Journal: bioRxiv

Article Title: A novel Gorilla-derived oncolytic Adenovirus with natural selective replication in cancer cells

doi: 10.64898/2026.02.26.708271

Figure Lengend Snippet: (A) A549 cells were infected with GRAd32 or GRAd25 at MOI 50 in the presence (w) or absence (w/o) of an anti-CD46 blocking antibody. Images were acquired 3 days post-infection. Competition with CD46 resulted in a delayed onset of cytopathic effect (CPE) exclusively in GRAd25-infected cells. (B) Flow cytometry analysis of surface CAR and CD46 expression levels in lung cancer (A549, NCI-H727, NCI-H1975, NCI-H1299) and normal (MRC5, HUVEC) cell lines. (C) Infection of MRC5 and A549 cells with a replication-defective GRAd32 GFP reporter virus. GFP expression was evaluated at 48 hours post-infection. The presence of GFP signal confirms that GRAd32 is capable of entering MRC5 cells despite its inability to replicate in this cell line.

Article Snippet: For the evaluation of CD46 and CAR cell surface levels, 2-3 x 10 5 cells (MRC5, HUVEC, A549, NCI-H1299, NCI-H1975, NCI-H727) were collected and incubated with anti-human CD46 (1:50; mouse monoclonal; #12239-MM05, Sino Biological) or with anti-human CAR (1:50; rabbit monoclonal; #10799-R271, Sino Biological) for 30 min at 4°C.

Techniques: Infection, Blocking Assay, Flow Cytometry, Expressing, Virus

Journal: bioRxiv

Article Title: Dynamics of the CD9 interactome during bacterial infection of epithelial cells by proximity labelling proteomics

doi: 10.1101/2024.12.13.628358

Figure Lengend Snippet: (A) CD9 knockout disrupts both meningococcal and staphylococcal adherence. WT and CD9 -/- cells were infected with either meningococci (MC58) or staphylococci (SH1000) for 60 mins at an MOI=50. Cells were disrupted after infection and adherent and internalised bacteria were enumerated by colony forming units (cfu). (B-C) CD9-derived peptide, 800C, reduces meningococcal adherence (B) and staphylcoccal adherence (C) to epithelial cells. WT or CD9 -/- cells were pre-treated with a scrambled peptide or 800C for 60 mins prior to infection. Cells were infected as above and adherence enumerated by cfu. (D) Cell surface expression of meningococcal receptors measured by flow cytometry. WT or CD9 -/- cells were treated with an anti-CD9 antibody (602.29), an anti-CD46 antibody, an anti-CD147 antibody. A mouse IgG isotype control (JC1) was included and expression was determined using a FITC-conjugated secondary antibody. % change was calculated by comparing WT to CD9 -/- cells. (E) Representative blot demonstrating expression of meningococcal and staphylococcal receptors. Whole cell lysates of WT and CD9 -/- cells were electrophoresed and blotted. Blots were probed with an anti-CD44 antibody. An anti-GAPDH antibody was used as a loading control. Densitometry was calculated through ImageJ analysis, removing background with an empty lane and normalising to the loading control. n > 3, mean + SEM.

Article Snippet: Mouse anti-human CD9 IgG1 (MM2/57; Merck, Germany), mouse anti-human CD9 IgG1 (602.29; kind gift of Lynda Partridge, University of Sheffield, Sheffield, UK), mouse anti-human CD147 IgG1 (HIM6; Biolegend, California, USA), mouse anti-human CD46 IgG1 (MEM-258; Thermo Fisher Scientific), mouse anti-human CEACAM1 IgG1 (B3-17; Merck), mouse anti-human CEACAM6 IgG1 (1H7-4B; Merck), mouse anti-human CD44 IgG2a (60224-1-Ig; Proteintech Group, Inc, Illinois, USA), mouse anti-human GAPDH (MAB374; Merck), mouse IgG1 (JC1; in house), mouse IgG2a (02-6200; Thermofisher Scientific), mouse anti-FLAG (M2; Merck), goat anti-mouse HRP (P0447; Agilent, California, USA) were used as described.

Techniques: Knock-Out, Infection, Bacteria, Derivative Assay, Expressing, Flow Cytometry, Control

Journal: bioRxiv

Article Title: Dynamics of the CD9 interactome during bacterial infection of epithelial cells by proximity labelling proteomics

doi: 10.1101/2024.12.13.628358

Figure Lengend Snippet: WT cells were treated with specific siRNAs 72 hours prior to infection. Cells were treated with 800C or a scrambled control peptide for 60 minutes prior to infection. Cells were infected with either meningococci (MC58) or staphylococci (SH1000) for 60 mins at an MOI=50. (A) Flow cytometry demonstrating efficient knockdown of meningococcal and staphylococcal receptors after siRNA treatment prior to infection. Cells were probed with anti-CD147, anti-CD46 and anti-CD44 antibodies. n=1, mean. (B) CD147 knockdown and CD9-derived peptide treatment demonstrate no additive effects on meningococcal adherence. WT and CD9 -/- siRNA treated cells were infected with meningococci (MC58) for 60 mins at MOI=50. (C) CD44 knockdown and CD9-derived peptide treatment demonstrate reduced additive effects in staphylococcal adherence. WT and CD9 -/- siRNA treated cells were infected with staphylococci (SH1000) for 60 mins at MOI=50. Cells were disrupted after infection and adherent and internalised bacteria were enumerated by cfu. n=3, mean + SEM, One-Way ANOVA.

Article Snippet: Mouse anti-human CD9 IgG1 (MM2/57; Merck, Germany), mouse anti-human CD9 IgG1 (602.29; kind gift of Lynda Partridge, University of Sheffield, Sheffield, UK), mouse anti-human CD147 IgG1 (HIM6; Biolegend, California, USA), mouse anti-human CD46 IgG1 (MEM-258; Thermo Fisher Scientific), mouse anti-human CEACAM1 IgG1 (B3-17; Merck), mouse anti-human CEACAM6 IgG1 (1H7-4B; Merck), mouse anti-human CD44 IgG2a (60224-1-Ig; Proteintech Group, Inc, Illinois, USA), mouse anti-human GAPDH (MAB374; Merck), mouse IgG1 (JC1; in house), mouse IgG2a (02-6200; Thermofisher Scientific), mouse anti-FLAG (M2; Merck), goat anti-mouse HRP (P0447; Agilent, California, USA) were used as described.

Techniques: Infection, Control, Flow Cytometry, Knockdown, Derivative Assay, Bacteria

Journal: Frontiers in Cell and Developmental Biology

Article Title: Spatial detection of mitochondrial DNA and RNA in tissues

doi: 10.3389/fcell.2024.1346778

Figure Lengend Snippet: Materials and equipment. Primary antibodies suitable for multiplexing IHC with RNAscope ™ in cryosections.

Article Snippet: Human , Plasma membrane , CD46 , Bio-Rad , MCA2113 , RRID:AB_323983 , 1:100.

Techniques: Multiplexing, RNAscope, Labeling, Transgenic Assay, Clinical Proteomics, Membrane

Journal: Frontiers in Cell and Developmental Biology

Article Title: Spatial detection of mitochondrial DNA and RNA in tissues

doi: 10.3389/fcell.2024.1346778

Figure Lengend Snippet: Materials and equipment. Primary antibodies suitable for multiplexing IHC with RNAscope ™ in cryosections.

Article Snippet: Human , Plasma membrane , CD46 , Bio-Rad , MCA2113 , RRID:AB_323983 , 1:100.

Techniques: Multiplexing, RNAscope, Labeling, Transgenic Assay, Clinical Proteomics, Membrane

Journal: Cancer Biology & Therapy

Article Title: CD46 and CD59 inhibitors enhance complement-dependent cytotoxicity of anti-CD38 monoclonal antibodies daratumumab and isatuximab in multiple myeloma and other B-cell malignancy cells

doi: 10.1080/15384047.2024.2314322

Figure Lengend Snippet: Flow cytometry analyses for CD38, CD46 and CD59. Daudi cells, SU-DHL-8, MOLP8, EJM, and MM.1 R cells were incubated with fluorophore-labeled anti-CD38, CD46 or CD59 antibodies. a) CD38. The gray peak in the flow histogram is the negative control (without antibody staining). The pink area represents cells stained with anti-CD38 antibody. b) CD46 and CD59. The gray curve is the negative control (without antibody staining). The blue curve represents cells detected by anti-CD46 or anti-CD59 antibodies. Shown is also the mean fluorescence intensity (MFI) of CD38, CD46 and CD59.

Article Snippet: The following antibodies were used: APC mouse anti-human CD46 (BD Biosciences), PE mouse anti-human CD59 (BD Biosciences), and PE anti-human CD38 (BioLegend).

Techniques: Flow Cytometry, Incubation, Labeling, Negative Control, Staining, Fluorescence

Journal: Cancer Biology & Therapy

Article Title: CD46 and CD59 inhibitors enhance complement-dependent cytotoxicity of anti-CD38 monoclonal antibodies daratumumab and isatuximab in multiple myeloma and other B-cell malignancy cells

doi: 10.1080/15384047.2024.2314322

Figure Lengend Snippet: Co-treatment of MOLP8 cells with Ad35K++ and rILYd4overcomes resistance to isatuximab after the second cycle of treatment in MOLP8 and SU-DHL-8 cells. a) schematic of the experiment. Test cells (MOLP8 and SU-DHL8) were incubated with isatuximab/daratumumab in the presence of NHS to trigger CDC. CD46 and CD59 MFIs were measured before and 16 hours after adding mAb/NHS (c and f). Cell viability was counted 16 hours after mAb/NHS (b, e, h). Cells were then either incubated with or without Ad35K++/rILYd4 and isatuximab or daratumumab (d, g, i). (b-d) studies with MOLP8 cells. e-i) studies with SU-DHL-8 cells. For each data set, three independent experiments with three technical replicas were performed. *** p < .01, n.S. not significant.

Article Snippet: The following antibodies were used: APC mouse anti-human CD46 (BD Biosciences), PE mouse anti-human CD59 (BD Biosciences), and PE anti-human CD38 (BioLegend).

Techniques: Incubation

Journal: International journal of molecular sciences

Article Title: Peritoneal Expression of Membrane Complement Regulators Is Decreased in Peritoneal Dialysis Patients with Infected Peritonitis.

doi: 10.3390/ijms24119146

Figure Lengend Snippet: Figure 1. Correlations of expressions of membrane complement regulators (CRegs), CD46, CD55, and CD59, deposition of complement (C) activation products such as aC3 and C5b-9, and levels of sC5b-9 in peritoneal dialysis fluid (PDF) with tissue injury score and comparison of sC5-9 levels in PDF between Groups P and NP. Expressions of each of the CRegs CD46, CD55, and CD59 showed good inverse correlations with the severity score for peritoneal injuries ((A–C) respectively). In contrast, deposition of C activation products such as activated C3 (aC3) and C5b-9 correlated with the severity score of peritoneal tissue injures ((D,E) respectively). Adjusted PDF levels of sC5b-9 (sC5b-9/TP) also correlated with the severity score for peritoneal injures (F). Between Groups P and NP, the tissue injury score and PDF levels of sC5b-9/TP were significantly different ((G,H), respectively). Dots in the graph show outliers. Outliers were defined as values greater than the 75th percentile plus 1.5 times the interquartile range. #, p < 0.05.

Article Snippet: Monoclonal Ab (mAb) mouse antihuman CD46 (MEM 258) was purchased from BIO-RAD (Hercules, CA, USA).

Techniques: Membrane, Activation Assay, Comparison

Journal: International journal of molecular sciences

Article Title: Peritoneal Expression of Membrane Complement Regulators Is Decreased in Peritoneal Dialysis Patients with Infected Peritonitis.

doi: 10.3390/ijms24119146

Figure Lengend Snippet: Figure 4. Expressions of CRegs CD46, CD55, and CD59 in those with and without peritonitis and depositions of complement (C) activation products. Frames (A–E) and (F–J) represent peritoneal tissues in Group P and Group NP, respectively. Frames (A,F), frames (B,G), frames (C,H), frames (D,I), and frames (E,J) show expressions of CD46, CD55, and CD59 and depositions of activated C3 (aC3) and C5b- 9, respectively. Graphs (K–O) show results for expressions of CD46, CD55, and CD59 and deposition of aC3 and C5b-9, respectively. Original magnification of (A–C) and (F–H) is ×400, while that of (D,E,I,J) is ×100. Inserted frames in right upper corners of frames (F–H) are at double the original magnification. Scale bars of 50 µm and 200 µm are shown in the left upper corner of frames (A,D), respectively. * shows peritoneal cavity side. Dots in the graph show outliers. Outliers were defined as values greater than the 75th percentile plus 1.5 times the interquartile range. # p < 0.05.

Article Snippet: Monoclonal Ab (mAb) mouse antihuman CD46 (MEM 258) was purchased from BIO-RAD (Hercules, CA, USA).

Techniques: Activation Assay

Journal: International journal of molecular sciences

Article Title: Peritoneal Expression of Membrane Complement Regulators Is Decreased in Peritoneal Dialysis Patients with Infected Peritonitis.

doi: 10.3390/ijms24119146

Figure Lengend Snippet: Figure 5. Pathological changes with accumulation of inflammatory cells of peritonitis and expression of membrane complement regulators (CRegs) caused by Candida spp. or P. aeruginosa (Group P1) and Gram-positive cocci (Group P2). Graphs (A,B) show the tissue injury scores and the proportion of cytokeratin-positive mesothelial cells in the peritoneum, respectively. Graph (C–E) shows the numbers of esterase-positive neutrophils, CD3-positive pan-T cells, and CD68-positive macrophages, respectively. Graphs (F,G) show the scores of fibrin deposition and fibrosis in the peritoneum, respectively. Graphs (H–J) show CRegs CD46, CD55, and CD59, respectively. Group P1 shows cases with fungal infection or P. aeruginosa and Group P2 shows cases with Gram-positive cocci. Dots in the graph show outliers. Outliers were defined as values greater than the 75th percentile plus 1.5 times the interquartile range. #, p < 0.05.

Article Snippet: Monoclonal Ab (mAb) mouse antihuman CD46 (MEM 258) was purchased from BIO-RAD (Hercules, CA, USA).

Techniques: Expressing, Membrane, Infection