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  • 99
    ATCC skmel28
    Knockdown of STAG2 or STAG3 decreases BRAFi sensitivity in BRAF mutant melanoma cells ( a ) Viability of A375 cells after treatment with varying concentrations of dabrafenib for 3 d. Experiment was performed 3 times. Data are mean ± s.e.m. ( b ) A375 cells were treated with dabrafenib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( c ) Viability of <t>SKMEL28</t> cells after treatment with varying concentrations of vemurafenib for 3 d. Experiment was performed 3 times. Data are mean ± s.e.m. ( d ) SKMEL28 cells were treated with vemurafenib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( e ) SKMEL30 cells were treated with trametinib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( f ) SKMEL30 cells were treated with trametinib as indicated in clonogenic growth assays. Experiment was performed 3 times. Scale bar: 5 mm. ( g ) Viability of A375 cells after treatment with varying concentrations of dabrafenib for 3 d. Experiment was performed 3 times. Data are mean ± s.e.m. ( h ) A375 cells were treated with vemurafenib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( i ) WM902-BR cells stably expressing control vector, FLAG-tagged wild-type STAG2 (WT), Lys1083* (K*) or Asp193Asn (DN) mutants were treated with 3 μM vemurafenib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( j ) WM902-BR cells were used in soft agar assays in the presence or absence of 3 μM vemurafenib. Experiment was performed 3 times. Scale bar: 5 mm.
    Skmel28, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 121 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore clorox
    Sodium hypochlorite treatment of glycoproteins to release N-glycans. a) Chemical scheme. b) TLC analysis of ovalbumin before and after bleach treatment. c) MALDI-MS profiles of N-glycans released from several glycoproteins by bleach treatment. d) MALDI-MS profiles of permethylated glycans released from fetuin by PNGase F digestion (top) or bleach treatment (bottom). e) Normal phase (NP) HPLC profiles of AEAB conjugated N-glycans released from fetuin by PNGase F digestion (top) or bleach treatment (bottom). Peaks at 10-17 min were nonglycan peaks presumably generated from oxidized peptide fragments. f) MALDI-MS profiles of human plasma permethylated N-glycans released by PNGase F digestion (top) and bleach (bottom). g) MALDI-MS profiles of N-glycans released by <t>NaClO</t> from egg white (top) and egg yolk (bottom). H: Hexose; N: N-acetylhexosamine; F: fucose; S: sialic acid. The Y-axis scales are not calibrated to reflect quantitative comparison.
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    Millipore sodium periodate
    Sodium hypochlorite treatment of glycoproteins to release N-glycans. a) Chemical scheme. b) TLC analysis of ovalbumin before and after bleach treatment. c) MALDI-MS profiles of N-glycans released from several glycoproteins by bleach treatment. d) MALDI-MS profiles of permethylated glycans released from fetuin by PNGase F digestion (top) or bleach treatment (bottom). e) Normal phase (NP) HPLC profiles of AEAB conjugated N-glycans released from fetuin by PNGase F digestion (top) or bleach treatment (bottom). Peaks at 10-17 min were nonglycan peaks presumably generated from oxidized peptide fragments. f) MALDI-MS profiles of human plasma permethylated N-glycans released by PNGase F digestion (top) and bleach (bottom). g) MALDI-MS profiles of N-glycans released by <t>NaClO</t> from egg white (top) and egg yolk (bottom). H: Hexose; N: N-acetylhexosamine; F: fucose; S: sialic acid. The Y-axis scales are not calibrated to reflect quantitative comparison.
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    Millipore sodium dithionite
    Sodium hypochlorite treatment of glycoproteins to release N-glycans. a) Chemical scheme. b) TLC analysis of ovalbumin before and after bleach treatment. c) MALDI-MS profiles of N-glycans released from several glycoproteins by bleach treatment. d) MALDI-MS profiles of permethylated glycans released from fetuin by PNGase F digestion (top) or bleach treatment (bottom). e) Normal phase (NP) HPLC profiles of AEAB conjugated N-glycans released from fetuin by PNGase F digestion (top) or bleach treatment (bottom). Peaks at 10-17 min were nonglycan peaks presumably generated from oxidized peptide fragments. f) MALDI-MS profiles of human plasma permethylated N-glycans released by PNGase F digestion (top) and bleach (bottom). g) MALDI-MS profiles of N-glycans released by <t>NaClO</t> from egg white (top) and egg yolk (bottom). H: Hexose; N: N-acetylhexosamine; F: fucose; S: sialic acid. The Y-axis scales are not calibrated to reflect quantitative comparison.
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    Millipore calcofluor white
    mRNA expression patterns of meristem and flower development genes revealed by RNA FISH. The mRNAs (magenta) were hybridized with digoxigenin-labeled antisense RNA probes and detected by TSA-Cy5. Cell walls were stained by <t>Calcofluor</t> White and are shown in blue. Left images show RNA FISH signals in a Fire format. Meristems and flowers are outlined by white dashed lines. A, Schematic representation of the Arabidopsis inflorescence shoot apex. CZ, Central zone; PZ, peripheral zone; RM, rib meristem. B to F, H, K, L, and N, The distribution of WUS mRNA in the shoot apex (B), CLV3 (C), STM (D), LFY (E), AP1 (F), AP2 (H), CUC2 (K), FIL (L), and REV (N). I, AP3 mRNA expression in a stage 3 flower. M, FIL mRNA expression in a stage 6 flower. ca, Carpel; pe, petal; se, sepal; st, stamen. G and J, Expression patterns of AP1 translational fusion reporter gAP1 : GFP ) in the shoot apex and AP3 transcriptional reporter pAP3 :: 2BFP for serial sections. Bars = 25 μm.
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    Millipore sodium chlorate
    mRNA expression patterns of meristem and flower development genes revealed by RNA FISH. The mRNAs (magenta) were hybridized with digoxigenin-labeled antisense RNA probes and detected by TSA-Cy5. Cell walls were stained by <t>Calcofluor</t> White and are shown in blue. Left images show RNA FISH signals in a Fire format. Meristems and flowers are outlined by white dashed lines. A, Schematic representation of the Arabidopsis inflorescence shoot apex. CZ, Central zone; PZ, peripheral zone; RM, rib meristem. B to F, H, K, L, and N, The distribution of WUS mRNA in the shoot apex (B), CLV3 (C), STM (D), LFY (E), AP1 (F), AP2 (H), CUC2 (K), FIL (L), and REV (N). I, AP3 mRNA expression in a stage 3 flower. M, FIL mRNA expression in a stage 6 flower. ca, Carpel; pe, petal; se, sepal; st, stamen. G and J, Expression patterns of AP1 translational fusion reporter gAP1 : GFP ) in the shoot apex and AP3 transcriptional reporter pAP3 :: 2BFP for serial sections. Bars = 25 μm.
    Sodium Chlorate, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 352 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore styrene
    mRNA expression patterns of meristem and flower development genes revealed by RNA FISH. The mRNAs (magenta) were hybridized with digoxigenin-labeled antisense RNA probes and detected by TSA-Cy5. Cell walls were stained by <t>Calcofluor</t> White and are shown in blue. Left images show RNA FISH signals in a Fire format. Meristems and flowers are outlined by white dashed lines. A, Schematic representation of the Arabidopsis inflorescence shoot apex. CZ, Central zone; PZ, peripheral zone; RM, rib meristem. B to F, H, K, L, and N, The distribution of WUS mRNA in the shoot apex (B), CLV3 (C), STM (D), LFY (E), AP1 (F), AP2 (H), CUC2 (K), FIL (L), and REV (N). I, AP3 mRNA expression in a stage 3 flower. M, FIL mRNA expression in a stage 6 flower. ca, Carpel; pe, petal; se, sepal; st, stamen. G and J, Expression patterns of AP1 translational fusion reporter gAP1 : GFP ) in the shoot apex and AP3 transcriptional reporter pAP3 :: 2BFP for serial sections. Bars = 25 μm.
    Styrene, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 235 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore sodium tripolyphosphate tpp
    Transfection efficiency of nanoparticles on Neuro2a cells after 4 hours of incubation at 37°C and 5% CO 2 : a ) <t>PEGylated-chitosan-TPP:siGLO</t> nanoparticles; b ) chitosan-TPP:siGLO nanoparticles; c ) siGLO only (negative control); d ) chitosan-TPP nanoparticles (negative control); e ) cells only (negative control) Abbreviation: TPP, sodium <t>tripolyphosphate.</t>
    Sodium Tripolyphosphate Tpp, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 145 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Agilent technologies 28s
    Representative electropherogram of total RNA extracted from pancreatic cancer obtained by endoscopic ultrasound-guided fine-needle aspiration biopsy . The ratio of <t>28S</t> to 18S of ribosomal RNA indicates good quality of total RNA.
    28s, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 94/100, based on 831 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore sodium chloride solution
    Representative electropherogram of total RNA extracted from pancreatic cancer obtained by endoscopic ultrasound-guided fine-needle aspiration biopsy . The ratio of <t>28S</t> to 18S of ribosomal RNA indicates good quality of total RNA.
    Sodium Chloride Solution, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 192 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    Carna cmpd 28
    <t>Cmpd</t> 28 treatment improves insulin-induced Akt phosphorylation and lowers the expression of gluconeogenic genes in obese mice. Ten-week-old ob/ob mice were injected with 0.2 mg/kg body wt i.p. cmpd 28 or vehicle (control [Con]) each day for 3 weeks ( n = 5 mice per group). A : The mice were fasted for 6 h and then injected with 2 IU/kg body wt insulin (ins) through the portal vein. After 3 min, livers were harvested, and total liver extracts were assayed for p-Akt, Akt, Trb3, and β-actin by immunoblot. Densitometric quantification of the immunoblot data is shown in the graph (* P
    Cmpd 28, supplied by Carna, used in various techniques. Bioz Stars score: 89/100, based on 89 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Agilent technologies 28s 18s ratio
    DIMT1L, WBSCR22, and TRMT112 are required for distinct pre-rRNA processing steps, and the pre-rRNA processing defects are conserved in different cell types and do not depend on p53. WI-38, RKO, and HCT116 cells were treated for 3 d with a specific siRNA targeting DIMT1L, WBSCR22, or TRMT112. Paired HCT116 cell lines expressing p53 or not were used (+/+ and –/–). Total RNA was extracted, resolved on denaturing agarose gel, transferred to a nylon membrane, and hybridized with probes. The probes used were as follows: (I, II) LD1844, (III) LD1827, and (IV) LD1828. The detected pre-rRNA species are indicated to the right and schematized. (V, VI) The mature <t>28S</t> and <t>18S</t> rRNAs stained with ethidium bromide. For each sample, the 28S/18S ratio was calculated from Agilent bioanalyzer electropherograms. All RNA species were quantified with a Phosphorimager normalized with respect to the nontargeting control (SCR), and their abundances represented as a heatmap using the color code indicated to the right. The sequences of the siRNAs used are listed in Supplemental Table S3 (WBSCR22#1, TRMT112#2, and DIMT1L#2). Note that in the heatmap for RKO cells, the signal for lane 8 was corrected for loading.
    28s 18s Ratio, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 91/100, based on 321 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore nahs
    Effect of treatment with <t>NaHS</t> on protein levels of Cx40 and Cx43 in peripheral blood lymphocytes of <t>SHR.</t> We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, peripheral blood lymphocytes were harvested and examined by Western blot for the expression levels of Cx40 ( A ) and Cx43 ( B ). After densitometric analysis, the data were expressed as ratios of Cxs to β-actin. The data represent the mean ±SEM of 3 experiments ( n =15 animals in each group). ** P
    Nahs, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 522 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    Agilent technologies pd l1 ihc 28 8 pharmdx
    Prevalence of PD-L1 protein in FFPE Melanoma Specimens using PD-L1 IHC 28-8 <t>pharmDx</t> assay for detection (n=104 specimens). PD-L1 indicates programmed cell death 1-ligand 1; IHC, immunohistochemistry; FFPE indicates formalin-fixed paraffin-embedded.
    Pd L1 Ihc 28 8 Pharmdx, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 96/100, based on 129 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam anti pd l1 antibody 28 8
    Removal of N-linked glycosylation enhances anti-PD-L1 signal in human cancer cells in a variety of bioassays. (A and B) Immunofluorescence confocal microscopy of BT-549 (A) and A549 (B) cells processed with or without deglycosylation by PNGase F (5%) pretreatment stained with DAPI and an <t>anti-PD-L1</t> antibody (Abcam, ab58810). Bar, 10 μm. Quantification is shown to the right. Data are representative of 3 independent experiments, randomly chosen in 3 different fields. (C and D) ELISA of Con A (C) and PD-L1 (clone <t>28-8</t> mAb) (D) levels in BT-549 cells processed with deglycosylation by increasing concentrations of PNGase F (1, 2, 5%) pretreatment for comparison with cells without deglycosylation (PNGase F; 0%). The intensity of Con A and PD-L1 was normalized to that without PNGase F pretreatment and set to 1. (E) ELISA of PD-L1 levels (clone 28-8 mAb) in lung cancer cells processed with deglycosylation by PNGase F (1%) pretreatment for comparison with cells without deglycosylation (0%). Negative control, secondary Ab only control. (F) Left: saturation binding assay of A549 cell lysates binding to anti-PD-L1 clone 28-8 mAb. Right: scatchard plot of cell number binding to anti-PD-L1 antibody transformed from the left. (G) Representative images (left) and quantification (right) of H-score of IHC staining for BLBC (BT-549, BT-20, and MDA-MB-231) and non-BLBC (MCF-7) cancer cell blocks processed with or without deglycosylation by PNGase F (5%) pretreatment. Bar, 50 μm. (H) Representative images (top) and quantification (bottom) of H-score of IHC staining for a panel of lung cancer cell blocks processed with or without deglycosylation by PNGase F (5%) pretreatment. Bar, 50 μm. (A–F) Results are presented as mean ± SD. *p
    Anti Pd L1 Antibody 28 8, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 66 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Santa Cruz Biotechnology vk 28
    <t>VK-28</t> attenuates ROS production, iron deposition, neuronal degeneration, lesion volume, and brain edema after ICH in vivo. Middle-aged male C57BL/6 mice underwent ICH by collagenase injection. DFX, VK-28, or vehicle (5% DMSO in saline) was administered i.p. at 6 h post-ICH and then daily until sacrifice. (a) HEt was injected i.p. 1 h before sacrifice. Representative images show HEt fluorescence intensity one day after ICH, and quantification shows the intensity as fold change of that in the vehicle group. Vehicle: n = 10, DFX: n = 5, VK-28: n = 5. (b) Perls’ staining and quantification of iron + cells at three days after ICH. n = 5 mice/group. (c) Fluoro-Jade B (FJB) was used to stain degenerating neurons at day 3. Representative images and quantification are shown. n = 5 mice/group. (d) Cresyl violet (CV) staining and quantification of surviving neurons. n = 5 mice/group. (e) CV/Luxol fast blue staining was performed on post-ICH day 3, and lesion volume (corrected for brain swilling) was calculated. n = 5 mice/group. (f) The percentage of brain swelling was measured at three days after ICH. n = 5 mice/group. (g) The percentage of brain water content was measured at three days after ICH. n = 6 mice/group. (h) Field selection for ICH brain. * p
    Vk 28, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 91/100, based on 59 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore sodium silicate
    <t>VK-28</t> attenuates ROS production, iron deposition, neuronal degeneration, lesion volume, and brain edema after ICH in vivo. Middle-aged male C57BL/6 mice underwent ICH by collagenase injection. DFX, VK-28, or vehicle (5% DMSO in saline) was administered i.p. at 6 h post-ICH and then daily until sacrifice. (a) HEt was injected i.p. 1 h before sacrifice. Representative images show HEt fluorescence intensity one day after ICH, and quantification shows the intensity as fold change of that in the vehicle group. Vehicle: n = 10, DFX: n = 5, VK-28: n = 5. (b) Perls’ staining and quantification of iron + cells at three days after ICH. n = 5 mice/group. (c) Fluoro-Jade B (FJB) was used to stain degenerating neurons at day 3. Representative images and quantification are shown. n = 5 mice/group. (d) Cresyl violet (CV) staining and quantification of surviving neurons. n = 5 mice/group. (e) CV/Luxol fast blue staining was performed on post-ICH day 3, and lesion volume (corrected for brain swilling) was calculated. n = 5 mice/group. (f) The percentage of brain swelling was measured at three days after ICH. n = 5 mice/group. (g) The percentage of brain water content was measured at three days after ICH. n = 6 mice/group. (h) Field selection for ICH brain. * p
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    Millipore sodium chlorite
    <t>VK-28</t> attenuates ROS production, iron deposition, neuronal degeneration, lesion volume, and brain edema after ICH in vivo. Middle-aged male C57BL/6 mice underwent ICH by collagenase injection. DFX, VK-28, or vehicle (5% DMSO in saline) was administered i.p. at 6 h post-ICH and then daily until sacrifice. (a) HEt was injected i.p. 1 h before sacrifice. Representative images show HEt fluorescence intensity one day after ICH, and quantification shows the intensity as fold change of that in the vehicle group. Vehicle: n = 10, DFX: n = 5, VK-28: n = 5. (b) Perls’ staining and quantification of iron + cells at three days after ICH. n = 5 mice/group. (c) Fluoro-Jade B (FJB) was used to stain degenerating neurons at day 3. Representative images and quantification are shown. n = 5 mice/group. (d) Cresyl violet (CV) staining and quantification of surviving neurons. n = 5 mice/group. (e) CV/Luxol fast blue staining was performed on post-ICH day 3, and lesion volume (corrected for brain swilling) was calculated. n = 5 mice/group. (f) The percentage of brain swelling was measured at three days after ICH. n = 5 mice/group. (g) The percentage of brain water content was measured at three days after ICH. n = 6 mice/group. (h) Field selection for ICH brain. * p
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    Image Search Results


    Knockdown of STAG2 or STAG3 decreases BRAFi sensitivity in BRAF mutant melanoma cells ( a ) Viability of A375 cells after treatment with varying concentrations of dabrafenib for 3 d. Experiment was performed 3 times. Data are mean ± s.e.m. ( b ) A375 cells were treated with dabrafenib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( c ) Viability of SKMEL28 cells after treatment with varying concentrations of vemurafenib for 3 d. Experiment was performed 3 times. Data are mean ± s.e.m. ( d ) SKMEL28 cells were treated with vemurafenib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( e ) SKMEL30 cells were treated with trametinib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( f ) SKMEL30 cells were treated with trametinib as indicated in clonogenic growth assays. Experiment was performed 3 times. Scale bar: 5 mm. ( g ) Viability of A375 cells after treatment with varying concentrations of dabrafenib for 3 d. Experiment was performed 3 times. Data are mean ± s.e.m. ( h ) A375 cells were treated with vemurafenib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( i ) WM902-BR cells stably expressing control vector, FLAG-tagged wild-type STAG2 (WT), Lys1083* (K*) or Asp193Asn (DN) mutants were treated with 3 μM vemurafenib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( j ) WM902-BR cells were used in soft agar assays in the presence or absence of 3 μM vemurafenib. Experiment was performed 3 times. Scale bar: 5 mm.

    Journal: Nature medicine

    Article Title: Loss of cohesin complex components STAG2 or STAG3 confers resistance to BRAF inhibition in melanoma

    doi: 10.1038/nm.4155

    Figure Lengend Snippet: Knockdown of STAG2 or STAG3 decreases BRAFi sensitivity in BRAF mutant melanoma cells ( a ) Viability of A375 cells after treatment with varying concentrations of dabrafenib for 3 d. Experiment was performed 3 times. Data are mean ± s.e.m. ( b ) A375 cells were treated with dabrafenib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( c ) Viability of SKMEL28 cells after treatment with varying concentrations of vemurafenib for 3 d. Experiment was performed 3 times. Data are mean ± s.e.m. ( d ) SKMEL28 cells were treated with vemurafenib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( e ) SKMEL30 cells were treated with trametinib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( f ) SKMEL30 cells were treated with trametinib as indicated in clonogenic growth assays. Experiment was performed 3 times. Scale bar: 5 mm. ( g ) Viability of A375 cells after treatment with varying concentrations of dabrafenib for 3 d. Experiment was performed 3 times. Data are mean ± s.e.m. ( h ) A375 cells were treated with vemurafenib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( i ) WM902-BR cells stably expressing control vector, FLAG-tagged wild-type STAG2 (WT), Lys1083* (K*) or Asp193Asn (DN) mutants were treated with 3 μM vemurafenib for 2 h. Cell lysates were used for western blotting with indicated antibodies. Experiment was performed 3 times. ( j ) WM902-BR cells were used in soft agar assays in the presence or absence of 3 μM vemurafenib. Experiment was performed 3 times. Scale bar: 5 mm.

    Article Snippet: A375 and SKMEL28 were purchased from ATCC.

    Techniques: Mutagenesis, Western Blot, Stable Transfection, Expressing, Plasmid Preparation

    Sodium hypochlorite treatment of glycoproteins to release N-glycans. a) Chemical scheme. b) TLC analysis of ovalbumin before and after bleach treatment. c) MALDI-MS profiles of N-glycans released from several glycoproteins by bleach treatment. d) MALDI-MS profiles of permethylated glycans released from fetuin by PNGase F digestion (top) or bleach treatment (bottom). e) Normal phase (NP) HPLC profiles of AEAB conjugated N-glycans released from fetuin by PNGase F digestion (top) or bleach treatment (bottom). Peaks at 10-17 min were nonglycan peaks presumably generated from oxidized peptide fragments. f) MALDI-MS profiles of human plasma permethylated N-glycans released by PNGase F digestion (top) and bleach (bottom). g) MALDI-MS profiles of N-glycans released by NaClO from egg white (top) and egg yolk (bottom). H: Hexose; N: N-acetylhexosamine; F: fucose; S: sialic acid. The Y-axis scales are not calibrated to reflect quantitative comparison.

    Journal: Nature methods

    Article Title: Oxidative Release of Natural Glycans for Functional Glycomics

    doi: 10.1038/nmeth.3861

    Figure Lengend Snippet: Sodium hypochlorite treatment of glycoproteins to release N-glycans. a) Chemical scheme. b) TLC analysis of ovalbumin before and after bleach treatment. c) MALDI-MS profiles of N-glycans released from several glycoproteins by bleach treatment. d) MALDI-MS profiles of permethylated glycans released from fetuin by PNGase F digestion (top) or bleach treatment (bottom). e) Normal phase (NP) HPLC profiles of AEAB conjugated N-glycans released from fetuin by PNGase F digestion (top) or bleach treatment (bottom). Peaks at 10-17 min were nonglycan peaks presumably generated from oxidized peptide fragments. f) MALDI-MS profiles of human plasma permethylated N-glycans released by PNGase F digestion (top) and bleach (bottom). g) MALDI-MS profiles of N-glycans released by NaClO from egg white (top) and egg yolk (bottom). H: Hexose; N: N-acetylhexosamine; F: fucose; S: sialic acid. The Y-axis scales are not calibrated to reflect quantitative comparison.

    Article Snippet: Sodium hypochlorite solutions are from Clorox (6.15% NaClO), Pure Bright (6% NaClO), Up & up (8.25% NaClO) or Sigma-Aldrich (5% chlorine) and prepared freshly by addition of water.

    Techniques: Thin Layer Chromatography, Mass Spectrometry, Normal Phase Liquid Chromatography, Generated

    Sodium hypochlorite treatment of glycoproteins to release O-glycans. a) Chemical scheme of O-glycan release and labeling. b) TLC analysis of porcine stomach mucin before and after NaClO treatment. c) MALDI-MS profile of O-glycans released from porcine stomach mucin. d) MALDI-MS profile of glycans from porcine stomach mucin and fetuin after NaClO treatment and permethylation. e) MALDI-MS profile and f) HPLC profile of O-glycans released from porcine stomach mucin by NaClO and labeled with monoFmoc-ethylenediamine. Fluorescence: Ex 254 xnm/Em 340 nm.

    Journal: Nature methods

    Article Title: Oxidative Release of Natural Glycans for Functional Glycomics

    doi: 10.1038/nmeth.3861

    Figure Lengend Snippet: Sodium hypochlorite treatment of glycoproteins to release O-glycans. a) Chemical scheme of O-glycan release and labeling. b) TLC analysis of porcine stomach mucin before and after NaClO treatment. c) MALDI-MS profile of O-glycans released from porcine stomach mucin. d) MALDI-MS profile of glycans from porcine stomach mucin and fetuin after NaClO treatment and permethylation. e) MALDI-MS profile and f) HPLC profile of O-glycans released from porcine stomach mucin by NaClO and labeled with monoFmoc-ethylenediamine. Fluorescence: Ex 254 xnm/Em 340 nm.

    Article Snippet: Sodium hypochlorite solutions are from Clorox (6.15% NaClO), Pure Bright (6% NaClO), Up & up (8.25% NaClO) or Sigma-Aldrich (5% chlorine) and prepared freshly by addition of water.

    Techniques: Labeling, Thin Layer Chromatography, Mass Spectrometry, High Performance Liquid Chromatography, Fluorescence

    mRNA expression patterns of meristem and flower development genes revealed by RNA FISH. The mRNAs (magenta) were hybridized with digoxigenin-labeled antisense RNA probes and detected by TSA-Cy5. Cell walls were stained by Calcofluor White and are shown in blue. Left images show RNA FISH signals in a Fire format. Meristems and flowers are outlined by white dashed lines. A, Schematic representation of the Arabidopsis inflorescence shoot apex. CZ, Central zone; PZ, peripheral zone; RM, rib meristem. B to F, H, K, L, and N, The distribution of WUS mRNA in the shoot apex (B), CLV3 (C), STM (D), LFY (E), AP1 (F), AP2 (H), CUC2 (K), FIL (L), and REV (N). I, AP3 mRNA expression in a stage 3 flower. M, FIL mRNA expression in a stage 6 flower. ca, Carpel; pe, petal; se, sepal; st, stamen. G and J, Expression patterns of AP1 translational fusion reporter gAP1 : GFP ) in the shoot apex and AP3 transcriptional reporter pAP3 :: 2BFP for serial sections. Bars = 25 μm.

    Journal: Plant Physiology

    Article Title: Visualization of Protein Coding, Long Noncoding, and Nuclear RNAs by Fluorescence in Situ Hybridization in Sections of Shoot Apical Meristems and Developing Flowers

    doi: 10.1104/pp.19.00980

    Figure Lengend Snippet: mRNA expression patterns of meristem and flower development genes revealed by RNA FISH. The mRNAs (magenta) were hybridized with digoxigenin-labeled antisense RNA probes and detected by TSA-Cy5. Cell walls were stained by Calcofluor White and are shown in blue. Left images show RNA FISH signals in a Fire format. Meristems and flowers are outlined by white dashed lines. A, Schematic representation of the Arabidopsis inflorescence shoot apex. CZ, Central zone; PZ, peripheral zone; RM, rib meristem. B to F, H, K, L, and N, The distribution of WUS mRNA in the shoot apex (B), CLV3 (C), STM (D), LFY (E), AP1 (F), AP2 (H), CUC2 (K), FIL (L), and REV (N). I, AP3 mRNA expression in a stage 3 flower. M, FIL mRNA expression in a stage 6 flower. ca, Carpel; pe, petal; se, sepal; st, stamen. G and J, Expression patterns of AP1 translational fusion reporter gAP1 : GFP ) in the shoot apex and AP3 transcriptional reporter pAP3 :: 2BFP for serial sections. Bars = 25 μm.

    Article Snippet: For cell wall staining, sectioned samples were treated with 0.1% (w/v) Calcofluor White (Fluorescent Brightener 28; Sigma) for 5 min. After briefly washing in TBS-T two times, samples were imaged with a Zeiss LSM700 confocal microscope with 405 nm excitation and 425 to 475 nm emission.

    Techniques: Expressing, Fluorescence In Situ Hybridization, Labeling, Staining

    Configuration of clamps and pseudoclamps and DAPI and Fluorescent Brightener 28 staining of nuclei and cell walls in the cotransformants. (A) Pseudoclamps in Hox2-1. (B) Clamps in Hox2-1. (C) Pseudoclamps with staining of nuclei and cell walls in Hox2-1.

    Journal: Eukaryotic Cell

    Article Title: A-Mating-Type Gene Expression Can Drive Clamp Formation in the Bipolar Mushroom Pholiota microspora ( Pholiota nameko) ▿

    doi: 10.1128/EC.00374-09

    Figure Lengend Snippet: Configuration of clamps and pseudoclamps and DAPI and Fluorescent Brightener 28 staining of nuclei and cell walls in the cotransformants. (A) Pseudoclamps in Hox2-1. (B) Clamps in Hox2-1. (C) Pseudoclamps with staining of nuclei and cell walls in Hox2-1.

    Article Snippet: The mycelium was put on the glass-containing agar, incubated for 5 to 7 days, and then stained for 20 min with a solution of 50 μg of DAPI (4′,6′-diamidino-2-phenylindole; Merck, Darmstadt, Germany)/ml, which stains nuclei, and 20 μg of Fluorescent Brightener 28 (Sigma-Aldrich, St. Louis, MO)/ml, which stains the cell wall.

    Techniques: Staining

    Congo-red and calcofluor binding assays on mono or two-species colonies formed by E. coli alone or with A. pleuropneumoniae . Congo-red assay evidenced curli-producing bacteria when E. coli was growing on congo-red-supplemented nutrient agar. Moreover, changes in cellulose production were detected by the calcofluor assay. The plus sings indicate differences on production observed in the colonies of one or two-species. The crosses indicate the qualitative production of cellulose seen in each of the biofilms (+ low, ++ medium, +++ medium high and ++++ high).

    Journal: Frontiers in Veterinary Science

    Article Title: Incorporation of Actinobacillus pleuropneumoniae in Preformed Biofilms by Escherichia coli Isolated From Drinking Water of Swine Farms

    doi: 10.3389/fvets.2018.00184

    Figure Lengend Snippet: Congo-red and calcofluor binding assays on mono or two-species colonies formed by E. coli alone or with A. pleuropneumoniae . Congo-red assay evidenced curli-producing bacteria when E. coli was growing on congo-red-supplemented nutrient agar. Moreover, changes in cellulose production were detected by the calcofluor assay. The plus sings indicate differences on production observed in the colonies of one or two-species. The crosses indicate the qualitative production of cellulose seen in each of the biofilms (+ low, ++ medium, +++ medium high and ++++ high).

    Article Snippet: For the assay, a 2 μl drop of bacterial culture was taken from liquid medium, and was placed on Luria-Bertani salt-free plates (LB; Difco Laboratories, Detroit, MI), containing 0.02% of Congo-red (Sigma®, C-6767) and 0.002% of Coomassie brilliant blue G (Sigma®, F3546-5G) for fimbriae detection, and containing 0.02% calcofluor (fluorescent brightener 28, Sigma-Aldrich® F-3543) dissolved in 1 mM HEPES for cellulose detection.

    Techniques: Binding Assay

    Transfection efficiency of nanoparticles on Neuro2a cells after 4 hours of incubation at 37°C and 5% CO 2 : a ) PEGylated-chitosan-TPP:siGLO nanoparticles; b ) chitosan-TPP:siGLO nanoparticles; c ) siGLO only (negative control); d ) chitosan-TPP nanoparticles (negative control); e ) cells only (negative control) Abbreviation: TPP, sodium tripolyphosphate.

    Journal: International Journal of Nanomedicine

    Article Title: A novel method for synthesizing PEGylated chitosan nanoparticles: strategy, preparation, and in vitro analysis

    doi: 10.2147/IJN.S17190

    Figure Lengend Snippet: Transfection efficiency of nanoparticles on Neuro2a cells after 4 hours of incubation at 37°C and 5% CO 2 : a ) PEGylated-chitosan-TPP:siGLO nanoparticles; b ) chitosan-TPP:siGLO nanoparticles; c ) siGLO only (negative control); d ) chitosan-TPP nanoparticles (negative control); e ) cells only (negative control) Abbreviation: TPP, sodium tripolyphosphate.

    Article Snippet: Poly (ethylene glycol) monomethyl ether (MW 5000), phthalic anhydride, sodium hydride (NaH), pyridine, hydrazine monohydrate, sodium tripolyphosphate (TPP), glacial acetic acid, agarose (low gelling temperature), and ethidium bromide (10 mg/mL) of analytical grade were obtained from Sigma-Aldrich (Oakville, ON, Canada).

    Techniques: Transfection, Incubation, Negative Control

    TEM images of a ) PEGylated chitosan polymer (mag. 57,000×); b ) chitosan–TPP nanoparticle (mag. 22,000×); c ) PEGylated chitosan–TPP nanoparticle (mag. 57,000×) and d ), PEGy lated chitosan-TPP nanoparticle (mag. 135000x). Abbreviation: TPP, sodium tripolyphosphate.

    Journal: International Journal of Nanomedicine

    Article Title: A novel method for synthesizing PEGylated chitosan nanoparticles: strategy, preparation, and in vitro analysis

    doi: 10.2147/IJN.S17190

    Figure Lengend Snippet: TEM images of a ) PEGylated chitosan polymer (mag. 57,000×); b ) chitosan–TPP nanoparticle (mag. 22,000×); c ) PEGylated chitosan–TPP nanoparticle (mag. 57,000×) and d ), PEGy lated chitosan-TPP nanoparticle (mag. 135000x). Abbreviation: TPP, sodium tripolyphosphate.

    Article Snippet: Poly (ethylene glycol) monomethyl ether (MW 5000), phthalic anhydride, sodium hydride (NaH), pyridine, hydrazine monohydrate, sodium tripolyphosphate (TPP), glacial acetic acid, agarose (low gelling temperature), and ethidium bromide (10 mg/mL) of analytical grade were obtained from Sigma-Aldrich (Oakville, ON, Canada).

    Techniques: Transmission Electron Microscopy

    A schematic diagram for the fabrication of phthalocyanine encapsulated chitosan/TPP nanoparticles (FNP) from chitosan, tripolyphosphate (TPP), and phthalocyanine-4,4′,4″,4‴-tetrasulfonic acid (FePC).

    Journal: Pharmaceutics

    Article Title: Sequential Photodynamic Therapy with Phthalocyanine Encapsulated Chitosan-Tripolyphosphate Nanoparticles and Flucytosine Treatment against Candida tropicalis

    doi: 10.3390/pharmaceutics11010016

    Figure Lengend Snippet: A schematic diagram for the fabrication of phthalocyanine encapsulated chitosan/TPP nanoparticles (FNP) from chitosan, tripolyphosphate (TPP), and phthalocyanine-4,4′,4″,4‴-tetrasulfonic acid (FePC).

    Article Snippet: Sodium tripolyphosphate (Sigma-Aldrich, St. Louis, MO, US) was dissolved in H2 O to reach a concentration of 1% (w /v ), and then filtered with a 0.22 μm membrane.

    Techniques:

    Representative electropherogram of total RNA extracted from pancreatic cancer obtained by endoscopic ultrasound-guided fine-needle aspiration biopsy . The ratio of 28S to 18S of ribosomal RNA indicates good quality of total RNA.

    Journal: Journal of Experimental & Clinical Cancer Research : CR

    Article Title: Gemcitabine sensitivity-related mRNA expression in endoscopic ultrasound-guided fine-needle aspiration biopsy of unresectable pancreatic cancer

    doi: 10.1186/1756-9966-28-83

    Figure Lengend Snippet: Representative electropherogram of total RNA extracted from pancreatic cancer obtained by endoscopic ultrasound-guided fine-needle aspiration biopsy . The ratio of 28S to 18S of ribosomal RNA indicates good quality of total RNA.

    Article Snippet: RNAs were examined for qualities by confirming the 28S and 18S ribosomal bands with an Agilent 2100 Bioanalyzer (Agilent Technologies, Inc., Santa Clara, CA, USA).

    Techniques:

    Cmpd 28 treatment improves insulin-induced Akt phosphorylation and lowers the expression of gluconeogenic genes in obese mice. Ten-week-old ob/ob mice were injected with 0.2 mg/kg body wt i.p. cmpd 28 or vehicle (control [Con]) each day for 3 weeks ( n = 5 mice per group). A : The mice were fasted for 6 h and then injected with 2 IU/kg body wt insulin (ins) through the portal vein. After 3 min, livers were harvested, and total liver extracts were assayed for p-Akt, Akt, Trb3, and β-actin by immunoblot. Densitometric quantification of the immunoblot data is shown in the graph (* P

    Journal: Diabetes

    Article Title: Treatment of Obese Insulin-Resistant Mice With an Allosteric MAPKAPK2/3 Inhibitor Lowers Blood Glucose and Improves Insulin Sensitivity

    doi: 10.2337/db14-1945

    Figure Lengend Snippet: Cmpd 28 treatment improves insulin-induced Akt phosphorylation and lowers the expression of gluconeogenic genes in obese mice. Ten-week-old ob/ob mice were injected with 0.2 mg/kg body wt i.p. cmpd 28 or vehicle (control [Con]) each day for 3 weeks ( n = 5 mice per group). A : The mice were fasted for 6 h and then injected with 2 IU/kg body wt insulin (ins) through the portal vein. After 3 min, livers were harvested, and total liver extracts were assayed for p-Akt, Akt, Trb3, and β-actin by immunoblot. Densitometric quantification of the immunoblot data is shown in the graph (* P

    Article Snippet: Most importantly, cmpd 28 had no effect on CaMKII at the doses where it can efficiently inhibit forskolin-induced p-hsp25 ( Supplementary Fig. 2C ).

    Techniques: Expressing, Mouse Assay, Injection

    Cmpd 28 treatment improves glucose homeostasis in ob/ob mice. Ten-week-old ob/ob mice were injected with 0.2 mg/kg body wt i.p. cmpd 28 or vehicle (control [Con]) each day for 3 weeks ( n = 5 mice per group). The mice were then assayed for hepatic p-hsp25, total hsp25, and β-actin by immunoblot ( A ). p, phosphorylated. B and C : Ten-week-old ob/ob mice were injected with 0.2 mg/kg body wt i.p. cmpd 28 or vehicle (Con) each day for 2 weeks ( n = 6 mice per group). Blood glucose and plasma insulin levels after a 6-h fast at day 3 are shown (* P

    Journal: Diabetes

    Article Title: Treatment of Obese Insulin-Resistant Mice With an Allosteric MAPKAPK2/3 Inhibitor Lowers Blood Glucose and Improves Insulin Sensitivity

    doi: 10.2337/db14-1945

    Figure Lengend Snippet: Cmpd 28 treatment improves glucose homeostasis in ob/ob mice. Ten-week-old ob/ob mice were injected with 0.2 mg/kg body wt i.p. cmpd 28 or vehicle (control [Con]) each day for 3 weeks ( n = 5 mice per group). The mice were then assayed for hepatic p-hsp25, total hsp25, and β-actin by immunoblot ( A ). p, phosphorylated. B and C : Ten-week-old ob/ob mice were injected with 0.2 mg/kg body wt i.p. cmpd 28 or vehicle (Con) each day for 2 weeks ( n = 6 mice per group). Blood glucose and plasma insulin levels after a 6-h fast at day 3 are shown (* P

    Article Snippet: Most importantly, cmpd 28 had no effect on CaMKII at the doses where it can efficiently inhibit forskolin-induced p-hsp25 ( Supplementary Fig. 2C ).

    Techniques: Mouse Assay, Injection

    Cmpd 28 treatment improves glucose homeostasis in DIO mice. DIO mice (17 weeks old) were injected with 0.2 mg/kg body wt i.p. cmpd 28 or vehicle (control [Con]) each day for 3 weeks ( n = 6 mice per group). The mice were then assayed for fasting blood glucose ( A ), fed blood glucose ( B ), fasting plasma insulin ( C ), and fed plasma insulin ( D ), and body weight was determined ( E ) (* P

    Journal: Diabetes

    Article Title: Treatment of Obese Insulin-Resistant Mice With an Allosteric MAPKAPK2/3 Inhibitor Lowers Blood Glucose and Improves Insulin Sensitivity

    doi: 10.2337/db14-1945

    Figure Lengend Snippet: Cmpd 28 treatment improves glucose homeostasis in DIO mice. DIO mice (17 weeks old) were injected with 0.2 mg/kg body wt i.p. cmpd 28 or vehicle (control [Con]) each day for 3 weeks ( n = 6 mice per group). The mice were then assayed for fasting blood glucose ( A ), fed blood glucose ( B ), fasting plasma insulin ( C ), and fed plasma insulin ( D ), and body weight was determined ( E ) (* P

    Article Snippet: Most importantly, cmpd 28 had no effect on CaMKII at the doses where it can efficiently inhibit forskolin-induced p-hsp25 ( Supplementary Fig. 2C ).

    Techniques: Mouse Assay, Injection

    The metabolic improvement by cmpd 28 and dominant-negative MK2 in obese mice is not additive. Ten-week-old ob/ob mice were transduced with adeno-LacZ or adeno–T222A-MK2 (dominant negative), which inhibits hepatic MK2. Mice were then treated with 0.2 mg/kg body wt i.p. cmpd 28 or vehicle (Veh) for 7 days ( n = 5 mice per group). After a 6-h fast, the mice were assayed for fasting blood glucose ( A ) and plasma insulin ( B ) (* P

    Journal: Diabetes

    Article Title: Treatment of Obese Insulin-Resistant Mice With an Allosteric MAPKAPK2/3 Inhibitor Lowers Blood Glucose and Improves Insulin Sensitivity

    doi: 10.2337/db14-1945

    Figure Lengend Snippet: The metabolic improvement by cmpd 28 and dominant-negative MK2 in obese mice is not additive. Ten-week-old ob/ob mice were transduced with adeno-LacZ or adeno–T222A-MK2 (dominant negative), which inhibits hepatic MK2. Mice were then treated with 0.2 mg/kg body wt i.p. cmpd 28 or vehicle (Veh) for 7 days ( n = 5 mice per group). After a 6-h fast, the mice were assayed for fasting blood glucose ( A ) and plasma insulin ( B ) (* P

    Article Snippet: Most importantly, cmpd 28 had no effect on CaMKII at the doses where it can efficiently inhibit forskolin-induced p-hsp25 ( Supplementary Fig. 2C ).

    Techniques: Dominant Negative Mutation, Mouse Assay, Transduction

    The metabolic improvement by cmpd 28 and metformin in obese mice is additive. Ten-week-old ob/ob mice were treated intraperitoneally with vehicle (control [Con]), 200 mg/kg body wt metformin (Met), 0.2 mg/kg body wt cmpd 28, or both compounds for 14 days ( n = 5 mice per group). The mice were assayed for fasting blood glucose ( A ), plasma insulin ( B ), hepatic G6pc mRNA expression levels ( C ), and body weight ( D ) (* P

    Journal: Diabetes

    Article Title: Treatment of Obese Insulin-Resistant Mice With an Allosteric MAPKAPK2/3 Inhibitor Lowers Blood Glucose and Improves Insulin Sensitivity

    doi: 10.2337/db14-1945

    Figure Lengend Snippet: The metabolic improvement by cmpd 28 and metformin in obese mice is additive. Ten-week-old ob/ob mice were treated intraperitoneally with vehicle (control [Con]), 200 mg/kg body wt metformin (Met), 0.2 mg/kg body wt cmpd 28, or both compounds for 14 days ( n = 5 mice per group). The mice were assayed for fasting blood glucose ( A ), plasma insulin ( B ), hepatic G6pc mRNA expression levels ( C ), and body weight ( D ) (* P

    Article Snippet: Most importantly, cmpd 28 had no effect on CaMKII at the doses where it can efficiently inhibit forskolin-induced p-hsp25 ( Supplementary Fig. 2C ).

    Techniques: Mouse Assay, Expressing

    Cmpd 28 is an effective inhibitor of MK2/3 activity in liver cells and inhibits forskolin-induced hsp25 phosphorylation and G6pc expression. A : Primary HCs from WT mice were pretreated with either vehicle (Control [Con]) or 500 nm cmpd 28 for 1 h followed by incubation with either BSA control or 0.3 mmol/L palmitate (palm) for 6 h. Lysates were probed for p-hsp25, hsp25, and β-actin by immunoblot. Densitometric quantification of the immunoblot data are shown in the graph (differing symbols indicate P

    Journal: Diabetes

    Article Title: Treatment of Obese Insulin-Resistant Mice With an Allosteric MAPKAPK2/3 Inhibitor Lowers Blood Glucose and Improves Insulin Sensitivity

    doi: 10.2337/db14-1945

    Figure Lengend Snippet: Cmpd 28 is an effective inhibitor of MK2/3 activity in liver cells and inhibits forskolin-induced hsp25 phosphorylation and G6pc expression. A : Primary HCs from WT mice were pretreated with either vehicle (Control [Con]) or 500 nm cmpd 28 for 1 h followed by incubation with either BSA control or 0.3 mmol/L palmitate (palm) for 6 h. Lysates were probed for p-hsp25, hsp25, and β-actin by immunoblot. Densitometric quantification of the immunoblot data are shown in the graph (differing symbols indicate P

    Article Snippet: Most importantly, cmpd 28 had no effect on CaMKII at the doses where it can efficiently inhibit forskolin-induced p-hsp25 ( Supplementary Fig. 2C ).

    Techniques: Activity Assay, Expressing, Mouse Assay, Incubation

    DIMT1L, WBSCR22, and TRMT112 are required for distinct pre-rRNA processing steps, and the pre-rRNA processing defects are conserved in different cell types and do not depend on p53. WI-38, RKO, and HCT116 cells were treated for 3 d with a specific siRNA targeting DIMT1L, WBSCR22, or TRMT112. Paired HCT116 cell lines expressing p53 or not were used (+/+ and –/–). Total RNA was extracted, resolved on denaturing agarose gel, transferred to a nylon membrane, and hybridized with probes. The probes used were as follows: (I, II) LD1844, (III) LD1827, and (IV) LD1828. The detected pre-rRNA species are indicated to the right and schematized. (V, VI) The mature 28S and 18S rRNAs stained with ethidium bromide. For each sample, the 28S/18S ratio was calculated from Agilent bioanalyzer electropherograms. All RNA species were quantified with a Phosphorimager normalized with respect to the nontargeting control (SCR), and their abundances represented as a heatmap using the color code indicated to the right. The sequences of the siRNAs used are listed in Supplemental Table S3 (WBSCR22#1, TRMT112#2, and DIMT1L#2). Note that in the heatmap for RKO cells, the signal for lane 8 was corrected for loading.

    Journal: Molecular Biology of the Cell

    Article Title: The human 18S rRNA base methyltransferases DIMT1L and WBSCR22-TRMT112 but not rRNA modification are required for ribosome biogenesis

    doi: 10.1091/mbc.E15-02-0073

    Figure Lengend Snippet: DIMT1L, WBSCR22, and TRMT112 are required for distinct pre-rRNA processing steps, and the pre-rRNA processing defects are conserved in different cell types and do not depend on p53. WI-38, RKO, and HCT116 cells were treated for 3 d with a specific siRNA targeting DIMT1L, WBSCR22, or TRMT112. Paired HCT116 cell lines expressing p53 or not were used (+/+ and –/–). Total RNA was extracted, resolved on denaturing agarose gel, transferred to a nylon membrane, and hybridized with probes. The probes used were as follows: (I, II) LD1844, (III) LD1827, and (IV) LD1828. The detected pre-rRNA species are indicated to the right and schematized. (V, VI) The mature 28S and 18S rRNAs stained with ethidium bromide. For each sample, the 28S/18S ratio was calculated from Agilent bioanalyzer electropherograms. All RNA species were quantified with a Phosphorimager normalized with respect to the nontargeting control (SCR), and their abundances represented as a heatmap using the color code indicated to the right. The sequences of the siRNAs used are listed in Supplemental Table S3 (WBSCR22#1, TRMT112#2, and DIMT1L#2). Note that in the heatmap for RKO cells, the signal for lane 8 was corrected for loading.

    Article Snippet: The 28S/18S ratio was calculated with the Agilent RNA 6000 nano kit (5067-1511) on a BioAnalyzer 2100 (Agilent).

    Techniques: Expressing, Agarose Gel Electrophoresis, Staining

    Dynamic analysis of pre-rRNA processing defects in cells depleted of DIMT1L, WBSCR22, or TRMT112. (I) HeLa cells were depleted of WBSCR22, TRMT112, or DIMT1L for 3 d by means of a specific siRNA, pulse labeled for 30 min with l -(methyl- 3 H)-methionine, chased with an excess of cold methionine, and collected at different time points over a 4-h period. Total RNA was extracted, resolved on an agarose gel, transferred to a GeneScreen membrane, and then exposed by fluorography. (II, III) The ethidium bromide–stained mature 28S and 18S rRNAs. The sequences of the siRNAs used are listed in Supplemental Table S3 (WBSCR22#1, TRMT112#2, and DIMT1L#2).

    Journal: Molecular Biology of the Cell

    Article Title: The human 18S rRNA base methyltransferases DIMT1L and WBSCR22-TRMT112 but not rRNA modification are required for ribosome biogenesis

    doi: 10.1091/mbc.E15-02-0073

    Figure Lengend Snippet: Dynamic analysis of pre-rRNA processing defects in cells depleted of DIMT1L, WBSCR22, or TRMT112. (I) HeLa cells were depleted of WBSCR22, TRMT112, or DIMT1L for 3 d by means of a specific siRNA, pulse labeled for 30 min with l -(methyl- 3 H)-methionine, chased with an excess of cold methionine, and collected at different time points over a 4-h period. Total RNA was extracted, resolved on an agarose gel, transferred to a GeneScreen membrane, and then exposed by fluorography. (II, III) The ethidium bromide–stained mature 28S and 18S rRNAs. The sequences of the siRNAs used are listed in Supplemental Table S3 (WBSCR22#1, TRMT112#2, and DIMT1L#2).

    Article Snippet: The 28S/18S ratio was calculated with the Agilent RNA 6000 nano kit (5067-1511) on a BioAnalyzer 2100 (Agilent).

    Techniques: Labeling, Agarose Gel Electrophoresis, Staining

    Effect of treatment with NaHS on protein levels of Cx40 and Cx43 in peripheral blood lymphocytes of SHR. We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, peripheral blood lymphocytes were harvested and examined by Western blot for the expression levels of Cx40 ( A ) and Cx43 ( B ). After densitometric analysis, the data were expressed as ratios of Cxs to β-actin. The data represent the mean ±SEM of 3 experiments ( n =15 animals in each group). ** P

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Hydrogen Sulfide Attenuates Hypertensive Inflammation via Regulating Connexin Expression in Spontaneously Hypertensive Rats

    doi: 10.12659/MSM.908761

    Figure Lengend Snippet: Effect of treatment with NaHS on protein levels of Cx40 and Cx43 in peripheral blood lymphocytes of SHR. We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, peripheral blood lymphocytes were harvested and examined by Western blot for the expression levels of Cx40 ( A ) and Cx43 ( B ). After densitometric analysis, the data were expressed as ratios of Cxs to β-actin. The data represent the mean ±SEM of 3 experiments ( n =15 animals in each group). ** P

    Article Snippet: SHR in the SHR+NaHS group were intraperitoneally injected with 56 μmol/kg−1 ·day−1 of NaHS (NaHS solution was freshly prepared in normal saline) (Cat. No. 161527; Sigma Aldrich, St. Louis, MO, USA) at the same time daily for 9 weeks.

    Techniques: Injection, Western Blot, Expressing

    Effect of long-term NaHS treatment on surface expressions of Cx40 and Cx43 in different T lymphocyte subtypes of SHR. A, Representative flow cytometry plots are presented for Cx40 and Cx43 expression levels on gated single-positive CD4 + T lymphocytes or CD8 + T lymphocyte populations in the peripheral blood from 15 SHR and 15 WKY rats. Fresh, resting PBMCs from SHR and WKY rats underwent surface staining with antibodies against CD3, CD4, and CD8 molecules. After surface staining, the cells were fixed, permeabilized, and stained with unlabeled anti-Cx40 or anti-Cx43 plus FITC-labeled secondary antibodies. Based on the CD4 + or CD8 + gate, the cells were further gated based on Cx40 and Cx43 expression levels, and the frequency of CD4 + or CD8 + T cells expressing Cx40 and Cx43 was determined. B, Bar graph shown are the percentage of CD4 + or CD8 + T cell population expressing Cx40 and Cx43. Both Cx40 and Cx43 expression levels are significantly increased in CD4 + or CD8 + T cells of SHR compared with those of WKY rats. Long-term NaHS treatment inhibited the expressions of Cx40 and Cx43 in CD4 + and CD8 + T cells from the peripheral blood of SHR, and their expressions in SHR returned to the levels seen in WKY rats. Values are mean ± SEM. * P

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Hydrogen Sulfide Attenuates Hypertensive Inflammation via Regulating Connexin Expression in Spontaneously Hypertensive Rats

    doi: 10.12659/MSM.908761

    Figure Lengend Snippet: Effect of long-term NaHS treatment on surface expressions of Cx40 and Cx43 in different T lymphocyte subtypes of SHR. A, Representative flow cytometry plots are presented for Cx40 and Cx43 expression levels on gated single-positive CD4 + T lymphocytes or CD8 + T lymphocyte populations in the peripheral blood from 15 SHR and 15 WKY rats. Fresh, resting PBMCs from SHR and WKY rats underwent surface staining with antibodies against CD3, CD4, and CD8 molecules. After surface staining, the cells were fixed, permeabilized, and stained with unlabeled anti-Cx40 or anti-Cx43 plus FITC-labeled secondary antibodies. Based on the CD4 + or CD8 + gate, the cells were further gated based on Cx40 and Cx43 expression levels, and the frequency of CD4 + or CD8 + T cells expressing Cx40 and Cx43 was determined. B, Bar graph shown are the percentage of CD4 + or CD8 + T cell population expressing Cx40 and Cx43. Both Cx40 and Cx43 expression levels are significantly increased in CD4 + or CD8 + T cells of SHR compared with those of WKY rats. Long-term NaHS treatment inhibited the expressions of Cx40 and Cx43 in CD4 + and CD8 + T cells from the peripheral blood of SHR, and their expressions in SHR returned to the levels seen in WKY rats. Values are mean ± SEM. * P

    Article Snippet: SHR in the SHR+NaHS group were intraperitoneally injected with 56 μmol/kg−1 ·day−1 of NaHS (NaHS solution was freshly prepared in normal saline) (Cat. No. 161527; Sigma Aldrich, St. Louis, MO, USA) at the same time daily for 9 weeks.

    Techniques: Flow Cytometry, Cytometry, Expressing, Staining, Labeling

    Effect of long-term NaHS treatment on the production of pro-inflammatory and anti-inflammatory cytokines in serum of SHR. We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, serum level of IL-2 (A), IL-6 (B) and IL-10 (C) were determined as described in “Materials and methods”. Data represented as total amount of cytokine produced in pg/ml in peripheral blood; the results shown are the mean ±SEM; * P

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Hydrogen Sulfide Attenuates Hypertensive Inflammation via Regulating Connexin Expression in Spontaneously Hypertensive Rats

    doi: 10.12659/MSM.908761

    Figure Lengend Snippet: Effect of long-term NaHS treatment on the production of pro-inflammatory and anti-inflammatory cytokines in serum of SHR. We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, serum level of IL-2 (A), IL-6 (B) and IL-10 (C) were determined as described in “Materials and methods”. Data represented as total amount of cytokine produced in pg/ml in peripheral blood; the results shown are the mean ±SEM; * P

    Article Snippet: SHR in the SHR+NaHS group were intraperitoneally injected with 56 μmol/kg−1 ·day−1 of NaHS (NaHS solution was freshly prepared in normal saline) (Cat. No. 161527; Sigma Aldrich, St. Louis, MO, USA) at the same time daily for 9 weeks.

    Techniques: Injection, Produced

    The inhibitory effect of NaHS on the contractile response in basilar arteries (BA) induced by 40 mM KCl. We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, BA were isolated and their contractile response were determined as described in “Materials and methods”. Contraction of BA in response to KCl was greater in SHR than in WKY rats (* P

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Hydrogen Sulfide Attenuates Hypertensive Inflammation via Regulating Connexin Expression in Spontaneously Hypertensive Rats

    doi: 10.12659/MSM.908761

    Figure Lengend Snippet: The inhibitory effect of NaHS on the contractile response in basilar arteries (BA) induced by 40 mM KCl. We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, BA were isolated and their contractile response were determined as described in “Materials and methods”. Contraction of BA in response to KCl was greater in SHR than in WKY rats (* P

    Article Snippet: SHR in the SHR+NaHS group were intraperitoneally injected with 56 μmol/kg−1 ·day−1 of NaHS (NaHS solution was freshly prepared in normal saline) (Cat. No. 161527; Sigma Aldrich, St. Louis, MO, USA) at the same time daily for 9 weeks.

    Techniques: Injection, Isolation

    Long-term NaHS treatment reversed the changes of different T lymphocyte subtypes in SHR. We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, PBMCs were harvested and subjected to flow cytometry analysis. Reported dot plots are generated gating on living PBMCs in the scatter (FSC vs. SSC) dot plot (not shown). A, Representative flow cytometry analysis showing percentages of circulating T lymphocytes subtypes in the peripheral blood of 15 SHR and 15 age-matched WKY rats. B, Bar graph shown are proportion of CD3 + , CD4 + , CD8 + and CD25 + T cells expressing CD4 + as well as the ratio of CD4 + /CD8 + in the peripheral blood of SHR and WKY rats. The vertical axis represents the frequency of various T lymphocyte subtypes. Quantitative analysis of the mean percentage of cells ±SEM. ** P

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Hydrogen Sulfide Attenuates Hypertensive Inflammation via Regulating Connexin Expression in Spontaneously Hypertensive Rats

    doi: 10.12659/MSM.908761

    Figure Lengend Snippet: Long-term NaHS treatment reversed the changes of different T lymphocyte subtypes in SHR. We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, PBMCs were harvested and subjected to flow cytometry analysis. Reported dot plots are generated gating on living PBMCs in the scatter (FSC vs. SSC) dot plot (not shown). A, Representative flow cytometry analysis showing percentages of circulating T lymphocytes subtypes in the peripheral blood of 15 SHR and 15 age-matched WKY rats. B, Bar graph shown are proportion of CD3 + , CD4 + , CD8 + and CD25 + T cells expressing CD4 + as well as the ratio of CD4 + /CD8 + in the peripheral blood of SHR and WKY rats. The vertical axis represents the frequency of various T lymphocyte subtypes. Quantitative analysis of the mean percentage of cells ±SEM. ** P

    Article Snippet: SHR in the SHR+NaHS group were intraperitoneally injected with 56 μmol/kg−1 ·day−1 of NaHS (NaHS solution was freshly prepared in normal saline) (Cat. No. 161527; Sigma Aldrich, St. Louis, MO, USA) at the same time daily for 9 weeks.

    Techniques: Injection, Flow Cytometry, Cytometry, Generated, Expressing

    Effect of NaHS on spontaneous hypertension induced increase in blood pressure (BP) in SHR. We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, systolic blood pressure in WKY, SHR, and NaHS+SHR was measured. SHR had significantly increased blood pressure compared to WKY (** P

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Hydrogen Sulfide Attenuates Hypertensive Inflammation via Regulating Connexin Expression in Spontaneously Hypertensive Rats

    doi: 10.12659/MSM.908761

    Figure Lengend Snippet: Effect of NaHS on spontaneous hypertension induced increase in blood pressure (BP) in SHR. We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, systolic blood pressure in WKY, SHR, and NaHS+SHR was measured. SHR had significantly increased blood pressure compared to WKY (** P

    Article Snippet: SHR in the SHR+NaHS group were intraperitoneally injected with 56 μmol/kg−1 ·day−1 of NaHS (NaHS solution was freshly prepared in normal saline) (Cat. No. 161527; Sigma Aldrich, St. Louis, MO, USA) at the same time daily for 9 weeks.

    Techniques: Injection

    Long-term NaHS treatment alleviates vascular remodeling and infiltration of inflammatory cells in BA and kidney tissues of SHR. We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, kidneys and basal cerebral arteries were harvested and were stained by hematoxylin-eosin. A, cross-sections of BA stained with hematoxylin-eosin staining (magnification×200. scalar bar=4.5 μm). B, longitudinal sections of kidney tissues stained with hematoxylin-eosin staining (magnification×100, scalar bar=4.5 μm) ( n =15).

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Hydrogen Sulfide Attenuates Hypertensive Inflammation via Regulating Connexin Expression in Spontaneously Hypertensive Rats

    doi: 10.12659/MSM.908761

    Figure Lengend Snippet: Long-term NaHS treatment alleviates vascular remodeling and infiltration of inflammatory cells in BA and kidney tissues of SHR. We treated 9-week-old male SHR and WKY rats with NaHS (56 μmol/kg −1 ·day −1 , i.p.) or the same volume of normal saline and continued every day. At 9 weeks after NaHS injection, kidneys and basal cerebral arteries were harvested and were stained by hematoxylin-eosin. A, cross-sections of BA stained with hematoxylin-eosin staining (magnification×200. scalar bar=4.5 μm). B, longitudinal sections of kidney tissues stained with hematoxylin-eosin staining (magnification×100, scalar bar=4.5 μm) ( n =15).

    Article Snippet: SHR in the SHR+NaHS group were intraperitoneally injected with 56 μmol/kg−1 ·day−1 of NaHS (NaHS solution was freshly prepared in normal saline) (Cat. No. 161527; Sigma Aldrich, St. Louis, MO, USA) at the same time daily for 9 weeks.

    Techniques: Injection, Staining

    Prevalence of PD-L1 protein in FFPE Melanoma Specimens using PD-L1 IHC 28-8 pharmDx assay for detection (n=104 specimens). PD-L1 indicates programmed cell death 1-ligand 1; IHC, immunohistochemistry; FFPE indicates formalin-fixed paraffin-embedded.

    Journal: Applied Immunohistochemistry & Molecular Morphology

    Article Title: Development of a Diagnostic Programmed Cell Death 1-Ligand 1 Immunohistochemistry Assay for Nivolumab Therapy in Melanoma

    doi: 10.1097/PAI.0000000000000605

    Figure Lengend Snippet: Prevalence of PD-L1 protein in FFPE Melanoma Specimens using PD-L1 IHC 28-8 pharmDx assay for detection (n=104 specimens). PD-L1 indicates programmed cell death 1-ligand 1; IHC, immunohistochemistry; FFPE indicates formalin-fixed paraffin-embedded.

    Article Snippet: All instructions on how to run the assay are found in the PD-L1 IHC 28-8 pharmDx instructions for use (IFU).

    Techniques: Formalin-fixed Paraffin-Embedded, Immunohistochemistry

    Removal of N-linked glycosylation enhances anti-PD-L1 signal in human cancer cells in a variety of bioassays. (A and B) Immunofluorescence confocal microscopy of BT-549 (A) and A549 (B) cells processed with or without deglycosylation by PNGase F (5%) pretreatment stained with DAPI and an anti-PD-L1 antibody (Abcam, ab58810). Bar, 10 μm. Quantification is shown to the right. Data are representative of 3 independent experiments, randomly chosen in 3 different fields. (C and D) ELISA of Con A (C) and PD-L1 (clone 28-8 mAb) (D) levels in BT-549 cells processed with deglycosylation by increasing concentrations of PNGase F (1, 2, 5%) pretreatment for comparison with cells without deglycosylation (PNGase F; 0%). The intensity of Con A and PD-L1 was normalized to that without PNGase F pretreatment and set to 1. (E) ELISA of PD-L1 levels (clone 28-8 mAb) in lung cancer cells processed with deglycosylation by PNGase F (1%) pretreatment for comparison with cells without deglycosylation (0%). Negative control, secondary Ab only control. (F) Left: saturation binding assay of A549 cell lysates binding to anti-PD-L1 clone 28-8 mAb. Right: scatchard plot of cell number binding to anti-PD-L1 antibody transformed from the left. (G) Representative images (left) and quantification (right) of H-score of IHC staining for BLBC (BT-549, BT-20, and MDA-MB-231) and non-BLBC (MCF-7) cancer cell blocks processed with or without deglycosylation by PNGase F (5%) pretreatment. Bar, 50 μm. (H) Representative images (top) and quantification (bottom) of H-score of IHC staining for a panel of lung cancer cell blocks processed with or without deglycosylation by PNGase F (5%) pretreatment. Bar, 50 μm. (A–F) Results are presented as mean ± SD. *p

    Journal: Cancer cell

    Article Title: Removal of N-linked glycosylation enhances PD-L1 detection and predicts anti-PD-1/PD-L1 therapeutic efficacy

    doi: 10.1016/j.ccell.2019.06.008

    Figure Lengend Snippet: Removal of N-linked glycosylation enhances anti-PD-L1 signal in human cancer cells in a variety of bioassays. (A and B) Immunofluorescence confocal microscopy of BT-549 (A) and A549 (B) cells processed with or without deglycosylation by PNGase F (5%) pretreatment stained with DAPI and an anti-PD-L1 antibody (Abcam, ab58810). Bar, 10 μm. Quantification is shown to the right. Data are representative of 3 independent experiments, randomly chosen in 3 different fields. (C and D) ELISA of Con A (C) and PD-L1 (clone 28-8 mAb) (D) levels in BT-549 cells processed with deglycosylation by increasing concentrations of PNGase F (1, 2, 5%) pretreatment for comparison with cells without deglycosylation (PNGase F; 0%). The intensity of Con A and PD-L1 was normalized to that without PNGase F pretreatment and set to 1. (E) ELISA of PD-L1 levels (clone 28-8 mAb) in lung cancer cells processed with deglycosylation by PNGase F (1%) pretreatment for comparison with cells without deglycosylation (0%). Negative control, secondary Ab only control. (F) Left: saturation binding assay of A549 cell lysates binding to anti-PD-L1 clone 28-8 mAb. Right: scatchard plot of cell number binding to anti-PD-L1 antibody transformed from the left. (G) Representative images (left) and quantification (right) of H-score of IHC staining for BLBC (BT-549, BT-20, and MDA-MB-231) and non-BLBC (MCF-7) cancer cell blocks processed with or without deglycosylation by PNGase F (5%) pretreatment. Bar, 50 μm. (H) Representative images (top) and quantification (bottom) of H-score of IHC staining for a panel of lung cancer cell blocks processed with or without deglycosylation by PNGase F (5%) pretreatment. Bar, 50 μm. (A–F) Results are presented as mean ± SD. *p

    Article Snippet: After wiping off normal serum, PD-L1 primary antibodies (1:100; Abcam, ab205921, clone 28-8 mAb) were added to the sections in a humid chamber at 40 °C overnight, washed with PBS three times, and incubated with an anti-rabbit secondary antibody (1:200) for 1 hr in a humid chamber at room temperature.

    Techniques: Immunofluorescence, Confocal Microscopy, Staining, Enzyme-linked Immunosorbent Assay, Negative Control, Saturation Assay, Binding Assay, Transformation Assay, Immunohistochemistry, Multiple Displacement Amplification

    VK-28 attenuates ROS production, iron deposition, neuronal degeneration, lesion volume, and brain edema after ICH in vivo. Middle-aged male C57BL/6 mice underwent ICH by collagenase injection. DFX, VK-28, or vehicle (5% DMSO in saline) was administered i.p. at 6 h post-ICH and then daily until sacrifice. (a) HEt was injected i.p. 1 h before sacrifice. Representative images show HEt fluorescence intensity one day after ICH, and quantification shows the intensity as fold change of that in the vehicle group. Vehicle: n = 10, DFX: n = 5, VK-28: n = 5. (b) Perls’ staining and quantification of iron + cells at three days after ICH. n = 5 mice/group. (c) Fluoro-Jade B (FJB) was used to stain degenerating neurons at day 3. Representative images and quantification are shown. n = 5 mice/group. (d) Cresyl violet (CV) staining and quantification of surviving neurons. n = 5 mice/group. (e) CV/Luxol fast blue staining was performed on post-ICH day 3, and lesion volume (corrected for brain swilling) was calculated. n = 5 mice/group. (f) The percentage of brain swelling was measured at three days after ICH. n = 5 mice/group. (g) The percentage of brain water content was measured at three days after ICH. n = 6 mice/group. (h) Field selection for ICH brain. * p

    Journal: Journal of Cerebral Blood Flow & Metabolism

    Article Title: Neuroprotection of brain-permeable iron chelator VK-28 against intracerebral hemorrhage in mice

    doi: 10.1177/0271678X17709186

    Figure Lengend Snippet: VK-28 attenuates ROS production, iron deposition, neuronal degeneration, lesion volume, and brain edema after ICH in vivo. Middle-aged male C57BL/6 mice underwent ICH by collagenase injection. DFX, VK-28, or vehicle (5% DMSO in saline) was administered i.p. at 6 h post-ICH and then daily until sacrifice. (a) HEt was injected i.p. 1 h before sacrifice. Representative images show HEt fluorescence intensity one day after ICH, and quantification shows the intensity as fold change of that in the vehicle group. Vehicle: n = 10, DFX: n = 5, VK-28: n = 5. (b) Perls’ staining and quantification of iron + cells at three days after ICH. n = 5 mice/group. (c) Fluoro-Jade B (FJB) was used to stain degenerating neurons at day 3. Representative images and quantification are shown. n = 5 mice/group. (d) Cresyl violet (CV) staining and quantification of surviving neurons. n = 5 mice/group. (e) CV/Luxol fast blue staining was performed on post-ICH day 3, and lesion volume (corrected for brain swilling) was calculated. n = 5 mice/group. (f) The percentage of brain swelling was measured at three days after ICH. n = 5 mice/group. (g) The percentage of brain water content was measured at three days after ICH. n = 6 mice/group. (h) Field selection for ICH brain. * p

    Article Snippet: Hb induced large amounts of ROS (vehicle: 0.35 ± 0.05 × 10E7 , Hb: 2.89 ± 0.51 × 10E7 ; n = 8 slices/group; p < 0.01; and ( )); DFX and VK-28 were both able to reverse Hb-induced ROS (Hb + DFX: 0.61 ± 0.56 × 10E7 , p < 0.05 vs. Hb; Hb + VK-28: 0.18 ± 0.13 × 10E7 , p < 0.001 vs. Hb; n = 5-8 slices/group; and ( )).

    Techniques: In Vivo, Mouse Assay, Injection, Fluorescence, Staining, Selection

    VK-28 attenuates white matter damage after ICH in vivo. Middle-aged male C57BL/6 mice underwent ICH by collagenase injection. DFX, VK-28, or vehicle was administered i.p. at 6 h after ICH and then daily until sacrifice. (a) Representative images show immunostaining of degraded myelin basic protein (dMBP). The percentage of positively stained area was quantified and is shown to the right. (b) Representative images show immunostaining of beta-amyloid precursor protein (β-APP). The percentage of positively stained area was quantified and is shown to the right. n = 5 mice/group. * p

    Journal: Journal of Cerebral Blood Flow & Metabolism

    Article Title: Neuroprotection of brain-permeable iron chelator VK-28 against intracerebral hemorrhage in mice

    doi: 10.1177/0271678X17709186

    Figure Lengend Snippet: VK-28 attenuates white matter damage after ICH in vivo. Middle-aged male C57BL/6 mice underwent ICH by collagenase injection. DFX, VK-28, or vehicle was administered i.p. at 6 h after ICH and then daily until sacrifice. (a) Representative images show immunostaining of degraded myelin basic protein (dMBP). The percentage of positively stained area was quantified and is shown to the right. (b) Representative images show immunostaining of beta-amyloid precursor protein (β-APP). The percentage of positively stained area was quantified and is shown to the right. n = 5 mice/group. * p

    Article Snippet: Hb induced large amounts of ROS (vehicle: 0.35 ± 0.05 × 10E7 , Hb: 2.89 ± 0.51 × 10E7 ; n = 8 slices/group; p < 0.01; and ( )); DFX and VK-28 were both able to reverse Hb-induced ROS (Hb + DFX: 0.61 ± 0.56 × 10E7 , p < 0.05 vs. Hb; Hb + VK-28: 0.18 ± 0.13 × 10E7 , p < 0.001 vs. Hb; n = 5-8 slices/group; and ( )).

    Techniques: In Vivo, Mouse Assay, Injection, Immunostaining, Staining

    VK-28 reduces Hb-induced cell death and ROS production in OHSCs. (a–c) OHSCs were treated under the conditions shown for 16 h. Slices were stained with propidium iodide (PI). Representative images of PI staining (a), the percentage of PI + cells (b), and LDH activity (c) are shown. Vehicle, n = 5; Hb, n = 7; DFX, n = 9; VK-28, n = 8. (d–e) OHSCs were incubated with hydroethidine (HEt) for 30 min, and fluorescence intensity was measured. Representative images (d) and quantification (e) are shown. n = 5–8 slices/group. * p

    Journal: Journal of Cerebral Blood Flow & Metabolism

    Article Title: Neuroprotection of brain-permeable iron chelator VK-28 against intracerebral hemorrhage in mice

    doi: 10.1177/0271678X17709186

    Figure Lengend Snippet: VK-28 reduces Hb-induced cell death and ROS production in OHSCs. (a–c) OHSCs were treated under the conditions shown for 16 h. Slices were stained with propidium iodide (PI). Representative images of PI staining (a), the percentage of PI + cells (b), and LDH activity (c) are shown. Vehicle, n = 5; Hb, n = 7; DFX, n = 9; VK-28, n = 8. (d–e) OHSCs were incubated with hydroethidine (HEt) for 30 min, and fluorescence intensity was measured. Representative images (d) and quantification (e) are shown. n = 5–8 slices/group. * p

    Article Snippet: Hb induced large amounts of ROS (vehicle: 0.35 ± 0.05 × 10E7 , Hb: 2.89 ± 0.51 × 10E7 ; n = 8 slices/group; p < 0.01; and ( )); DFX and VK-28 were both able to reverse Hb-induced ROS (Hb + DFX: 0.61 ± 0.56 × 10E7 , p < 0.05 vs. Hb; Hb + VK-28: 0.18 ± 0.13 × 10E7 , p < 0.001 vs. Hb; n = 5-8 slices/group; and ( )).

    Techniques: Staining, Activity Assay, Incubation, Fluorescence

    VK-28 treatment decreases microglial activation and M1-like polarization after ICH in vivo. Middle-aged male C57BL/6 mice underwent ICH by collagenase injection. DFX, VK-28, or vehicle was administered i.p. at 6 h after ICH and then daily until sacrifice. (a) Immunostaining of Iba-1 reveals microglia and recruited macrophage around the lesion at three days post-ICH. Quantification of activated Iba-1 + cells is shown. * p

    Journal: Journal of Cerebral Blood Flow & Metabolism

    Article Title: Neuroprotection of brain-permeable iron chelator VK-28 against intracerebral hemorrhage in mice

    doi: 10.1177/0271678X17709186

    Figure Lengend Snippet: VK-28 treatment decreases microglial activation and M1-like polarization after ICH in vivo. Middle-aged male C57BL/6 mice underwent ICH by collagenase injection. DFX, VK-28, or vehicle was administered i.p. at 6 h after ICH and then daily until sacrifice. (a) Immunostaining of Iba-1 reveals microglia and recruited macrophage around the lesion at three days post-ICH. Quantification of activated Iba-1 + cells is shown. * p

    Article Snippet: Hb induced large amounts of ROS (vehicle: 0.35 ± 0.05 × 10E7 , Hb: 2.89 ± 0.51 × 10E7 ; n = 8 slices/group; p < 0.01; and ( )); DFX and VK-28 were both able to reverse Hb-induced ROS (Hb + DFX: 0.61 ± 0.56 × 10E7 , p < 0.05 vs. Hb; Hb + VK-28: 0.18 ± 0.13 × 10E7 , p < 0.001 vs. Hb; n = 5-8 slices/group; and ( )).

    Techniques: Activation Assay, In Vivo, Mouse Assay, Injection, Immunostaining

    VK-28 improves neurologic function and corner turn preferences after ICH in vivo. Middle-aged male C57BL/6 mice underwent ICH by collagenase injection. DFX, VK-28, or vehicle was administered i.p. at 6 h after ICH and then daily until sacrifice. (a) Neurologic deficit score of ICH mice at days 1, 3, 7, and 28. Vehicle: n = 16, DFX: n = 10, VK-28: n = 6. (b) Mouse behavior in the corner turn test was assessed before ICH and on day 1, 7, and 28 after ICH. n = 6 mice/group * p

    Journal: Journal of Cerebral Blood Flow & Metabolism

    Article Title: Neuroprotection of brain-permeable iron chelator VK-28 against intracerebral hemorrhage in mice

    doi: 10.1177/0271678X17709186

    Figure Lengend Snippet: VK-28 improves neurologic function and corner turn preferences after ICH in vivo. Middle-aged male C57BL/6 mice underwent ICH by collagenase injection. DFX, VK-28, or vehicle was administered i.p. at 6 h after ICH and then daily until sacrifice. (a) Neurologic deficit score of ICH mice at days 1, 3, 7, and 28. Vehicle: n = 16, DFX: n = 10, VK-28: n = 6. (b) Mouse behavior in the corner turn test was assessed before ICH and on day 1, 7, and 28 after ICH. n = 6 mice/group * p

    Article Snippet: Hb induced large amounts of ROS (vehicle: 0.35 ± 0.05 × 10E7 , Hb: 2.89 ± 0.51 × 10E7 ; n = 8 slices/group; p < 0.01; and ( )); DFX and VK-28 were both able to reverse Hb-induced ROS (Hb + DFX: 0.61 ± 0.56 × 10E7 , p < 0.05 vs. Hb; Hb + VK-28: 0.18 ± 0.13 × 10E7 , p < 0.001 vs. Hb; n = 5-8 slices/group; and ( )).

    Techniques: In Vivo, Mouse Assay, Injection