rabbit polyclonal anti metap2  (Proteintech)

Journal: Molecular cell

Article Title: Decoding the Function of Expansion Segments in Ribosomes

doi: 10.1016/j.molcel.2018.11.023

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit polyclonal anti-MetAP2 , Proteintech , Cat# 17040-1-AP, RRID: AB_2144162.

Techniques: Virus, Recombinant, Reverse Transcription, SYBR Green Assay, Reporter Assay, Plasmid Preparation, Software, Negative Control

Journal:

Article Title: Methionine Aminopeptidase-2 Regulates Human Mesothelioma Cell Survival

doi:

Figure Lengend Snippet: Bcl-2 overexpression and zVAD-fmk protect mesothelioma cells from caspase 3 activation and apoptosis induced by MetAP2 inhibition. A: HM cells were exposed to either vehicle or 0.5 μg/ml of fumagillin for 96 hours either in the presence or in the absence of 100 nmol/L of zVAD-fmk. Nucleosome formation was determined as indicated in Materials and Methods. Results are the mean ± SD from four separate experiments. B: HM cells, HM-PKneo, or HM-Bcl-2.2 clones were incubated with 100 nmol/L of MetAP2 anti-sense oligonucleotide either in the presence or the absence of 100 nmol/L of zVAD-fmk. Caspase 3 activity was determined as indicated in Materials and Methods.

Article Snippet: These were incubated for a minimum of 2 hours with either a monoclonal antibody against Bcl-2 (Santa Cruz Biotechnology) or with a rabbit polyclonal anti-MetAP2 antibody (CM33) (Zymed Inc., Histo-Line Laboratories, Milan, Italy) and subsequently exposed to a horseradish peroxidase-conjugated secondary antibody.

Techniques: Over Expression, Activation Assay, Inhibition, Clone Assay, Incubation, Activity Assay

Journal:

Article Title: Methionine Aminopeptidase-2 Regulates Human Mesothelioma Cell Survival

doi:

Figure Lengend Snippet: MetAP expression in normal and malignant mesothelial cells. A: Total RNA, extracted from HUVEC (lane 1), three mesothelioma cell lines (MM-B1, MM-F1, and HM) (lanes 2 to 4), and three primary normal mesothelial cells (NM-1, NM-2, and NM-3) (lanes 5 to 7), was subjected to Northern blotting using specific probes for MetAP1 and MetAP2. β-actin was used as an internal control. B: Densitometric analysis of results in A. C: Forty μg of lysates from cells listed in A were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotted using a specific MetAP2 polyclonal antibody. The MetAP-2 protein was visualized as a band of 67 kd. β-Actin was used as a control for equal protein loading.

Article Snippet: These were incubated for a minimum of 2 hours with either a monoclonal antibody against Bcl-2 (Santa Cruz Biotechnology) or with a rabbit polyclonal anti-MetAP2 antibody (CM33) (Zymed Inc., Histo-Line Laboratories, Milan, Italy) and subsequently exposed to a horseradish peroxidase-conjugated secondary antibody.

Techniques: Expressing, Northern Blot, Polyacrylamide Gel Electrophoresis

Journal:

Article Title: Methionine Aminopeptidase-2 Regulates Human Mesothelioma Cell Survival

doi:

Figure Lengend Snippet: Impact of MetAP2 anti-sense oligonucleotide on mesothelioma cell viability and nucleosome formation. A: Mesothelioma cells (HM) were incubated with increasing concentrations of either a specific MetAP2 anti-sense or a scrambled oligonucleotide. After 24 hours total RNA was extracted and blotted with a specific probe for MetAP2. MetAP2 protein levels were determined by Western blotting after 48 hours exposure to the oligonucleotides. β-actin was used as an internal control. B: HM cells were treated for different periods of time with either lipofectin alone (mock) or with 100 nmol/L of the MetAP2 anti-sense or of a scrambled oligonucleotide. At the end of the incubation, cell viability was assessed by trypan blue exclusion. Data points depict mean values ± SD from two experiments with quadruplicate determinations (*, P = 0.02; **, P < 0.001). C: Normal mesothelial (NM-1) and HM cells were treated as indicated in the legend to B. Nucleosome formation in the presence of lipofectin alone, the MetAP2 anti-sense, or the scrambled oligonucleotide was determined after 96 hours using the cell death detection ELISAPLUS kit as indicated in the Materials and Methods section. Results are the average ± SD from three experiments with duplicate determinations. (*, P = 0.018).

Article Snippet: These were incubated for a minimum of 2 hours with either a monoclonal antibody against Bcl-2 (Santa Cruz Biotechnology) or with a rabbit polyclonal anti-MetAP2 antibody (CM33) (Zymed Inc., Histo-Line Laboratories, Milan, Italy) and subsequently exposed to a horseradish peroxidase-conjugated secondary antibody.

Techniques: Incubation, Western Blot

Journal:

Article Title: Methionine Aminopeptidase-2 Regulates Human Mesothelioma Cell Survival

doi:

Figure Lengend Snippet: MetAP2 inhibition reduces Bcl-2 expression in mesothelioma cells. A: Total RNA was extracted from malignant (HM) and normal (NM-1) mesothelial cells treated with either 0.5 μg/ml of fumagillin or 100 nmol/L of MetAP2 anti-sense oligonucleotide for varying periods of time and hybridized with bcl-2-, bax-, and β-actin-specific probes. RNA from HeLa cells expressing bcl-2 and bax was used as a positive control (+). Results are from a representative experiment (n = 3). B: Bcl-2 protein levels (open circle) and nucleosome formation (filled circle) were determined in fumagillin-treated HM cells as described in Experimental Procedures. Data points represent the mean ± SD from three separate experiments with duplicate determinations. (*, P < 0.05; **, P < 0.001).

Article Snippet: These were incubated for a minimum of 2 hours with either a monoclonal antibody against Bcl-2 (Santa Cruz Biotechnology) or with a rabbit polyclonal anti-MetAP2 antibody (CM33) (Zymed Inc., Histo-Line Laboratories, Milan, Italy) and subsequently exposed to a horseradish peroxidase-conjugated secondary antibody.

Techniques: Inhibition, Expressing, Positive Control

Journal:

Article Title: Methionine Aminopeptidase-2 Regulates Human Mesothelioma Cell Survival

doi:

Figure Lengend Snippet: MetAP2 inhibition decreases telomerase activity in mesothelioma cells. A: Mesothelioma (HM) cells were exposed to either 0.5 μg/ml of fumagillin or 100 nmol/L of MetAP2 anti-sense for the indicated periods of time. Telomerase activity in cell extracts (0.1 μg) was determined as described in Experimental Procedures. Values are expressed as percentage of the telomerase activity present in untreated or scrambled oligo-treated HM cells (white bar) and represent the mean ± SD from three experiments with duplicate determinations (*, P = 0.015; **, P < 0.001). B: TRAP assay of mesothelioma (HM) and normal mesothelial (NM-2) cell extracts from cultures treated for 96 hours with either vehicle or 0.5 μg/ml of fumagillin. Negative control refers to HM cell extracts that were incubated with 200 μg/ml of RNase before the TRAP assay. Positive control was 0.02 μg of HeLa cell extract. Results are from one experiment representative of three.

Article Snippet: These were incubated for a minimum of 2 hours with either a monoclonal antibody against Bcl-2 (Santa Cruz Biotechnology) or with a rabbit polyclonal anti-MetAP2 antibody (CM33) (Zymed Inc., Histo-Line Laboratories, Milan, Italy) and subsequently exposed to a horseradish peroxidase-conjugated secondary antibody.

Techniques: Inhibition, Activity Assay, TRAP Assay, Negative Control, Incubation, Positive Control

Journal:

Article Title: Methionine Aminopeptidase-2 Regulates Human Mesothelioma Cell Survival

doi:

Figure Lengend Snippet: Schematic representation of intracellular pathways controlled by MetAP2. MetAP2 positively regulates Bcl-2 expression. This, on one side contributes to the up-regulation of telomerase activity in the nucleus, and on the other inhibits caspase activation. On the other hand, telomerase activity down-regulates caspase activity. Whether telomerase activity may have also an impact on Bcl-2 expression in this model remains to be established.

Article Snippet: These were incubated for a minimum of 2 hours with either a monoclonal antibody against Bcl-2 (Santa Cruz Biotechnology) or with a rabbit polyclonal anti-MetAP2 antibody (CM33) (Zymed Inc., Histo-Line Laboratories, Milan, Italy) and subsequently exposed to a horseradish peroxidase-conjugated secondary antibody.

Techniques: Expressing, Activity Assay, Activation Assay