fxa protease  (New England Biolabs)


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    Name:
    Factor Xa Protease
    Description:
    Factor Xa Protease 250 ug
    Catalog Number:
    P8010L
    Price:
    305
    Category:
    Proteases
    Size:
    250 ug
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    Structured Review

    New England Biolabs fxa protease
    Factor Xa Protease
    Factor Xa Protease 250 ug
    https://www.bioz.com/result/fxa protease/product/New England Biolabs
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    fxa protease - by Bioz Stars, 2021-05
    99/100 stars

    Images

    1) Product Images from "Using Cystine Knot Proteins as a Novel Approach to Retarget Oncolytic Measles Virus"

    Article Title: Using Cystine Knot Proteins as a Novel Approach to Retarget Oncolytic Measles Virus

    Journal: Molecular Therapy Oncolytics

    doi: 10.1016/j.omto.2017.09.005

    Proteolytic Removal of the CKP Prevents Infection by MV-CKPint Virus MV-CKPint was treated with FXa protease at 100 and 200 μg/mL for 2 hr. Untreated and protease-treated virus was used to infect human glioblastoma (U87), melanoma (MDAMB-435) and vero cells for 3 hr at 37°C. The number of live cells was quantified 96 hr post-infection using the trypan blue exclusion method (bar graphs). The experiments were performed in three independent repeats. Error bars represent SEM. *p
    Figure Legend Snippet: Proteolytic Removal of the CKP Prevents Infection by MV-CKPint Virus MV-CKPint was treated with FXa protease at 100 and 200 μg/mL for 2 hr. Untreated and protease-treated virus was used to infect human glioblastoma (U87), melanoma (MDAMB-435) and vero cells for 3 hr at 37°C. The number of live cells was quantified 96 hr post-infection using the trypan blue exclusion method (bar graphs). The experiments were performed in three independent repeats. Error bars represent SEM. *p

    Techniques Used: Infection

    2) Product Images from ""

    Article Title:

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M110.193490

    Schematic diagram of the topology of Gwt1p. A , summary of the results of glycosylation ( Glyco ) mapping and fXa cleavage analysis. Four conserved regions ( A–D ) and essential residues (Asp-145, Lys-155, Asp-165, and Arg-216) are also shown. Asp-145
    Figure Legend Snippet: Schematic diagram of the topology of Gwt1p. A , summary of the results of glycosylation ( Glyco ) mapping and fXa cleavage analysis. Four conserved regions ( A–D ) and essential residues (Asp-145, Lys-155, Asp-165, and Arg-216) are also shown. Asp-145

    Techniques Used:

    Determination of membrane topology of Gwt1p by factor Xa protease cleavage analysis. Membrane fractions from GC1 cells expressing wild-type ( WT ) or Gwt1p fusion proteins with fXa cleavage sequences, tagged with an HA epitope, were prepared and digested
    Figure Legend Snippet: Determination of membrane topology of Gwt1p by factor Xa protease cleavage analysis. Membrane fractions from GC1 cells expressing wild-type ( WT ) or Gwt1p fusion proteins with fXa cleavage sequences, tagged with an HA epitope, were prepared and digested

    Techniques Used: Expressing

    3) Product Images from "Recombinant Measles Viruses Efficiently Entering Cells through Targeted Receptors"

    Article Title: Recombinant Measles Viruses Efficiently Entering Cells through Targeted Receptors

    Journal: Journal of Virology

    doi:

    Genomic structure, protein composition, and replication of recombinant MV. (A) Plasmid p(+)MV-NSe coding for the MV antigenome (top), Pac I- Spe I fragments used for subcloning (center), and amino acid sequences (one-letter code) of the junctions between the H protein ectodomain and the specificity domains (bottom). Coding regions of the six MV cistrons are represented by solid black boxes, the transmembrane segment of the F and H proteins by a horizontally lined box, the FXa cleavage site and the flexible linker by a hatched box, and the ligand by a gray box. Arginine residues in parentheses have been deleted. For details see text. (B) Protein composition of recombinant viruses. Viral particles were purified by centrifugation through a 20% sucrose layer onto a 60% sucrose cushion and subsequent pelleting. The viruses were titrated, 5,000 PFU was subjected to lysis, and proteins were separated by SDS-PAGE. For immunodetection, a MV-specific antiserum was used. (C) FXa protease sensitivity of hybrid H proteins. Purified viral particles (5,000 PFU) were lysed and incubated for 1 h without (−) or with (+) FXa protease at room temperature. Proteins were separated by SDS-PAGE and detected with an H-specific antiserum. (D) FACS analysis of CD46 (interrupted line), IGF1r (dotted line), and EGFr (continuous line) expression on Vero cells. Vertical axis, cell number; horizontal axis, fluorescence intensity. The greyed profile represents incubation of the cells without the primary antibody. (E) Time course of released (left panel) and cell-associated (right panel) virus production in Vero cells infected with parental MV (diamonds), MV-H/XhEGF (squares), and MV-H/XhIGF1 (triangles). Cells were infected at a MOI of 3 and were incubated at 32°C for the times indicated. Viral titers were determined by 50% end-point dilution. Indicated values are averages of three experiments.
    Figure Legend Snippet: Genomic structure, protein composition, and replication of recombinant MV. (A) Plasmid p(+)MV-NSe coding for the MV antigenome (top), Pac I- Spe I fragments used for subcloning (center), and amino acid sequences (one-letter code) of the junctions between the H protein ectodomain and the specificity domains (bottom). Coding regions of the six MV cistrons are represented by solid black boxes, the transmembrane segment of the F and H proteins by a horizontally lined box, the FXa cleavage site and the flexible linker by a hatched box, and the ligand by a gray box. Arginine residues in parentheses have been deleted. For details see text. (B) Protein composition of recombinant viruses. Viral particles were purified by centrifugation through a 20% sucrose layer onto a 60% sucrose cushion and subsequent pelleting. The viruses were titrated, 5,000 PFU was subjected to lysis, and proteins were separated by SDS-PAGE. For immunodetection, a MV-specific antiserum was used. (C) FXa protease sensitivity of hybrid H proteins. Purified viral particles (5,000 PFU) were lysed and incubated for 1 h without (−) or with (+) FXa protease at room temperature. Proteins were separated by SDS-PAGE and detected with an H-specific antiserum. (D) FACS analysis of CD46 (interrupted line), IGF1r (dotted line), and EGFr (continuous line) expression on Vero cells. Vertical axis, cell number; horizontal axis, fluorescence intensity. The greyed profile represents incubation of the cells without the primary antibody. (E) Time course of released (left panel) and cell-associated (right panel) virus production in Vero cells infected with parental MV (diamonds), MV-H/XhEGF (squares), and MV-H/XhIGF1 (triangles). Cells were infected at a MOI of 3 and were incubated at 32°C for the times indicated. Viral titers were determined by 50% end-point dilution. Indicated values are averages of three experiments.

    Techniques Used: Recombinant, Plasmid Preparation, Subcloning, Purification, Centrifugation, Lysis, SDS Page, Immunodetection, Incubation, FACS, Expressing, Fluorescence, Infection

    Competition of viral entry by soluble receptors (A) and the effect of proteolytic cleavage of the specificity domain on entry (B). (A) Competition of viral infections by soluble EGF (sEGF; top panel) or soluble IGF1 (sIGF1; bottom panel). CHO-hEGFr and Vero cells pretreated or not with sEGF were infected with MV green or MV green -H/XhEGF at a MOI of 0.3. 3T3-hIGF1r and Vero cells pretreated or not with sIGF1 were infected with MV green and MV green -H/XhIGF1 at a MOI of 0.1. The number of infected cells was determined at 24 h p.i. by counting GFP-expressing cells in a standard area. Percentages are relative to the number of GFP-expressing cells in infections without added soluble receptor. Results obtained with MV Edmonston on Vero cells are indicated by gray columns. Results obtained with recombinant MV on Vero cells are indicated by white columns and on rodent cells by black columns. (B) Effects of the treatment of recombinant viruses with FXa protease prior to infection. Vero and CHO-hEGFr or Vero and 3T3-hIGF1r cells were infected with MV green -H/XhEGF or MV green -H/XhIGF1 (10 4 PFU/well), respectively, and were pretreated with 0, 5, or 50 μg of FXa protease/ml. The number of GFP-expressing cells was determined 42 h p.i. Percentages refer to the number of cells infected with untreated virus. For details, see text.
    Figure Legend Snippet: Competition of viral entry by soluble receptors (A) and the effect of proteolytic cleavage of the specificity domain on entry (B). (A) Competition of viral infections by soluble EGF (sEGF; top panel) or soluble IGF1 (sIGF1; bottom panel). CHO-hEGFr and Vero cells pretreated or not with sEGF were infected with MV green or MV green -H/XhEGF at a MOI of 0.3. 3T3-hIGF1r and Vero cells pretreated or not with sIGF1 were infected with MV green and MV green -H/XhIGF1 at a MOI of 0.1. The number of infected cells was determined at 24 h p.i. by counting GFP-expressing cells in a standard area. Percentages are relative to the number of GFP-expressing cells in infections without added soluble receptor. Results obtained with MV Edmonston on Vero cells are indicated by gray columns. Results obtained with recombinant MV on Vero cells are indicated by white columns and on rodent cells by black columns. (B) Effects of the treatment of recombinant viruses with FXa protease prior to infection. Vero and CHO-hEGFr or Vero and 3T3-hIGF1r cells were infected with MV green -H/XhEGF or MV green -H/XhIGF1 (10 4 PFU/well), respectively, and were pretreated with 0, 5, or 50 μg of FXa protease/ml. The number of GFP-expressing cells was determined 42 h p.i. Percentages refer to the number of cells infected with untreated virus. For details, see text.

    Techniques Used: Infection, Expressing, Recombinant

    Related Articles

    Purification:

    Article Title: Species-specific immune responses generated by histidyl-tRNA synthetase immunization are associated with muscle and lung inflammation
    Article Snippet: Expressed proteins were purified with amylose resin per the manufacturer's protocol (New England Biolabs, Ipswich, MA), filter sterilized, and then subjected to additional column purification for endotoxin removal (Profos AG, Regensburg, Germany) prior to use in proliferation assays. .. As previously described [ ], full length versions of Jo-1 were cleaved with Factor Xa (New England Biolabs, Ipswich, MA) to release the MBP moiety and further purified using ion exchange chromatography. .. Overlapping peptides (18-20 mers) comprising the amino terminal 120 amino acids of murine Jo-1 were synthesized and HPLC purified by the University of Pittsburgh Molecular Medicine Institute using Fmoc chemistry.

    Article Title: Protective efficacy of vaccination with Neospora caninum multiple recombinant antigens against experimental Neospora caninum infection
    Article Snippet: The expressions of recombinant proteins were induced by adding isopropyl-1-thio-β-D-galactopyranoside (IPTG) to a final concentration of 1 mM, and they were purified using an amylase affinity chromatography column (New England BioLab). .. The maltose-binding protein (MBP) was removed from the purified recombinant proteins using Factor Xa (New England BioLab) according to the manufacturer's instructions. .. The expressions and purities of the recombinant proteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE).

    Ion Exchange Chromatography:

    Article Title: Species-specific immune responses generated by histidyl-tRNA synthetase immunization are associated with muscle and lung inflammation
    Article Snippet: Expressed proteins were purified with amylose resin per the manufacturer's protocol (New England Biolabs, Ipswich, MA), filter sterilized, and then subjected to additional column purification for endotoxin removal (Profos AG, Regensburg, Germany) prior to use in proliferation assays. .. As previously described [ ], full length versions of Jo-1 were cleaved with Factor Xa (New England Biolabs, Ipswich, MA) to release the MBP moiety and further purified using ion exchange chromatography. .. Overlapping peptides (18-20 mers) comprising the amino terminal 120 amino acids of murine Jo-1 were synthesized and HPLC purified by the University of Pittsburgh Molecular Medicine Institute using Fmoc chemistry.

    Incubation:

    Article Title: GTP-dependent formation of straight tubulin oligomers leads to microtubule nucleation
    Article Snippet: The tubulin eluted from the FLAG-affinity column was concentrated to > 5 mg/ml with an Amicon Ultracel-30K filter (EMD Millipore), centrifuged at 100,000 rpm (∼540,000 g ; Beckman TLA-100.3 rotor) for 10 min at 4°C, and the supernatant was incubated at 30°C after adding glycerol (final 33% vol/vol) and GTP (2 mM). .. After 2 h of incubation, Factor Xa protease (New England Biolabs) was added to the MT solution at a molar ratio of 1:22 (Factor Xa:tubulin) and reacted overnight at 25°C. .. The next morning, the sample solution was centrifuged at 100,000 rpm (Beckman TLA-100.3 rotor) for 15 min at 30°C, and the precipitated MTs were suspended in BRB80 buffer solution (80 mM Pipes, 2 mM MgCl2 , and 1 mM EGTA, pH 6.8) containing 1 mM GTP (for WT tubulin) or 1 mM GDP (for Y222F tubulin) at 4°C, and left for 30 min on ice.

    Article Title: Using Cystine Knot Proteins as a Novel Approach to Retarget Oncolytic Measles Virus
    Article Snippet: .. For the FXa proteolysis assay, prior to cellular infection, the number of MV-CKPint virus equivalent to an MOI of 1 was incubated with FXa protease (New England Biolabs, Ipswich, MA, USA) at final concentrations of 100 and 200 μg/mL in Opti-MEM media. .. The virus was incubated with FXa at room temperature for 1 hr and then for an additional 1 hr at 37°C in the water bath.

    other:

    Article Title:
    Article Snippet: When fXa cleavage sites were inserted into loop 5–6 ( 6 ), loop 7–8 ( 9 ), or loop 9–10 ( 12 ) of Gwt1p (see also A ), the fXa protease was accessible to the fusion proteins in the presence or absence of digitonin, suggesting that these positions were located on the cytosolic side of the ER membrane.

    Recombinant:

    Article Title: Protective efficacy of vaccination with Neospora caninum multiple recombinant antigens against experimental Neospora caninum infection
    Article Snippet: The expressions of recombinant proteins were induced by adding isopropyl-1-thio-β-D-galactopyranoside (IPTG) to a final concentration of 1 mM, and they were purified using an amylase affinity chromatography column (New England BioLab). .. The maltose-binding protein (MBP) was removed from the purified recombinant proteins using Factor Xa (New England BioLab) according to the manufacturer's instructions. .. The expressions and purities of the recombinant proteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE).

    Proteolysis Assay:

    Article Title: Using Cystine Knot Proteins as a Novel Approach to Retarget Oncolytic Measles Virus
    Article Snippet: .. For the FXa proteolysis assay, prior to cellular infection, the number of MV-CKPint virus equivalent to an MOI of 1 was incubated with FXa protease (New England Biolabs, Ipswich, MA, USA) at final concentrations of 100 and 200 μg/mL in Opti-MEM media. .. The virus was incubated with FXa at room temperature for 1 hr and then for an additional 1 hr at 37°C in the water bath.

    Infection:

    Article Title: Using Cystine Knot Proteins as a Novel Approach to Retarget Oncolytic Measles Virus
    Article Snippet: .. For the FXa proteolysis assay, prior to cellular infection, the number of MV-CKPint virus equivalent to an MOI of 1 was incubated with FXa protease (New England Biolabs, Ipswich, MA, USA) at final concentrations of 100 and 200 μg/mL in Opti-MEM media. .. The virus was incubated with FXa at room temperature for 1 hr and then for an additional 1 hr at 37°C in the water bath.

    Article Title: Recombinant Measles Viruses Efficiently Entering Cells through Targeted Receptors
    Article Snippet: When MV-H/XhEGF particles were treated with 5 or 50 μg of FXa protease/ml for 1 h, the number of green fluorescent CHO-hEGFr cells diminished by almost two orders of magnitude (Fig. B, left panel, black column), whereas FXa treatment did not significantly change the infectivity of these particles on Vero cells (Fig. B, left panel, white columns). .. Analogously, proteolytic cleavage of MV-H/XhIGF1 virus particles with FXa protease resulted in the loss of more than 80% of the infectivity selectively in 3T3-hIGF1r cells (Fig. B, right panel, compare black and white columns). .. Thus, by two different approaches, competition with a soluble form of the displayed domain and proteolytic cleavage of that domain from viral particles, it was confirmed that entry of recombinant MV in rodent cells depends on the interaction of the specificity domain with the cognate receptor.

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    New England Biolabs fxa protease
    Proteolytic Removal of the CKP Prevents Infection by <t>MV-CKPint</t> Virus MV-CKPint was treated with <t>FXa</t> protease at 100 and 200 μg/mL for 2 hr. Untreated and protease-treated virus was used to infect human glioblastoma (U87), melanoma (MDAMB-435) and vero cells for 3 hr at 37°C. The number of live cells was quantified 96 hr post-infection using the trypan blue exclusion method (bar graphs). The experiments were performed in three independent repeats. Error bars represent SEM. *p
    Fxa Protease, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/fxa protease/product/New England Biolabs
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    fxa protease - by Bioz Stars, 2021-05
    99/100 stars
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    Proteolytic Removal of the CKP Prevents Infection by MV-CKPint Virus MV-CKPint was treated with FXa protease at 100 and 200 μg/mL for 2 hr. Untreated and protease-treated virus was used to infect human glioblastoma (U87), melanoma (MDAMB-435) and vero cells for 3 hr at 37°C. The number of live cells was quantified 96 hr post-infection using the trypan blue exclusion method (bar graphs). The experiments were performed in three independent repeats. Error bars represent SEM. *p

    Journal: Molecular Therapy Oncolytics

    Article Title: Using Cystine Knot Proteins as a Novel Approach to Retarget Oncolytic Measles Virus

    doi: 10.1016/j.omto.2017.09.005

    Figure Lengend Snippet: Proteolytic Removal of the CKP Prevents Infection by MV-CKPint Virus MV-CKPint was treated with FXa protease at 100 and 200 μg/mL for 2 hr. Untreated and protease-treated virus was used to infect human glioblastoma (U87), melanoma (MDAMB-435) and vero cells for 3 hr at 37°C. The number of live cells was quantified 96 hr post-infection using the trypan blue exclusion method (bar graphs). The experiments were performed in three independent repeats. Error bars represent SEM. *p

    Article Snippet: For the FXa proteolysis assay, prior to cellular infection, the number of MV-CKPint virus equivalent to an MOI of 1 was incubated with FXa protease (New England Biolabs, Ipswich, MA, USA) at final concentrations of 100 and 200 μg/mL in Opti-MEM media.

    Techniques: Infection

    Schematic diagram of the topology of Gwt1p. A , summary of the results of glycosylation ( Glyco ) mapping and fXa cleavage analysis. Four conserved regions ( A–D ) and essential residues (Asp-145, Lys-155, Asp-165, and Arg-216) are also shown. Asp-145

    Journal: The Journal of Biological Chemistry

    Article Title:

    doi: 10.1074/jbc.M110.193490

    Figure Lengend Snippet: Schematic diagram of the topology of Gwt1p. A , summary of the results of glycosylation ( Glyco ) mapping and fXa cleavage analysis. Four conserved regions ( A–D ) and essential residues (Asp-145, Lys-155, Asp-165, and Arg-216) are also shown. Asp-145

    Article Snippet: When fXa cleavage sites were inserted into loop 5–6 ( 6 ), loop 7–8 ( 9 ), or loop 9–10 ( 12 ) of Gwt1p (see also A ), the fXa protease was accessible to the fusion proteins in the presence or absence of digitonin, suggesting that these positions were located on the cytosolic side of the ER membrane.

    Techniques:

    Determination of membrane topology of Gwt1p by factor Xa protease cleavage analysis. Membrane fractions from GC1 cells expressing wild-type ( WT ) or Gwt1p fusion proteins with fXa cleavage sequences, tagged with an HA epitope, were prepared and digested

    Journal: The Journal of Biological Chemistry

    Article Title:

    doi: 10.1074/jbc.M110.193490

    Figure Lengend Snippet: Determination of membrane topology of Gwt1p by factor Xa protease cleavage analysis. Membrane fractions from GC1 cells expressing wild-type ( WT ) or Gwt1p fusion proteins with fXa cleavage sequences, tagged with an HA epitope, were prepared and digested

    Article Snippet: When fXa cleavage sites were inserted into loop 5–6 ( 6 ), loop 7–8 ( 9 ), or loop 9–10 ( 12 ) of Gwt1p (see also A ), the fXa protease was accessible to the fusion proteins in the presence or absence of digitonin, suggesting that these positions were located on the cytosolic side of the ER membrane.

    Techniques: Expressing

    Genomic structure, protein composition, and replication of recombinant MV. (A) Plasmid p(+)MV-NSe coding for the MV antigenome (top), Pac I- Spe I fragments used for subcloning (center), and amino acid sequences (one-letter code) of the junctions between the H protein ectodomain and the specificity domains (bottom). Coding regions of the six MV cistrons are represented by solid black boxes, the transmembrane segment of the F and H proteins by a horizontally lined box, the FXa cleavage site and the flexible linker by a hatched box, and the ligand by a gray box. Arginine residues in parentheses have been deleted. For details see text. (B) Protein composition of recombinant viruses. Viral particles were purified by centrifugation through a 20% sucrose layer onto a 60% sucrose cushion and subsequent pelleting. The viruses were titrated, 5,000 PFU was subjected to lysis, and proteins were separated by SDS-PAGE. For immunodetection, a MV-specific antiserum was used. (C) FXa protease sensitivity of hybrid H proteins. Purified viral particles (5,000 PFU) were lysed and incubated for 1 h without (−) or with (+) FXa protease at room temperature. Proteins were separated by SDS-PAGE and detected with an H-specific antiserum. (D) FACS analysis of CD46 (interrupted line), IGF1r (dotted line), and EGFr (continuous line) expression on Vero cells. Vertical axis, cell number; horizontal axis, fluorescence intensity. The greyed profile represents incubation of the cells without the primary antibody. (E) Time course of released (left panel) and cell-associated (right panel) virus production in Vero cells infected with parental MV (diamonds), MV-H/XhEGF (squares), and MV-H/XhIGF1 (triangles). Cells were infected at a MOI of 3 and were incubated at 32°C for the times indicated. Viral titers were determined by 50% end-point dilution. Indicated values are averages of three experiments.

    Journal: Journal of Virology

    Article Title: Recombinant Measles Viruses Efficiently Entering Cells through Targeted Receptors

    doi:

    Figure Lengend Snippet: Genomic structure, protein composition, and replication of recombinant MV. (A) Plasmid p(+)MV-NSe coding for the MV antigenome (top), Pac I- Spe I fragments used for subcloning (center), and amino acid sequences (one-letter code) of the junctions between the H protein ectodomain and the specificity domains (bottom). Coding regions of the six MV cistrons are represented by solid black boxes, the transmembrane segment of the F and H proteins by a horizontally lined box, the FXa cleavage site and the flexible linker by a hatched box, and the ligand by a gray box. Arginine residues in parentheses have been deleted. For details see text. (B) Protein composition of recombinant viruses. Viral particles were purified by centrifugation through a 20% sucrose layer onto a 60% sucrose cushion and subsequent pelleting. The viruses were titrated, 5,000 PFU was subjected to lysis, and proteins were separated by SDS-PAGE. For immunodetection, a MV-specific antiserum was used. (C) FXa protease sensitivity of hybrid H proteins. Purified viral particles (5,000 PFU) were lysed and incubated for 1 h without (−) or with (+) FXa protease at room temperature. Proteins were separated by SDS-PAGE and detected with an H-specific antiserum. (D) FACS analysis of CD46 (interrupted line), IGF1r (dotted line), and EGFr (continuous line) expression on Vero cells. Vertical axis, cell number; horizontal axis, fluorescence intensity. The greyed profile represents incubation of the cells without the primary antibody. (E) Time course of released (left panel) and cell-associated (right panel) virus production in Vero cells infected with parental MV (diamonds), MV-H/XhEGF (squares), and MV-H/XhIGF1 (triangles). Cells were infected at a MOI of 3 and were incubated at 32°C for the times indicated. Viral titers were determined by 50% end-point dilution. Indicated values are averages of three experiments.

    Article Snippet: Analogously, proteolytic cleavage of MV-H/XhIGF1 virus particles with FXa protease resulted in the loss of more than 80% of the infectivity selectively in 3T3-hIGF1r cells (Fig. B, right panel, compare black and white columns).

    Techniques: Recombinant, Plasmid Preparation, Subcloning, Purification, Centrifugation, Lysis, SDS Page, Immunodetection, Incubation, FACS, Expressing, Fluorescence, Infection

    Competition of viral entry by soluble receptors (A) and the effect of proteolytic cleavage of the specificity domain on entry (B). (A) Competition of viral infections by soluble EGF (sEGF; top panel) or soluble IGF1 (sIGF1; bottom panel). CHO-hEGFr and Vero cells pretreated or not with sEGF were infected with MV green or MV green -H/XhEGF at a MOI of 0.3. 3T3-hIGF1r and Vero cells pretreated or not with sIGF1 were infected with MV green and MV green -H/XhIGF1 at a MOI of 0.1. The number of infected cells was determined at 24 h p.i. by counting GFP-expressing cells in a standard area. Percentages are relative to the number of GFP-expressing cells in infections without added soluble receptor. Results obtained with MV Edmonston on Vero cells are indicated by gray columns. Results obtained with recombinant MV on Vero cells are indicated by white columns and on rodent cells by black columns. (B) Effects of the treatment of recombinant viruses with FXa protease prior to infection. Vero and CHO-hEGFr or Vero and 3T3-hIGF1r cells were infected with MV green -H/XhEGF or MV green -H/XhIGF1 (10 4 PFU/well), respectively, and were pretreated with 0, 5, or 50 μg of FXa protease/ml. The number of GFP-expressing cells was determined 42 h p.i. Percentages refer to the number of cells infected with untreated virus. For details, see text.

    Journal: Journal of Virology

    Article Title: Recombinant Measles Viruses Efficiently Entering Cells through Targeted Receptors

    doi:

    Figure Lengend Snippet: Competition of viral entry by soluble receptors (A) and the effect of proteolytic cleavage of the specificity domain on entry (B). (A) Competition of viral infections by soluble EGF (sEGF; top panel) or soluble IGF1 (sIGF1; bottom panel). CHO-hEGFr and Vero cells pretreated or not with sEGF were infected with MV green or MV green -H/XhEGF at a MOI of 0.3. 3T3-hIGF1r and Vero cells pretreated or not with sIGF1 were infected with MV green and MV green -H/XhIGF1 at a MOI of 0.1. The number of infected cells was determined at 24 h p.i. by counting GFP-expressing cells in a standard area. Percentages are relative to the number of GFP-expressing cells in infections without added soluble receptor. Results obtained with MV Edmonston on Vero cells are indicated by gray columns. Results obtained with recombinant MV on Vero cells are indicated by white columns and on rodent cells by black columns. (B) Effects of the treatment of recombinant viruses with FXa protease prior to infection. Vero and CHO-hEGFr or Vero and 3T3-hIGF1r cells were infected with MV green -H/XhEGF or MV green -H/XhIGF1 (10 4 PFU/well), respectively, and were pretreated with 0, 5, or 50 μg of FXa protease/ml. The number of GFP-expressing cells was determined 42 h p.i. Percentages refer to the number of cells infected with untreated virus. For details, see text.

    Article Snippet: Analogously, proteolytic cleavage of MV-H/XhIGF1 virus particles with FXa protease resulted in the loss of more than 80% of the infectivity selectively in 3T3-hIGF1r cells (Fig. B, right panel, compare black and white columns).

    Techniques: Infection, Expressing, Recombinant