pre immune serum igg  (Cell Signaling Technology Inc)

Journal: PLoS ONE

Article Title: The Antioxidant Protein Peroxiredoxin 4 Is Epigenetically Down Regulated in Acute Promyelocytic Leukemia

doi: 10.1371/journal.pone.0016340

Figure Lengend Snippet: ChIP using αH3K27me3 and an IgG control ( A ) and αH3K4me3 and an IgG control ( B ) showing an enrichment of both H3K27me3 and H3K4me3 at the TSS of PRDX4 in APL cells. To rule out a possible involvement of X-inactivation, and thereby H3K27me3, in silencing of PRDX4 expression, male APL samples are compared to a male AML control and female APL samples are compared to a female AML control.

Article Snippet: For immunoprecipitation, protein A dynabeads (Invitrogen, Breda, the Netherlands) were pre-incubated with antibodies against H3K27me3 (07-449, Upstate Biotechnology Inc, Charlottesville, VA) or IgG pre-immune serum as control (Sigma Aldrich, St Louis, MO).

Techniques: Expressing

Journal: Current biology : CB

Article Title: SIRPA-Inhibited, Marrow-Derived Macrophages Engorge, Accumulate, and Differentiate in Antibody-Targeted Regression of Solid Tumors

doi: 10.1016/j.cub.2017.06.005

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Anti-hum Ab and serum purified rabbit IgG (Sigma, pre-immune) were reconstituted per manufacturer’s direction and further diluted using sterile PBS.

Techniques: Recombinant, Lysis, shRNA, Plasmid Preparation, Software

Journal: Genome Biology

Article Title: Post-transcriptional regulation of fruit ripening and disease resistance in tomato by the vacuolar protease SlVPE3

doi: 10.1186/s13059-017-1178-2

Figure Lengend Snippet: SlVPE3 does not bind to the promoters of genes with abnormal expression in SlVPE3 silenced fruit. For the ChIP assay, genomic DNA and proteins from pericarp of tomato fruit at 41 dpa were cross-linked, and the chromatin complexes were co-immunoprecipitated with anti-SlVPE3 antibodies. The promoter regions of the analyzed genes are indicated. Blue boxes represent the binding motifs and numbers indicate the position of these motifs relative to the translational start site. The green fragments with upper-case letters represent the regions used for ChIP-quantitative PCR. Values represent the percentage of DNA fragments that were co-immunoprecipitated with anti-SlVPE3 antibodies or preimmune serum (rabbit IgG) relative to the input DNA. Error bars represent the standard deviation of three independent experiments

Article Snippet: After centrifugation at 12,000 × g for 10 min, the supernatant containing the proteins was immunoprecipitated overnight at 4 °C with 50 μg of anti-SlVPE3 or pre-immune serum IgG (negative control) that was coupled to an agarose support, as described in the Pierce® Co-Immunoprecipitation (Co-IP) Kit (Pierce Biotechnology).

Techniques: Expressing, Immunoprecipitation, Binding Assay, Real-time Polymerase Chain Reaction, Standard Deviation

Journal: Genome Biology

Article Title: Post-transcriptional regulation of fruit ripening and disease resistance in tomato by the vacuolar protease SlVPE3

doi: 10.1186/s13059-017-1178-2

Figure Lengend Snippet: Identification of endogenous SlVPE3-interacting proteins. a Proteins isolated from tomato fruit at 41 days post-anthesis ( dpa ) were immunoprecipitated with anti-SlVPE3 antibodies or pre-immune serum IgG (negative control). The enriched proteins were eluted and submitted to Sequential Window Acquisition of all Theoretical Mass Spectra (SWATH-MS) analysis. Each circle represents a single protein. Most identified proteins showed no difference in abundance, indicating nonspecifically bound proteins. Three proteins ( red circle ) were identified as the SlVPE3-interacting proteins with significant increases in abundance ( P < 0.01; t -test) by using anti-SlVPE3 antibodies compared with pre-immune serum IgG. Additional information related to protein identification is listed in Additional file : Table S8. b The changes in protein abundance in the SlVPE3 RNAi fruit revealed by isobaric tags for relative and absolute quantification (iTRAQ) analysis. The abundance ratios of selected proteins in the SlVPE3 RNAi fruit versus wild-type ( WT ) at 41 and 44 dpa are shown. Values represent means of two biological replicates, and error bars represent standard deviation. Asterisks indicate significant differences at a twofold cut-off between WT and SlVPE3 RNAi fruit

Article Snippet: After centrifugation at 12,000 × g for 10 min, the supernatant containing the proteins was immunoprecipitated overnight at 4 °C with 50 μg of anti-SlVPE3 or pre-immune serum IgG (negative control) that was coupled to an agarose support, as described in the Pierce® Co-Immunoprecipitation (Co-IP) Kit (Pierce Biotechnology).

Techniques: Isolation, Immunoprecipitation, Negative Control, Data-independent acquisition, Quantitative Proteomics, Multiplex sample analysis, Standard Deviation

Journal: Genome Biology

Article Title: Post-transcriptional regulation of fruit ripening and disease resistance in tomato by the vacuolar protease SlVPE3

doi: 10.1186/s13059-017-1178-2

Figure Lengend Snippet: Regulation of SlVPE3 by the RIN transcription factor. a SlVPE3 expression in wild type ( WT ) and the rin mutant during fruit ripening, as determined by quantitative RT-PCR. The gene transcript levels were normalized against the ACTIN gene, followed by normalization against WT at 35 days post-anthesis ( dpa ). Values are shown as the means ± standard deviation (SD). Asterisks indicate significant differences ( P < 0.05; t -test) between WT and the rin mutant. b ChIP-quantitative PCR assays indicated that RIN directly binds to the promoter of SlVPE3 . The promoter structure of SlVPE3 is shown. Blue boxes represent CArG box elements and numbers indicate the position of these motifs relative to the translational start site. Green fragments with upper-case letters represent the regions used for ChIP-quantitative PCR. Values are shown as the means ± SD. Asterisks indicate significant differences ( P < 0.05; t -test) between samples co-immunoprecipitated with anti-RIN antibodies and pre-immune serum. c Gel mobility shift assays revealed the direct binding of RIN to the CArG box element in the promoter region of SlVPE3 . The probe sequences corresponding to the SlVPE3 promoter are shown, with red letters representing the CArG box. The mutated bases in the probes are represented by blue letters. wt , probe with intact CArG box element; mt , probe with mutated CArG box element. One thousand-fold excess amounts of unlabeled probes were added to the binding reaction as a competitor. The specific complexes formed are indicated by arrowheads

Article Snippet: After centrifugation at 12,000 × g for 10 min, the supernatant containing the proteins was immunoprecipitated overnight at 4 °C with 50 μg of anti-SlVPE3 or pre-immune serum IgG (negative control) that was coupled to an agarose support, as described in the Pierce® Co-Immunoprecipitation (Co-IP) Kit (Pierce Biotechnology).

Techniques: Expressing, Mutagenesis, Quantitative RT-PCR, Standard Deviation, Real-time Polymerase Chain Reaction, Immunoprecipitation, Mobility Shift, Binding Assay