mouse cd164 cdna  (OriGene)

Journal: mBio

Article Title: Genome-Wide Knockout Screen Identifies Human Sialomucin CD164 as an Essential Entry Factor for Lymphocytic Choriomeningitis Virus

doi: 10.1128/mbio.00205-22

Figure Lengend Snippet: Genome-wide CRISPR loss-of-function screen in human cells identify host factors important for LCMV infection. (A) Schematic of CRISPR-based KO screen done in A549 lung epithelial cells for the identification of LCMV host factors. (B) Gene enrichment for CRISPR screen of rLCMV-mCherry infection. Enrichment scores were determined by MaGECK analysis, and genes were colored by biological function. Dotted line indicates −log 10 (enrichment score) = 4. All genes and their enrichment scores can be found in Table S1 . (C) Percentage of infected cells as determined by flow cytometry following infection of A549 homozygous knockouts (CD164, SPR14, IL2RA, and KHNYN) or heterozygous knockouts (ARFRP1, YKT6, ACKR4, RAB10, EMC1, and SYS1) with rLCMV-mCherry. Wild-type cells were used as normalization controls. Cells were infected at an MOI of 1 and harvested at 24 hpi. Error bars indicate standard errors from three independent experiments. (D) Quantification of viral infection in WT, Δ CD164 , Δ CD164 complemented with human CD164 (Δ CD164 + hCD164 ), and Δ CD164 complemented with mouse Cd164 (Δ CD164 + mCd164 ) in A549, 293T, and 3T6 cell type backgrounds. Cells were infected with rLCMV-mCherry at an MOI of 1 and harvested at 24 hpi. Error bars indicate standard errors from three independent experiments.

Article Snippet: Human CD164 (number RC202234; Origene) and mouse Cd164 (number MR201951; Origene) cDNAs were cloned into EcoRV-cut plenti-CMV-Puro-DEST (Addgene number 17452; gift from Eric Campeau and Paul Kaufman) ( ) using NEBuilder HiFi DNA assembly master mix (NEB).

Techniques: Genome Wide, CRISPR, Infection, Flow Cytometry

Journal: mBio

Article Title: Genome-Wide Knockout Screen Identifies Human Sialomucin CD164 as an Essential Entry Factor for Lymphocytic Choriomeningitis Virus

doi: 10.1128/mbio.00205-22

Figure Lengend Snippet: Infection of KO cell lines with a panel of mammarenavirus GP pseudotyped virus. (A to E) Percent infection of Δ CD164 , Δ DAG1 , and Δ CD164 Δ DAG1 double-KO cells relative to WT in either A549 or 293T cell type backgrounds following inoculation with low-DAG1-affinity LCMV strain Armstrong 53b-GP (A) or WE2.2-GP (B) and high-DAG1-affinity strain Armstrong clone 13-GP (C), W54-GP (D), or WE-GP (E) pseudotyped virus as determined by flow cytometry for GFP positivity. Cells were infected at an MOI of 1 and measured 24 hpi. Error bars indicate standard errors from three independent experiments. (F to H) Percent infection of ΔCD164, ΔDAG1, and ΔCD164 ΔDAG1 double-KO cells relative to WT in either A549 or 293T cell type backgrounds following inoculation with LASV-GP (D), GTOV-GP (E), or MACV-GP (F) pseudotyped virus as determined by flow cytometry for GFP positivity. Cells were infected at an MOI of 1 and measured 24 hpi. Error bars indicate standard errors from three independent experiments.

Article Snippet: Human CD164 (number RC202234; Origene) and mouse Cd164 (number MR201951; Origene) cDNAs were cloned into EcoRV-cut plenti-CMV-Puro-DEST (Addgene number 17452; gift from Eric Campeau and Paul Kaufman) ( ) using NEBuilder HiFi DNA assembly master mix (NEB).

Techniques: Infection, Virus, Flow Cytometry

Journal: mBio

Article Title: Genome-Wide Knockout Screen Identifies Human Sialomucin CD164 as an Essential Entry Factor for Lymphocytic Choriomeningitis Virus

doi: 10.1128/mbio.00205-22

Figure Lengend Snippet: CD164 functional region determination through antibody binding, domain deletion, and alanine mutagenesis. (A, left) Schematic of wild-type, Δ CD164 , Δ CD164 + hCD164 (ΔE1), Δ CD164 + hCD164 (ΔE1-2), Δ CD164 + hCD164 (ΔE1-3), Δ CD164 + hCD164 (ΔE1-4), Δ CD164 + hCD164 (ΔE1-5), and Δ CD164 + hCD164 (ΔE2-6). (Right) Complemented A549 and 293T cells were challenged with rLCMV-mCherry (MOI, 1) and infection was measured by flow cytometry at 24 hpi. Percent infection was normalized to wild type. Error bars represent standard errors from three independent experiments. (B, left) Schematic of wild type, ΔCD164 KO + hCD164(N72A), ΔCD164 + hCD164(N77A), ΔCD164 + hCD164(N94A), and ΔCD164 + hCD164(N104A). (Right) Complemented A549 and 293T cells were challenged with rLCMV-mCherry (MOI, 1) and infection was measured by flow cytometry at 24 hpi. Percent infection was normalized to the wild type. Error bars represent standard errors from three independent experiments. (C) Amino acid similarities of the cysteine-rich region in human CD164 and mouse Cd164 determined using the ClustalW program on SnapGene. Yellow circles indicate cysteine residues, red N symbolizes N-linked glycosylation sites, and identical amino acids are highlighted in green. (D) Blockade of LCMV infection with serial dilutions of anti-human CD164 monoclonal mouse antibody clone N6B6 or mouse IgG2a-κ isotope control in wild-type A549 cells. Cells were infected at an MOI of 1 and infection measured at 24 hpi. Error bars indicate standard errors from three independent experiments.

Article Snippet: Human CD164 (number RC202234; Origene) and mouse Cd164 (number MR201951; Origene) cDNAs were cloned into EcoRV-cut plenti-CMV-Puro-DEST (Addgene number 17452; gift from Eric Campeau and Paul Kaufman) ( ) using NEBuilder HiFi DNA assembly master mix (NEB).

Techniques: Functional Assay, Binding Assay, Mutagenesis, Infection, Flow Cytometry, Glycoproteomics, Control

Journal: mBio

Article Title: Genome-Wide Knockout Screen Identifies Human Sialomucin CD164 as an Essential Entry Factor for Lymphocytic Choriomeningitis Virus

doi: 10.1128/mbio.00205-22

Figure Lengend Snippet: Characterization of CD164 as a therapeutic target in human placenta. (A) Double immunofluorescence staining for CD164 and CK7 or isotype staining followed by counterstaining with DAPI in villous trophoblastic tissue. Original images were taken by confocal microscopy at ×100 magnification. Scale bar represents 20 μm. (B) Immunofluorescence imaging of JEG-3 placenta cells preincubated with various concentrations of anti-CD164 MAb N6B6 and infected with r3LCMV-mCherry at an MOI of 0.5. Cells were fixed and imaged at ×10 magnification 24 hpi. Scale bar represents 20 μm. (C) Quantification of percent infection of JEG-3 placenta cells preincubated with various concentrations of anti-CD164 MAb N6B6 and infected with r3LCMV-mCherry at MOI of 0.5. Analysis was done on 4 fields of view in 2 independent infections and normalized to infection control.

Article Snippet: Human CD164 (number RC202234; Origene) and mouse Cd164 (number MR201951; Origene) cDNAs were cloned into EcoRV-cut plenti-CMV-Puro-DEST (Addgene number 17452; gift from Eric Campeau and Paul Kaufman) ( ) using NEBuilder HiFi DNA assembly master mix (NEB).

Techniques: Double Immunofluorescence Staining, Staining, Confocal Microscopy, Immunofluorescence, Imaging, Infection, Control

Journal: mBio

Article Title: Genome-Wide Knockout Screen Identifies Human Sialomucin CD164 as an Essential Entry Factor for Lymphocytic Choriomeningitis Virus

doi: 10.1128/mbio.00205-22

Figure Lengend Snippet: CD164 functional region determination through antibody binding, domain deletion, and alanine mutagenesis. (A, left) Schematic of wild-type, Δ CD164 , Δ CD164 + hCD164 (ΔE1), Δ CD164 + hCD164 (ΔE1-2), Δ CD164 + hCD164 (ΔE1-3), Δ CD164 + hCD164 (ΔE1-4), Δ CD164 + hCD164 (ΔE1-5), and Δ CD164 + hCD164 (ΔE2-6). (Right) Complemented A549 and 293T cells were challenged with rLCMV-mCherry (MOI, 1) and infection was measured by flow cytometry at 24 hpi. Percent infection was normalized to wild type. Error bars represent standard errors from three independent experiments. (B, left) Schematic of wild type, ΔCD164 KO + hCD164(N72A), ΔCD164 + hCD164(N77A), ΔCD164 + hCD164(N94A), and ΔCD164 + hCD164(N104A). (Right) Complemented A549 and 293T cells were challenged with rLCMV-mCherry (MOI, 1) and infection was measured by flow cytometry at 24 hpi. Percent infection was normalized to the wild type. Error bars represent standard errors from three independent experiments. (C) Amino acid similarities of the cysteine-rich region in human CD164 and mouse Cd164 determined using the ClustalW program on SnapGene. Yellow circles indicate cysteine residues, red N symbolizes N-linked glycosylation sites, and identical amino acids are highlighted in green. (D) Blockade of LCMV infection with serial dilutions of anti-human CD164 monoclonal mouse antibody clone N6B6 or mouse IgG2a-κ isotope control in wild-type A549 cells. Cells were infected at an MOI of 1 and infection measured at 24 hpi. Error bars indicate standard errors from three independent experiments.

Article Snippet: Briefly, primary CD164 mouse anti-human clone N6B6 (BD Pharmingen), mouse isotype (BD Pharmingen), or cytokeratin-7 rat anti-human (Sigma Millipore) were used to probe sections overnight at 4°C.

Techniques: Functional Assay, Binding Assay, Mutagenesis, Infection, Flow Cytometry

Journal: mBio

Article Title: Genome-Wide Knockout Screen Identifies Human Sialomucin CD164 as an Essential Entry Factor for Lymphocytic Choriomeningitis Virus

doi: 10.1128/mbio.00205-22

Figure Lengend Snippet: Characterization of CD164 as a therapeutic target in human placenta. (A) Double immunofluorescence staining for CD164 and CK7 or isotype staining followed by counterstaining with DAPI in villous trophoblastic tissue. Original images were taken by confocal microscopy at ×100 magnification. Scale bar represents 20 μm. (B) Immunofluorescence imaging of JEG-3 placenta cells preincubated with various concentrations of anti-CD164 MAb N6B6 and infected with r3LCMV-mCherry at an MOI of 0.5. Cells were fixed and imaged at ×10 magnification 24 hpi. Scale bar represents 20 μm. (C) Quantification of percent infection of JEG-3 placenta cells preincubated with various concentrations of anti-CD164 MAb N6B6 and infected with r3LCMV-mCherry at MOI of 0.5. Analysis was done on 4 fields of view in 2 independent infections and normalized to infection control.

Article Snippet: Briefly, primary CD164 mouse anti-human clone N6B6 (BD Pharmingen), mouse isotype (BD Pharmingen), or cytokeratin-7 rat anti-human (Sigma Millipore) were used to probe sections overnight at 4°C.

Techniques: Double Immunofluorescence Staining, Staining, Confocal Microscopy, Immunofluorescence, Imaging, Infection

Journal: eLife

Article Title: Anatomical basis and physiological role of cerebrospinal fluid transport through the murine cribriform plate

doi: 10.7554/eLife.44278

Figure Lengend Snippet:

Article Snippet: Antibody , Mouse monoclonal anti-CD164 , Santa Cruz Biotechnology , Cat# sc-271179; RRID: AB_10613973 , IF (1:250).

Techniques: Software