Journal: Purinergic Signalling

Article Title: A new role for the P2X7 receptor: a scavenger receptor for bacteria and apoptotic cells in the absence of serum and extracellular ATP

doi: 10.1007/s11302-012-9308-5

Figure Lengend Snippet: The binding of short peptides mimicking P2X7 extracellular domain sequences to a range of phagocytic targets. 24 biotin-tagged peptides were incubated at a concentration of 10 μg/mL with either YG beads (3 μm, 5 μL), Alexa 488 conjugated Staphylococcus aureus or Escherichia coli (2 mg/mL, 5 μL), live S. aureus or E. coli (OD690 = 0.9, 10 μL each) or apoptotic HPB cells (5 × 106/mL, 200 μL) for 30 min, followed by washing and incubation with HRP-labeled streptavidin for 30 min. Particles were washed twice, and resuspended in 100 μL of HRP-labeled streptavidin (Jackson ImmunoResearch Lab, 1:2,000 diluted with PBS containing 1% BSA) for 30 min. After two washes, 500 μL of SuperSignal West Pico was added and the chemiluminescence was measured by a Glo 20/20 luminometer (Promega). From Gu et al. [29]

Article Snippet: Particles were washed twice, and resuspended in 100 μL of HRP-labeled streptavidin (Jackson ImmunoResearch Lab, 1:2,000 diluted with PBS containing 1% BSA) for 30 min. After two washes, 500 μL of SuperSignal West Pico was added and the chemiluminescence was measured by a Glo 20/20 luminometer (Promega).

Techniques: Binding Assay, Incubation, Concentration Assay, Labeling

Journal: Methods in Molecular Biology

Article Title: Transcription Factors

doi: 10.1007/978-1-60761-738-9

Figure Lengend Snippet: Fig. 2. Inhibition of the IGF-1-mediated HIF-1alpha induction by the PI(3)-kinase inhibitor LY294002 and the MEK inhibitor U0126. Serum-starved HepG2 cells were pretreated with 10 µM LY294002 or 10 µM U0126 for 30 min and then treated either with or without 100 nM human IGF-1 (Sigma) and exposed to normoxia (16% O2) or hypoxia (8% O2) for 4 h. Acetic acid was used in controls at a final concentration of 100 nM to keep the pH constant. 100 µg of protein from HepG2 cell lysates were analyzed by Western Blotting with antibodies against HIF-1alpha (Novus Biological Transduction Lab, 1:2,000), or against phospho-ERK1/2 (cell signaling, 1:1,000) where HepG2 cells were stimulated for 15 min with IGF-1. Autoradiographic signals were detected by chemiluminescence (77).

Article Snippet: 100 μg of protein from HepG2 cell lysates were analyzed by Western Blotting with antibodies against HIF-1alpha (Novus Biological Transduction Lab, 1:2,000), or against phospho-ERK1/2 (cell signaling, 1:1,000) where HepG2 cells were stimulated for 15 min with IGF-1.

Techniques: Inhibition, Concentration Assay, Western Blot, Transduction

Journal: Methods in Molecular Biology

Article Title: Transcription Factors

doi: 10.1007/978-1-60761-738-9

Figure Lengend Snippet: Fig. 3. HIF-1alpha is phosphorylated by GSK-3. (a, b) The positive control peptide (CP), the GST and the GST-HIF-1a-TADN wild-type fusion proteins were incubated with 50 mU active GSK-3b and 1 µCi (32P-g ATP) for 30 min at 30°C. Afterwards the phospho- rylated proteins were separated from unbound radioactivity by electrophoresis on a 10% SDS gel. Radioactive proteins were visualized by phosphoimaging. After autora- diography, the membrane was used to detect the respective GST-fusion proteins with an antibody against GST (86).

Article Snippet: 100 μg of protein from HepG2 cell lysates were analyzed by Western Blotting with antibodies against HIF-1alpha (Novus Biological Transduction Lab, 1:2,000), or against phospho-ERK1/2 (cell signaling, 1:1,000) where HepG2 cells were stimulated for 15 min with IGF-1.

Techniques: Positive Control, Incubation, Radioactivity, Electrophoresis, SDS-Gel, Membrane

Journal: Cell Reports Medicine

Article Title: Surface CD52, CD84, and PTGER2 mark mature PMN-MDSCs from cancer patients and G-CSF-treated donors

doi: 10.1016/j.xcrm.2023.101380

Figure Lengend Snippet:

Article Snippet: anti-human PTGER2-FITC (polyclonal) – Lab 1/2 , biorbyt , Cat# orb7558; RRID: AB_10938289.

Techniques: Recombinant, Enzyme-linked Immunosorbent Assay, Staining, Cell Isolation, Amplification, Multiplexing, Expressing, Software