Journal: Cell reports

Article Title: Derlin rhomboid pseudoproteases employ substrate engagement and lipid distortion to enable the retrotranslocation of ERAD membrane substrates

doi: 10.1016/j.celrep.2021.109840

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Mouse anti-GAPDH , BIO-RAD , Cat#MCA4740; RRID:AB_2107457.

Techniques: Virus, Recombinant, Mutagenesis, Positive Control, Software

Journal:

Article Title: Reduced Sensitivity to Human Serum Inactivation of Enveloped Viruses Produced by Pig Cells Transgenic for Human CD55 or Deficient for the Galactosyl-?(1-3) Galactosyl Epitope

doi: 10.1128/JVI.78.11.5812-5819.2004

Figure Lengend Snippet: hCD55 and α-Gal expression on porcine cells. (a) Expression of CD55 was analyzed by FACs on both transgenic (TgPAE) and nontransgenic (PAE) pig cells. Staining by anti-hCD55 (shaded histogram), no primary antibody (negative control; solid line), or anti-hCD46 (dotted line, similar profile to that of the no-antibody control) followed by FITC-labeled secondary anti-mouse immunoglobulin antibodies (upper panels) is shown. Staining for α-Gal was a one-step incubation; hence, −lectin on the histogram represents cells only, and +lectin represents cells incubated with IB-4 lectin conjugated to FITC (lower panels). (b) α-Gal expression on ST-IOWA wild-type and ST-IOWA Gal-null (−/−) cells. Expression of the α-Gal antigen was analyzed by FACs, using the IB-4 lectin. A rightward shift of the histogram in the presence of lectin (+lectin) compared to the histogram generated in the absence of lectin (−lectin) indicates expression of the α-Gal antigen. The level of expression was determined using the MFI shift, generated by the Cell Quest FACScan software. The MFI shift was calculated by determining test MFI and subtracting that of negative controls (no primary antibodies for hCD55 and hCD46 expression and no lectin for α-Gal expression), and all the mean shifts are shown in Table ​Table11.

Article Snippet: Samples were then incubated on ice with either primary mouse anti-human CD55 antibody (BRIC110) or mouse anti-human CD46 antibody (J4-48) from Cymbus Bioscience Ltd., diluted to 1:20 in PBS/BA, for 1 h. After three washes in PBS/BA, samples were incubated with fluorescein isothiocyanate (FITC)-conjugated anti-mouse secondary antibody (Jackson ImmunoResearch), diluted to 1:200 in PBS/BA and incubated on ice for 1 h. For α-Gal expression, cells were detached by a cell scraper, washed, resuspended, and stained in a single 1-h incubation with FITC-conjugated Bandeiraea simplicifolia isolectin (IB-4) (Sigma) at 10 μg/ml in PBS/BA on ice, as this lectin is specific for the terminal α-Gal sugar.

Techniques: Expressing, Transgenic Assay, Staining, Negative Control, Labeling, Incubation, Generated, Software

Journal:

Article Title: Reduced Sensitivity to Human Serum Inactivation of Enveloped Viruses Produced by Pig Cells Transgenic for Human CD55 or Deficient for the Galactosyl-?(1-3) Galactosyl Epitope

doi: 10.1128/JVI.78.11.5812-5819.2004

Figure Lengend Snippet: Summary of virus sensitivity and cell surface molecule expression

Article Snippet: Samples were then incubated on ice with either primary mouse anti-human CD55 antibody (BRIC110) or mouse anti-human CD46 antibody (J4-48) from Cymbus Bioscience Ltd., diluted to 1:20 in PBS/BA, for 1 h. After three washes in PBS/BA, samples were incubated with fluorescein isothiocyanate (FITC)-conjugated anti-mouse secondary antibody (Jackson ImmunoResearch), diluted to 1:200 in PBS/BA and incubated on ice for 1 h. For α-Gal expression, cells were detached by a cell scraper, washed, resuspended, and stained in a single 1-h incubation with FITC-conjugated Bandeiraea simplicifolia isolectin (IB-4) (Sigma) at 10 μg/ml in PBS/BA on ice, as this lectin is specific for the terminal α-Gal sugar.

Techniques: Expressing

Journal:

Article Title: Reduced Sensitivity to Human Serum Inactivation of Enveloped Viruses Produced by Pig Cells Transgenic for Human CD55 or Deficient for the Galactosyl-?(1-3) Galactosyl Epitope

doi: 10.1128/JVI.78.11.5812-5819.2004

Figure Lengend Snippet: Demonstration of hCD55 incorporation on VSV particles by a viral pull-down assay. VSV harvested through HeLa, TgPAE A, and PAE E cells in the presence of antibody was incubated with protein G-expressing bacterial cells (OMNISORB). Antibodies used were three anti-human CD55 antibodies, BRIC 216, BRIC 471, and anti-DAF; anti-human CD46 J4-48 (hCD46); and anti-CD59 BRA-10G (hCD59). Five micrograms of anti-DAF and 1 μg of the other antibodies were used for incubation with virus and protein G cells. Titers of VSV for pulled-down and input particles were determined by TCID50 assay. The ratio of these titers is shown as percent binding.

Article Snippet: Samples were then incubated on ice with either primary mouse anti-human CD55 antibody (BRIC110) or mouse anti-human CD46 antibody (J4-48) from Cymbus Bioscience Ltd., diluted to 1:20 in PBS/BA, for 1 h. After three washes in PBS/BA, samples were incubated with fluorescein isothiocyanate (FITC)-conjugated anti-mouse secondary antibody (Jackson ImmunoResearch), diluted to 1:200 in PBS/BA and incubated on ice for 1 h. For α-Gal expression, cells were detached by a cell scraper, washed, resuspended, and stained in a single 1-h incubation with FITC-conjugated Bandeiraea simplicifolia isolectin (IB-4) (Sigma) at 10 μg/ml in PBS/BA on ice, as this lectin is specific for the terminal α-Gal sugar.

Techniques: Pull Down Assay, Incubation, Expressing, TCID50 Assay, Binding Assay