ (middle), and YEPD, pH 7.5 (right). The growth assay shown is representative of two independent experiments.

Article Snippet: Functional effects of STV1 mutations have been analyzed in vph1∆ mutant background ( Finnigan et al. , 2012 ).

Techniques: Mutagenesis, Staining, Serial Dilution, Growth Assay