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Immunohistochemistry of paraffin-embedded and frozen tissues methods

J. Clin. Invest. 118(1): 89-99 (2007)

Immunohistochemistry of paraffin-embedded and frozen tissues. Paraffin-embedded tumor tissues were sectioned (5 μm thick), mounted on poly-l-lysine-coated glass slides, and allowed to dry overnight at 23°C. These sections were used for detection of PCNA and apoptosis by the ApopTag TUNEL assay, as previously reported (50, 51).

ROP2 Gene Expression in Arabidopsis Guard Cells. results

Plant Cell. 2008 January; 20(1): 75–87.

Figure 1. ROP2 Gene Expression in Arabidopsis Guard Cells. (A) ROP2 PCR products were amplified from 1 μg of total RNA from guard cell (G) and mesophyll cell (M) protoplasts. PCR was repeated twice with similar results. Actin was used as a loading control. (B) Histochemical assay of GUS activity in 4-week-old Arabidopsis seedlings expressing ROP2 promoter:GUS fusions. Bar = 50 μm. Plant Cell. 2008 January; 20(1): 75–87.

Promotion of in vivo neovascularization by transplantation of putative EPCs stimulated by Jag-1–mediated signals. results

Circulation. 2008;118:157-165

Figure 5. Promotion of in vivo neovascularization by transplantation of putative EPCs stimulated by Jag-1–mediated signals. A, Representative laser Doppler perfusion imaging findings in nude mice receiving PBS (no cells) or BM-Lin– cells cocultured with empty-vector– or specific Notch ligand (Jag-1, Dll-1)–transfected 3T3 stromal cells at days 0 and 14 (upper panel). Hindlimb perfusion recovery was significantly enhanced in the Jag-1 group compared with the Dll-1, empty-vector, and PBS groups (n=6 per group, lower panel). *P<0.05 vs PBS; **P<0.01 vs PBS; ***P<0.001 vs PBS; P<0.05 vs vector; P<0.01 vs vector; P<0.05 vs Dll-1; P<0.01 vs Dll-1. B, Histological capillary density by isolectin B4 staining revealed augmented neovascularization in the Jag-1 group but not the Dll-1 group compared with the PBS group. *P<0.05; **P<0.01 (n=4 per group). HPF indicates high-power field. C, Histological density of putative EPCs (BM-Lin– cells obtained from GFP transgenic mice) incorporating into vasculature of ischemic tissue. The density of the incorporating EPCs identified as CD31+/GFP+ cells was significantly greater in the Jag-1 group than in the Dll-1, empty-vector, and PBS groups. Green fluorescence indicates GFP; red signal, CD31. *P<0.05; **P<0.01 (n=4 per group).

biomarker abstract

xue, r. et al. investigation of volatile biomarkers in liver cancer blood using solid-phase microextraction and gas chromatography / mass spectrometry. rapid communications in mass spectrometry 1181-1

The emerging field of biomarkers has applications in the diagnosis, staging, prognosis and monitoring of disease progression, as well as in the monitoring of clinical responses to a therapeutic intervention and the development and delivery of personalized treatments to reduce attrition in clinical trials.

Statistical Analysis statistic

Circulation Research. 2004; 94: 46-52

Statistical Analysis All results are expressed as the mean±SEM. Data were analyzed either by unpaired t test or by ANOVA followed by Fisher’s post hoc test for multiple comparisons. Values of P<0.05 were considered to be statistically significant.

Downregulation of Six MicroRNAs Is Associated with Advanced Stage , Lymph Node Metastasis and Poor Prognosis in Small Cell Carcinoma of the Cervix methods

PLoS One. 2012; 7(3): e33762.

Ethics StatementAll patients agreed to participate in the study and gave written informed consent. This study was approved by the medical ethics committee of Cancer Center of Sun Yat-Sen University and complied with the Declaration of Helsinki.

Obesity, diabetes mellitus, and the risk of female breast cancer in Eastern China statistic

World J Surg Oncol. 2013; 11: 71.

Statistical analysisIn this study, the data were analyzed using SPSS software (version 16.0; SPSS Inc., Chicago, IL, USA). The χ2 and P-values were calculated to compare whether there were significant differences in frequencies and percentages of the analyzed variables between the breast-cancer case and control groups. Univariate and multivariate Cox regression models, whose effects were equal to those of conditional logistic regression models in the analysis to obtain P-values and odds ratios (ORs) with 95% confidence intervals (95% CIs) of the variables. The statistical significance was set at a two-sided P-value of 0.05.

Oxidative modification of peroxiredoxin is associated with drug-induced apoptotic signaling in experimental models of Parkinson disease. methods

J Biol Chem. 2008 Apr 11;283(15):9986-98.

Stable Transfection—To establish a stable cell line overexpressing PRX1 (MN9D/PRX1), MN9D cells were transfected with pCI eukaryotic expression vector (Promega) containing full-length human PRX1 cDNA using Lipofectamine 2000 (Invitrogen) as previously described with minor modifications (32). For gene silencing studies, MN9D cells were stably transfected with pRNAT-U6.1 containing small hairpin RNA of PRX1 designed as a small hairpin RNA and green fluorescent protein as a reporter sequence (MN9D/shPRX1; Genscript). The sequence of the RNA was 5′-GTGATAGAGCCGATGAATTTATTGATATCCGTAAATTCATCGGCTCTATCACTTTTTT-3′ (sense-loop-antisense-termination signal). Approximately 2 weeks after transfection, G-418-resistant cells (MN9D/PRX1 and MN9D/shPRX1) were selected and expanded in Dulbecco's modified Eagle's medium supplemented with 10% heat-inactivated fetal bovine serum and 500 μg/ml G-418. MN9D/PRX1 and MN9D/shPRX1 cells were characterized by immunoblot analysis as described above. MN9D cells stably transfected with an equivalent plasmid vector alone (MN9D/Neo and MN9D/RNeo) were used as controls.

CT With Hepatic Arterioportography as a Pretreatment Examination for Hepatocellular Carcinoma Patients: A Randomized Controlled Trial methods

Am J Gastroenterol. 2013 August; 108(8): 1305–1313.

Radiofrequency ablationRFA was performed on an inpatient basis. The precise procedure of RFA is described elsewhere (30). All RFA procedures were performed percutaneously under ultrasonographic guidance. We used a 17-gauge cooled-tip electrode (Cool-Tip; RF Ablation System, Covidien, Boulder, Colombia, CO) for RFA. Radiofrequency energy was delivered for 6–12min for each application. For large tumors, the electrode was repeatedly inserted into different sites, such that the entire tumor could be enveloped by assumed necrotic volumes. A CT scan with a 5-mm section thickness was performed 1–3 days after RFA to evaluate technical effectiveness. Complete ablation was defined as hypoattenuation of the entire tumor. We intended to ablate not only the tumor but also some of the liver parenchyma surrounding it. When we suspected that some portion of tumor remained nonablated, RFA was repeated. We did not predefine the procedure number in a treatment: treatment was generally continued until CT imaging demonstrated necrosis of the entire tumor.

ceramics in multilayers introduction

Compared to metals, ceramics are lacking in room temperature plasticity and fracture toughness, because of a high lattice fraction stress on most slip systems at low temperatures. In the competation between dislocation glide and brittle fracture in ceramics at low temperatures, brittle fracture wins since the stresses necessary to activate dislocation glide are significantly higher than fracture stress. However, localized dislocation activity in ceramics at room temperature has been observed underneath the indenter.

Triple-negative breast cancer (TNBC) introduction

Triple-negative breast cancer (TNBC) is a subtype of breast tumor that is negative for estrogen receptors (ER) and progesterone receptors (PR) and does not show over-expression of Her-2/neu (HER2). Higher occurrences of TNBC have been noted in younger and African-American women. Few drug choices with the exception of chemotherapy such as anthracycline, taxane and platinum are currently available for the treatment of TNBC. Patients with TNBC have a poor prognosis [1] because of the tendency for early relapse and visceral metastasis [2, 3]. TNBC is also insensitive to some of the effective therapies for breast cancer, including HER2-directed therapies, such as trastuzumab, and endocrine therapies, such as tamoxifen or aromatase inhibitors. Hence, treatment of TNBC remains a great challenge. Gene expression analysis has revealed six subtypes of TNBC [4], including two basal-like subtypes (BL1 and BL2) and immunomodulatory, mesenchymal (M), mesenchymal stem-like (MSL), and luminal androgen receptor (LAR) subtypes. Current and future treatment options include modulation of the DNA repair machinery, p53 family signaling, androgen receptor and MAPK/MEK signaling and PI3K/AKT/mTOR signaling. PI3K/AKT/mTOR inhibitors are one of these therapeutic regimens [5], and some of these inhibitors, such as GDC-0941, BKM120 and Everolimus, have been used as the therapeutic choice in recent clinical trials of TNBC (https://clinicaltrials.gov). It was recently reported that a PI3K inhibitor sensitized tumors to PARP inhibition by impairing BRCA1/2 expression [6].

A novel and generalizable organotypic slice platform to evaluate stem cell potential for targeting pediatric brain tumors introduction

Cancer Cell Int. 2008; 8: 9

One emerging strategy is to use the tumor tracking capacity inherent in many stem cell populations to deliver therapeutic agents to the brain cancer cells. Current limitations of the stem cell therapy strategy include that stem cells are treated as a single entity and lack of uniform technology is adopted for selection of clinically relevant sub-populations of stem cells.,

The correlation between endometrial thickness and outcome of in vitro fertilization and embryo transfer (IVF-ET) outcome title

Reproductive Biology and Endocrinology 2008, 6:40

The correlation between endometrial thickness and outcome of in vitro fertilization and embryo transfer (IVF-ET) outcome

Endothelial cell isolation and proliferation assay. methods

J. Clin. Invest. 118(1): 89-99 (2007)

Endothelial cell isolation and proliferation assay. Primary endothelial cells were isolated from adult mouse livers following mechanical mincing and collagenase (2 mg/ml) digestion for 30 min at 37°C. Cells were isolated by incubating for 30 min at 23°C with magnetic Dynabeads (Invitrogen) labeled with anti-CD31 antibodies and using a magnetic separator (53). Cells were then incubated in DMEM supplemented with 20% FBS, penicillin/streptomycin (100 U/ml), heparin (100 μg/ml), endothelial cell growth factor (100 μg/ml), 1× nonessential amino acids, 2 mM l-glutamine, 1× sodium pyruvate, gentamicin (58 μg/ml), and amphotericin B (25 ng/ml), termed “complete endothelium medium.” To assess the effects of VEGF-A on endothelial cell proliferation, cells from passage 1 or 2 were plated for 4 h in 96-well cell culture plates coated with fibronectin (5 μg/ml) at a density of 4,000 cells per well. The medium was then replaced with fresh complete endothelium medium containing 2% FBS and 0.5% BSA. Cells were incubated at 37°C for 48 h in the absence or presence of 50 ng/ml VEGF-A, prior to the addition of One Solution reagent (Promega) for 2 h. Optical density was then determined at 490 nm on a Molecular Devices microplate reader.

Localization of ROP2 in V. faba Guard Cells. results

Plant Cell. 2008 January; 20(1): 75–87.

Figure 2. Localization of ROP2 in V. faba Guard Cells. (A) An intact V. faba guard cell transformed with RFP-CA-ROP2. (B) An intact V. faba guard cell transformed with RFP-DN-ROP2. (C) An intact V. faba guard cell transformed with RFP alone. (D) An intact V. faba guard cell transformed with GFP-ROP2. (E) An intact V. faba guard cell transformed with GFP alone. In all panels, the guard cells at left were transformed by biolistic bombardment and the cells at right were not. A fluorescence image (right) and the corresponding bright-field image (left) of the same cell are shown. The guard cells shown in (D) and (E) were kept under darkness after bombardment until observation, and those in (A) to (C) were irradiated with white light for 3 h before observation. Black dots in bright-field images are gold particles. Focus was on the mid-plane of the cells. Bars = 10 μm. Plant Cell. 2008 January; 20(1): 75–87.

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