Journal: BMC Complementary and Alternative Medicine

Article Title: Protective effect of genistein on radiation-induced intestinal injury in tumor bearing mice

doi: 10.1186/1472-6882-13-103

Figure Lengend Snippet: Photomicrographs of progenitor cells in the jejunum circumference stained with an antibody for Ki-67. ( A and B ) Sham control group, ( C and D ) irradiation control group, and ( E and F ) genistein-pretreatment group 3.5 d after 10 Gy irradiation. Most of the Ki-67-positive cells were detected in the jejunal crypts. The number of Ki-67-positive cells decreased in irradiated mice, which corresponded to surviving crypts at 3.5 d after 10 Gy irradiation. However, higher Ki-67 expression was observed in crypts of increased depth in irradiated groups compared with the sham control group. A greater number of Ki-67-positive cells was observed in the genistein-treated group than in the irradiation control group at this magnification. ( A , C , and E ) magnification × 100. ( B , D , and F ) magnification × 400.

Article Snippet: To assess cell proliferation in tissue samples, the proliferation antigen Ki-67 was analyzed by immunocytochemistry using a polyclonal rabbit anti-Ki-67 antibody (Acris Antibodies GmbH, Hiddenhausen, Germany; diluted 1:500).

Techniques: Staining, Irradiation, Expressing

Journal: Viruses

Article Title: Differential Efficacy of Novel Antiviral Substances in 3D and Monolayer Cell Culture

doi: 10.3390/v12111294

Figure Lengend Snippet: Cultivation of primary human keratinocytes (NHEK) on decellularized extracellular matrix (ECM). The 8 µm sections of ECM were analyzed by microscopy. NHEK grown on decellularized ECM for 2 days were treated with CF594 (red) coupled wheat germ agglutinin (WGA) for staining of sialic acid. N-acetylglucosamin on cell membranes and ECM structure was visualized with a brightfield channel ( A ). After permeabilization, KI-67 ( B ) as a proliferation marker (proliferative cells indicated by arrows), integrin beta 1 (ITGB1) as basal polarization marker ( C ) and the atypical protein kinase C (aPKC) as apical polarization marker ( D ) were visualized with polyclonal antibodies and AF647 (yellow) coupled anti-rabbit secondary antibodies. Cellular nuclei were counterstained with DAPI (blue). Scale bar = 100 µm.

Article Snippet: Proteins were stained with antibodies against integrin beta 1 (ITGB1) (Abcam, Cambridge, UK; #30388, mouse), atypical protein kinase C (aPKC) (Santa Cruz Biotechnology, Dallas, TX, USA; sc-208, rabbit), phosphorylated EGFR (pEGFR) (Abcam; #40815, rabbit), EGFR (Abcam; #52894, rabbit), and KI-67 (Abcam; #16667, rabbit) diluted at 1/100 in PBS with 2% BSA and 0.2% NaN 3 .

Techniques: Microscopy, Staining, Marker

Journal: Viruses

Article Title: Differential Efficacy of Novel Antiviral Substances in 3D and Monolayer Cell Culture

doi: 10.3390/v12111294

Figure Lengend Snippet: Effect of cidofovir and tecovirimat treatment on Cowpox virus (CPXV) infection in primary 3D and 2D cultures. NHEK were grown on decellularized extracellular matrix and infected with CPXV. Infected cells were treated with five concentrations ( n = 3) of cidofovir or tecovirimat. At 48 h post infection (p.i.) viral genome equivalents (GE) were quantified via qPCR and expressed as GE per cell. All receptor tyrosine kinase inhibitor (RTKI)-treated samples were normalized to infected untreated controls ( A ). Expression of the cellular proliferation marker KI-67 ( B ) was determined via qPCR, normalized to the cellular reference gene MYC , and presented relative to the untreated controls. Statistical significance of reduction relative to the respective controls was calculated by two-tailed Student’s t -test (* p < 0.05, *** p < 0.001). Cell viability was determined by ATP measurement with the cell viability assay CellTiter-Glo ® (Promega). Infected but untreated controls ( n = 2) were taken as 100%. Ratios of treated samples ( n = 2) to corresponding controls are illustrated as relative viability ( C ).

Article Snippet: Proteins were stained with antibodies against integrin beta 1 (ITGB1) (Abcam, Cambridge, UK; #30388, mouse), atypical protein kinase C (aPKC) (Santa Cruz Biotechnology, Dallas, TX, USA; sc-208, rabbit), phosphorylated EGFR (pEGFR) (Abcam; #40815, rabbit), EGFR (Abcam; #52894, rabbit), and KI-67 (Abcam; #16667, rabbit) diluted at 1/100 in PBS with 2% BSA and 0.2% NaN 3 .

Techniques: Infection, Expressing, Marker, Two Tailed Test, Viability Assay

Journal: Viruses

Article Title: Differential Efficacy of Novel Antiviral Substances in 3D and Monolayer Cell Culture

doi: 10.3390/v12111294

Figure Lengend Snippet: Effect of different RTKIs on CPXV infection in primary 3D and 2D cultures. NHEK were grown either in monolayer or as 3D cultures for 48 h. Cells were infected at an MOI of 0.1 and treated with different RTKI concentrations ( n = 3). At 48 h p.i. viral GE were quantified via qPCR and described as GE per cell. All RTKI-treated samples were normalized to infected untreated controls ( A ). Expression of the cellular proliferation marker KI-67 ( B ) was determined via qPCR, normalized to the cellular reference gene MYC , and expressed relative to the untreated controls. Statistical significance of reduction relative to the respective controls was calculated by two-tailed Student’s t -test (* p < 0.05, ** p < 0.01, *** p < 0.001). Cell viability was determined by ATP measurement with the cell viability assay CellTiter-Glo ® (Promega). Infected but untreated controls ( n = 2) were taken as 100%. Ratios of treated samples ( n = 2) to corresponding controls are illustrated as relative viability ( C ).

Article Snippet: Proteins were stained with antibodies against integrin beta 1 (ITGB1) (Abcam, Cambridge, UK; #30388, mouse), atypical protein kinase C (aPKC) (Santa Cruz Biotechnology, Dallas, TX, USA; sc-208, rabbit), phosphorylated EGFR (pEGFR) (Abcam; #40815, rabbit), EGFR (Abcam; #52894, rabbit), and KI-67 (Abcam; #16667, rabbit) diluted at 1/100 in PBS with 2% BSA and 0.2% NaN 3 .

Techniques: Infection, Expressing, Marker, Two Tailed Test, Viability Assay

Journal: Viruses

Article Title: Differential Efficacy of Novel Antiviral Substances in 3D and Monolayer Cell Culture

doi: 10.3390/v12111294

Figure Lengend Snippet: Effect of cetuximab treatment on CPXV infection in primary 3D and 2D cultures. NHEK were grown on decellularized extracellular matrix and infected with CPXV. Infected cells were treated with five concentrations ( n = 3) of cetuximab. At 48 h p.i. viral GE were quantified via qPCR and described as GE per cell. All RTKI-treated samples were normalized to infected untreated controls ( A ). Expression of the cellular proliferation marker KI-67 ( B ) was determined via qPCR, normalized to the cellular reference gene MYC , and expressed relative to the untreated controls. Statistical significance of reduction relative to the respective controls was calculated by two-tailed Student’s t -test (* p < 0.05, ** p < 0.01, *** p < 0.001). Cell viability was determined by ATP measurement with the cell viability assay CellTiter-Glo ® (Promega). Infected but untreated controls ( n = 2) were taken as 100%. Ratios of treated samples ( n = 2) to corresponding controls are illustrated as relative viability ( C ).

Article Snippet: Proteins were stained with antibodies against integrin beta 1 (ITGB1) (Abcam, Cambridge, UK; #30388, mouse), atypical protein kinase C (aPKC) (Santa Cruz Biotechnology, Dallas, TX, USA; sc-208, rabbit), phosphorylated EGFR (pEGFR) (Abcam; #40815, rabbit), EGFR (Abcam; #52894, rabbit), and KI-67 (Abcam; #16667, rabbit) diluted at 1/100 in PBS with 2% BSA and 0.2% NaN 3 .

Techniques: Infection, Expressing, Marker, Two Tailed Test, Viability Assay

Journal: Viruses

Article Title: Differential Efficacy of Novel Antiviral Substances in 3D and Monolayer Cell Culture

doi: 10.3390/v12111294

Figure Lengend Snippet: Immunofluorescence assay (IFA) analysis of CPXV infected 2D and 3D cultures treated with different antiviral substances. NHEK cultures were cultured with DMSO as a control (ctrl), 2 µM erlotinib, 4 nM cetuximab or 40 nM tecovirimat and infected with CPXV for 48 h. In 2D cultures and 3D cryosections infected cells were visualized by GFP (green) expression ( A , B ). In 3D cultures cellular nuclei were counterstained with DAPI (blue). Phosphorylated EGFR ( C ) and the cellular proliferation marker KI-67 ( D ) were labelled with specific rabbit polyclonal antibodies and visualized with AF647 (yellow) conjugated anti-rabbit secondary antibodies. Scale bar = 100 µm.

Article Snippet: Proteins were stained with antibodies against integrin beta 1 (ITGB1) (Abcam, Cambridge, UK; #30388, mouse), atypical protein kinase C (aPKC) (Santa Cruz Biotechnology, Dallas, TX, USA; sc-208, rabbit), phosphorylated EGFR (pEGFR) (Abcam; #40815, rabbit), EGFR (Abcam; #52894, rabbit), and KI-67 (Abcam; #16667, rabbit) diluted at 1/100 in PBS with 2% BSA and 0.2% NaN 3 .

Techniques: Immunofluorescence, Infection, Cell Culture, Expressing, Marker

Journal: Viruses

Article Title: Differential Efficacy of Novel Antiviral Substances in 3D and Monolayer Cell Culture

doi: 10.3390/v12111294

Figure Lengend Snippet: Immunofluorescence assay (IFA) analysis of CPXV infected 2D and 3D cultures treated with different antiviral substances. NHEK cultures were cultured with DMSO as a control (ctrl), 2 µM erlotinib, 4 nM cetuximab or 40 nM tecovirimat and infected with CPXV for 48 h. In 2D cultures and 3D cryosections infected cells were visualized by GFP (green) expression ( A , B ). In 3D cultures cellular nuclei were counterstained with DAPI (blue). Phosphorylated EGFR ( C ) and the cellular proliferation marker KI-67 ( D ) were labelled with specific rabbit polyclonal antibodies and visualized with AF647 (yellow) conjugated anti-rabbit secondary antibodies. Scale bar = 100 µm.

Article Snippet: Proteins were stained with antibodies against integrin beta 1 (ITGB1) (Abcam, Cambridge, UK; #30388, mouse), atypical protein kinase C (aPKC) (Santa Cruz Biotechnology, Dallas, TX, USA; sc-208, rabbit), phosphorylated EGFR (pEGFR) (Abcam; #40815, rabbit), EGFR (Abcam; #52894, rabbit), and KI-67 (Abcam; #16667, rabbit) diluted at 1/100 in PBS with 2% BSA and 0.2% NaN 3 .

Techniques: Immunofluorescence, Infection, Cell Culture, Expressing, Marker

Journal: Cell death & disease

Article Title: Loss of thyroid gland circadian PER2 rhythmicity in aged mice and its potential association with thyroid cancer development.

doi: 10.1038/s41419-022-05342-2

Figure Lengend Snippet: Fig. 6 Relationship between PER2 immunoexpression and aging in the human thyroid gland. A In non-tumor thyroid, PER2 immunostaining is more frequently observed in the thyroid follicular epithelium of individuals aged <60 years than in those aged >60 years. B PER2 is more frequently expressed in PTCs of individuals aged <60 years than in those aged >60 years. Scale bar, 20 μm.

Article Snippet: The primary antibody was anti-rabbit PER2 (biorbyt, 1:100) and Ki-67 (Cell Signaling Technology, 1:200).

Techniques: Immunostaining

Journal: Scientific Reports

Article Title: Breast Cancer Cell Invasion into a Three Dimensional Tumor-Stroma Microenvironment

doi: 10.1038/srep34094

Figure Lengend Snippet: ( A ) Cells were stained for EGFR (red), pEGFR (green), and nuclei (blue) (scale bar: 20 μm). ( B ) Representative images of EGFR clusters with corresponding heat maps of relative intensities. ( C ) (+) EGF demonstrated significantly lower EGFR to cell area ratio. ( D , E ) (+) EGF displayed significantly higher pEGFR to cell area ratio and pEGFR to EGFR area ratio ( p < 0.05 calculated from Student’s t -test with n > 18 from more than three devices for each condition).

Article Snippet: Next, IF buffer (0.2% (v/v) Triton X-100 + 0.1% (v/v) BSA (radioimmunoassay grade) + 0.05% Tween 20, 7.7 mM NaN 3 in PBS) + 10% (v/v) goat serum was added into the channels and the devices were incubated at room temperature for 1.5 h. Later, primary antibodies, monoclonal Anti-α-Tubulin (1:500, T9026, Sigma-Aldrich), Ki-67 (1:100, (DSHB Hybridoma Product AFFN-KI67-3E6)), EGFR (1:1000, MA5-13319, Thermo Scientific), or pEGFR (1:100, 3777S, Cell Signaling Technology ® ) were diluted at in IF buffer and devices were parafilmed and kept at 4 °C overnight.

Techniques: Staining

Journal: bioRxiv

Article Title: Resistance to chemical carcinogenesis induction via a dampened inflammatory response in naked mole-rats

doi: 10.1101/2021.10.21.465383

Figure Lengend Snippet: A , Schematic diagram for investigating short-term responses to 3MC after subcutaneous (s.c.) injection into the back skin. B , HE staining and immunohistochemical staining for phospho-histone H2A.X (pH2AX, brown) of the skin of mice and NMRs at 1 week after s.c. injection of 3MC. Scale bars: 100 μm (HE) and 50 μm (pH2AX). Red arrowheads indicate positive cells. C , Quantification of pH2AX-, 8-hydroxy-2′-deoxyguanosine (8-OHdG)-, and TUNEL-positive cells after s.c. injection of 3MC. D , Schematic diagram for investigating short-term responses to DMBA treatment on the back skin. E , HE staining and immunohistochemical staining of pH2AX (brown) in the skin of mice and NMRs at 24 h after DMBA treatment. Scale bars: 100 μm (HE) and 50 μm (pH2AX). Red arrowheads indicate positive cells. F , Quantification of pH2AX-positive cells at 24 h after DMBA treatment. For quantification in C and F , data are presented as the mean ± SD of n = 3–4 animals. Unpaired t -test versus untreated control (Ctrl).

Article Snippet: The sections were incubated with 1% bovine serum albumin in Tris-buffered saline with 0.1% NaN 3 for blocking, and stained with primary antibodies against CD45 (Abcam, ab10558), MPO (DAKO, A0398), IBA1 (FUJIFILM WAKO, 019-19741), CD3 (Nichirei, 413591), Ki67 (Abcam, ab16667), 8-OHdG (Santa Cruz Biotechnology, sc-393871), pH2AX (Cell Signaling Technology [CST], 9718), or HMGB1 (Abcam, ab79823).

Techniques: Injection, Staining, Immunohistochemical staining, TUNEL Assay, Control