Journal: bioRxiv

Article Title: SARS-CoV-2 comprehensive receptor profiling: mechanistic insight to drive new therapeutic strategies

doi: 10.1101/2021.03.11.434937

Figure Lengend Snippet: A. Schematic representation of spike protein constructs. SP, signal peptide; RBD, receptor binding domain; TM, transmembrane domain; CT, cytoplasmic domain; T4, T4 foldon trimerization domain; Fc, Fc region of human IgG; His, hexahistidine tag. B. Reducing SDS-PAGE analysis of purified recombinant spike proteins. C. Size exclusion chromatography UV absorbance trace of purified recombinant spike proteins, with retention volume of molecular weight standard thyroglobulin marked as a dashed line. D. Schematic of the experimental approach take to identify interactors of the SARS-CoV-2 spike protein using cell microarray-based expression library screens, followed by cell microarray and flow cytometric confirmatory screens.

Article Snippet: Identical slides were treated before cell fixation with 5 μg/mL SARS-CoV-2 S1-sheep Fc (Native Antigen Company, UK, catalogue # REC31806) or PBS, each pre-incubated at a 2:1 molar ratio with AlexaFluor647 donkey anti-sheep IgG (H+L) (Thermo Fisher, UK, catalogue # A-21448); or 5 μg/mL SARS-CoV-2 S test protein (Peak Proteins, UK, described above), 5 μg/mL MERS S1-His (Native Antigen Company, UK, catalogue # REC31760) or PBS, each pre-incubated at a 2:1 molar ratio with Penta His Alexa Fluor 647 Conjugate (Qiagen, UK).

Techniques: Construct, Binding Assay, SDS Page, Purification, Recombinant, Size-exclusion Chromatography, Molecular Weight, Microarray, Expressing

Journal: bioRxiv

Article Title: SARS-CoV-2 comprehensive receptor profiling: mechanistic insight to drive new therapeutic strategies

doi: 10.1101/2021.03.11.434937

Figure Lengend Snippet: Representative images from one cell microarray-based confirmatory screen. Expression vectors encoding both ZsGreen 1 and individual library ‘hits’ were reverse-transfected into human HEK293 cells, and live transfectants were probed with the SARS-CoV-2 S test protein, SARS CoV S control protein, or other control treatments. This identified interactions that were specific to SARS-CoV-2 S test protein (bold and underlined), and those that were specific to SARS-CoV-2 S and SARS-CoV S spike proteins (green, not underlined). ZsGreen1 was used for spot localization. * indicates that these are secreted proteins which were synthetically cell surface-tethered.

Article Snippet: Identical slides were treated before cell fixation with 5 μg/mL SARS-CoV-2 S1-sheep Fc (Native Antigen Company, UK, catalogue # REC31806) or PBS, each pre-incubated at a 2:1 molar ratio with AlexaFluor647 donkey anti-sheep IgG (H+L) (Thermo Fisher, UK, catalogue # A-21448); or 5 μg/mL SARS-CoV-2 S test protein (Peak Proteins, UK, described above), 5 μg/mL MERS S1-His (Native Antigen Company, UK, catalogue # REC31760) or PBS, each pre-incubated at a 2:1 molar ratio with Penta His Alexa Fluor 647 Conjugate (Qiagen, UK).

Techniques: Microarray, Expressing, Transfection

Journal: bioRxiv

Article Title: SARS-CoV-2 comprehensive receptor profiling: mechanistic insight to drive new therapeutic strategies

doi: 10.1101/2021.03.11.434937

Figure Lengend Snippet: A. Representative of all cell microarray screening data arising library screens with 10 μg/mL SARS-CoV-2 S test protein using 4 different incubation approaches, and from the cell microarray-based confirmatory screens using 6 different incubation approaches. Either a fluorescent anti-Fc detection antibody or a fluorescent anti-His detection antibody was used, as indicated. Interaction intensities against the 10 SARS-CoV-2 S test protein-specific hits were graded from very weak to strong intensity. White boxes indicate no interaction was observed. * indicates that these are secreted proteins which were cell surface-tethered. B. The 10 SARS-CoV-2 S test protein-specific hits were tested for cross-reactivity with three control proteins, 10 μg/mL SARS-CoV S protein (using anti-Fc detection), 5 μg/mL SARS-COV-2 S1 fragment-sheep Fc fusion protein (using an anti-sheep detection antibody) or 5 μg/mL His-tagged MERS-S1 protein (using an anti-His detection antibody). Interaction intensities were graded from very weak to strong intensity. White boxes indicate no interaction was observed. P/ means added prior to fixation, F/ means added after fixation, N/ means no fixation, /P means a control protein was pre-incubated with the detection antibody at a 2:1 molar ratio, and /S means a control protein was incubated with the cells first, followed by the sequential incubation with detection antibody. Interactions exclusive to SARS-CoV-2 are highlighted in blue.

Article Snippet: Identical slides were treated before cell fixation with 5 μg/mL SARS-CoV-2 S1-sheep Fc (Native Antigen Company, UK, catalogue # REC31806) or PBS, each pre-incubated at a 2:1 molar ratio with AlexaFluor647 donkey anti-sheep IgG (H+L) (Thermo Fisher, UK, catalogue # A-21448); or 5 μg/mL SARS-CoV-2 S test protein (Peak Proteins, UK, described above), 5 μg/mL MERS S1-His (Native Antigen Company, UK, catalogue # REC31760) or PBS, each pre-incubated at a 2:1 molar ratio with Penta His Alexa Fluor 647 Conjugate (Qiagen, UK).

Techniques: Microarray, Incubation

Journal: bioRxiv

Article Title: SARS-CoV-2 comprehensive receptor profiling: mechanistic insight to drive new therapeutic strategies

doi: 10.1101/2021.03.11.434937

Figure Lengend Snippet: Live human HEK293 cells transiently over-expressing both ZsGreen and each specific hit identified by cell microarray, were incubated with 0 to 150 μg/mL of SARS-CoV-2 S protein (left hand panels) or SARS-CoV S protein (right hand panels), and interactions were investigated by flow cytometry. After correcting for background binding, binding curves were generated against each target, from which EC50 values were calculated.

Article Snippet: Identical slides were treated before cell fixation with 5 μg/mL SARS-CoV-2 S1-sheep Fc (Native Antigen Company, UK, catalogue # REC31806) or PBS, each pre-incubated at a 2:1 molar ratio with AlexaFluor647 donkey anti-sheep IgG (H+L) (Thermo Fisher, UK, catalogue # A-21448); or 5 μg/mL SARS-CoV-2 S test protein (Peak Proteins, UK, described above), 5 μg/mL MERS S1-His (Native Antigen Company, UK, catalogue # REC31760) or PBS, each pre-incubated at a 2:1 molar ratio with Penta His Alexa Fluor 647 Conjugate (Qiagen, UK).

Techniques: Expressing, Microarray, Incubation, Flow Cytometry, Binding Assay, Generated

Journal: BMC Cardiovascular Disorders

Article Title: Circular RNA expression profile and its potential regulative role in human abdominal aortic aneurysm

doi: 10.1186/s12872-020-01374-8

Figure Lengend Snippet: circRNAs expression profiles detected by microarray in the AAA group and control group. a The box plot shows the nearly identical distributions of normalized intensity values from the aortic samples of the AAA and control group. b The scatter plot is built to assess the expression variation of circRNAs between the two groups. The X and Y axes indicate the normalized intensity values of each circRNAs from the AAA and control group. The dots above the upper green line and below the lower green line represent the dysregulated circRNAs with a fold change (FC) > 2.0 between the two groups. c The volcano plot presents differentially expressed circRNAs in AAA. The vertical lines correspond to 2-fold upregulation and downregulation, the horizontal line indicates P value of 0.05. The red dots represent the differentially expressed circRNAs (FC > 2.0 and P value < 0.05). d Hierarchical clustering analysis reveals a distinguishable expression profile of circRNAs between the AAA and control group. Each column indicates an aortic sample, each row represents a circRNA. The red and green color indicate high and low expression level, respectively. e Chromosomal distribution of the differentially expressed circRNAs between the two groups

Article Snippet: The fragmented labeled cRNAs were hybridized onto the circRNA expression microarray slide (Arraystar Human circRNA Array V2).

Techniques: Expressing, Microarray, Control

Journal: BMC Cardiovascular Disorders

Article Title: Circular RNA expression profile and its potential regulative role in human abdominal aortic aneurysm

doi: 10.1186/s12872-020-01374-8

Figure Lengend Snippet: Validation of six randomly selected dysregulated circRNAs by qRT-PCR. Each circRNA was evaluated at least three times and compared with the results of microarray. The Y axis indicates the fold change of AAA vs control of each circRNA

Article Snippet: The fragmented labeled cRNAs were hybridized onto the circRNA expression microarray slide (Arraystar Human circRNA Array V2).

Techniques: Biomarker Discovery, Quantitative RT-PCR, Microarray, Control

Journal: BMC Cardiovascular Disorders

Article Title: Circular RNA expression profile and its potential regulative role in human abdominal aortic aneurysm

doi: 10.1186/s12872-020-01374-8

Figure Lengend Snippet: The predicted circRNA/miRNA interaction networks for six randomly selected circRNAs. a , b The red nodes indicate upregulated circRNAs. c - f The blue nodes represent downregulated circRNAs. The green nodes are five complementary binding miRNAs of each circRNA

Article Snippet: The fragmented labeled cRNAs were hybridized onto the circRNA expression microarray slide (Arraystar Human circRNA Array V2).

Techniques: Binding Assay

Journal: Cancer Cell International

Article Title: Spatial and single-cell explorations uncover prognostic significance and immunological functions of mitochondrial calcium uniporter in breast cancer

doi: 10.1186/s12935-024-03327-z

Figure Lengend Snippet: Bee swarm representation of differential expression in breast cancer patients based on various classified parameters. A – B Illustrate MCU mRNA expression levels in breast cancer patients using bee swarm plots in DNA microarray datasets and RNA-sequencing datasets. ( ER estrogen receptor, PR progesterone receptor, HER2 human epidermal growth factor receptor 2, TNBC triple-negative breast cancer)

Article Snippet: The research involved the analysis of human specimens through the utilization of tissue microarray (TMA) slides procured from SuperBioChips Laboratories in Seoul, Republic of Korea.

Techniques: Quantitative Proteomics, Expressing, Microarray, RNA Sequencing