Journal:

Article Title: Immunophenotyping of macrophages in human pulmonary tuberculosis and sarcoidosis

doi: 10.1111/j.0959-9673.2003.00365.x

Figure Lengend Snippet: Antibodies used in this study

Article Snippet: Rabbit anti-human SP-D , , K. Reid (University of Oxford) , Donkey anti-rabbit F(ab′) 2 fragment , Jackson Immuno-research Laboratory , ( Holmskov 2000 ).

Techniques: Control

Journal: Frontiers in Pharmacology

Article Title: Fusion assays for screening of fusion inhibitors targeting SARS-CoV-2 entry and syncytia formation

doi: 10.3389/fphar.2022.1007527

Figure Lengend Snippet: Specificity of different modes of fusion. (A) 19,000 target cells (293T-ACE2, 293T-ACE2-TMPRSS2) transfected with the luciferase and β-galactosidase reporter genes were pre-incubated with 10 μg/ml of anti-spike monoclonal antibody (ACROB SPD-M128), 75 μg/ml soybean trypsin inhibitor (SBTi), 10 μg/ml leupeptin or 10 μM of individual drugs before co-cultured with 19,000 donor 293T cells transfected with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike proteins (SARS-2-S) together with a plasmid encoding the T7 polymerase. The co-cultures were treated as indicated. Fusion activity was measured as luciferase activity normalized against β-galactosidase activity and expressed as a ratio to the solvent control in the respective cell type and fusion condition which is set as 1. Data are presented as mean+/-SD of >3 repeats. (B) Photomicrographs of fusion cell morphology. 50,000 target cells (293T-ACE2, 293T-ACE2-TMPRSS2) pre-incubated with 28 μg/ml anti-spike monoclonal antibody (Sino Biol, 40592-R001) or 10 μM of individual drugs were co-cultured with 50,000 donor 293T cells transfected with an empty vector or the SARS-CoV-2 spike protein. The co-cultures were treated as indicated. Cells were fixed and stained with methylene blue. Bright-field images are of the same magnification x100 and scale. Fields of entire or mostly syncytia formation are framed with red squares. Fields of entire or mostly intact cells are framed with orange squares. Scattered syncytia are circled or outlined red and labelled S. Scattered intact cells are circled or outlined orange and labelled C. Sporadic intact cells and syncytia are marked by orange and red arrowheads, respectively. (C) Viability was measured by XTT assays in fusion cells (293T + 293T-ACE2 or 293T + 293T-ACE2-TMPRSS2) under physiological pH and treated with compounds as in (A) . Data are expressed as % viability to solvent control which is set as 100%. Data are presented as mean+/-SD of three repeats. (D) Negative screen. One of the three cell types: 293T, 293T-ACE2 and 293T-ACE2-TMPRSS2 (30,000 cells) was co-transfected with plasmids encoding the T7 polymerase, luciferase and β-galactosidase reporter genes and then treated with compounds as in (A) . Luciferase, β-galactosidase and the luciferase/β-galactosidase ratio are presented as ratios to the solvent controls which are set as 1. Data are presented as mean+/-SD of three repeats * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: Neutralizing antibodies were used at 10 μg/ml (ACROB SPD-M128) and 28 μg/ml (Sino Biol, 40592-R001), respectively.

Techniques: Transfection, Luciferase, Incubation, Cell Culture, Plasmid Preparation, Activity Assay, Staining