Structured Review

The Jackson Laboratory biotinylated secondary antibodies
Effect of WNK2 and WNK3 on the protein level and surface expression of NKCC1 and KCC4. Western blot analysis of the total and <t>biotinylated</t> fraction of proteins extracted from oocytes injected with NKCC1 ( A ) or KCC4 ( B ) cRNA in the absence or presence of WNK2 or WNK3 cRNA. C and D depict the results of the functional expression performed the same day using oocytes from the same batch for NKCC1 or KCC4, respectively, expressed as bumetanide-sensitive ( C ) or chloride-dependent ( D ) 86 Rb + uptake.
Biotinylated Secondary Antibodies, supplied by The Jackson Laboratory, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/biotinylated secondary antibodies/product/The Jackson Laboratory
Average 86 stars, based on 1 article reviews
Price from $9.99 to $1999.99
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Images

1) Product Images from "WNK2 Kinase Is a Novel Regulator of Essential Neuronal Cation-Chloride Cotransporters *"

Article Title: WNK2 Kinase Is a Novel Regulator of Essential Neuronal Cation-Chloride Cotransporters *

Journal: The Journal of Biological Chemistry

doi: 10.1074/jbc.M111.222893

Effect of WNK2 and WNK3 on the protein level and surface expression of NKCC1 and KCC4. Western blot analysis of the total and biotinylated fraction of proteins extracted from oocytes injected with NKCC1 ( A ) or KCC4 ( B ) cRNA in the absence or presence of WNK2 or WNK3 cRNA. C and D depict the results of the functional expression performed the same day using oocytes from the same batch for NKCC1 or KCC4, respectively, expressed as bumetanide-sensitive ( C ) or chloride-dependent ( D ) 86 Rb + uptake.
Figure Legend Snippet: Effect of WNK2 and WNK3 on the protein level and surface expression of NKCC1 and KCC4. Western blot analysis of the total and biotinylated fraction of proteins extracted from oocytes injected with NKCC1 ( A ) or KCC4 ( B ) cRNA in the absence or presence of WNK2 or WNK3 cRNA. C and D depict the results of the functional expression performed the same day using oocytes from the same batch for NKCC1 or KCC4, respectively, expressed as bumetanide-sensitive ( C ) or chloride-dependent ( D ) 86 Rb + uptake.

Techniques Used: Expressing, Western Blot, Injection, Functional Assay

Related Articles

other:

Article Title: CD34+ Testicular Stromal Cells Support Long-Term Expansion of Embryonic and Adult Stem and Progenitor Cells
Article Snippet: The next day, cells were washed with PBS before incubating for 2 hours at room temperature with directly conjugated secondary antibodies or biotinylated secondary antibodies (Jackson Laboratory).

Article Title: CD34+ Testicular Stromal Cells Support Long-Term Expansion of Embryonic and Adult Stem and Progenitor Cells
Article Snippet: Biotinylated secondary antibodies were detected with either streptavidin-Alexa488 or streptavidin conjugated to horseradish peroxidase (HRP) (Jackson Laboratory).

Article Title: Brain radiation injury leads to a dose- and time-dependent recruitment of peripheral myeloid cells that depends on CCR2 signaling
Article Snippet: Biotinylated secondary antibodies utilized included goat anti-Armenian hamster IgG (Jackson Laboratories, 1:1000); goat anti-mouse F(ab′)2 (Jackson Laboratories, 1:2000); goat anti-rabbit IgG (Vector Laboratories, 1:1000); and goat anti-rat IgG (Vector Laboratories, 1:2000).

Article Title: Cranial Irradiation Leads to Acute and Persistent Neuroinflammation with Delayed Increases in T-Cell Infiltration and CD11c Expression in C57BL/6 Mouse Brain
Article Snippet: Biotinylated secondary antibodies used included goat anti-Armenian Hamster IgG (Jackson Laboratory, 1:1000); goat anti-mouse F(ab′)2 (Jackson Laboratory, 1:2000); goat anti-rabbit IgG (Vector Laboratory, 1:1000); and goat anti-rat IgG (Vector Laboratories, 1:2000).

Incubation:

Article Title: Oxaliplatin-induced loss of phosphorylated heavy neurofilament subunit neuronal immunoreactivity in rat DRG tissue
Article Snippet: Each slide was incubated overnight in a humidity chamber in both mouse anti-pNF-H (1:100) and rabbit anti-parvalbumin (1:1000) primary antibodies. .. The slides were then washed and incubated in the dark for 4 hours in both anti-rabbit cy3 (1:200; Jackson Laboratories, West Grove, PA, USA) and biotinylated anti-mouse secondary antibodies (1:200, Sigma). .. The slides were rewashed and incubated in the dark for 3 hours in FITC tertiary antibody (1:200; Sigma-Aldrich).

Article Title: MKP-1 overexpression is associated with chemoresistance in bladder cancer via the MAPK pathway
Article Snippet: Primary rabbit polyclonal anti-MKP-1 antibodies (dilution: 1:500, catlog: NBP2-67909, Novus Biologicals, LLC) were then added and incubated overnight at 4°C. .. After being washed three times, samples were further incubated with biotinylated secondary antibodies (the Jackson laboratory, Bar Harbor, ME, USA) for 30 min at room temperature. ..

Article Title: Distinct Behaviors of Neural Stem and Progenitor Cells Underlie Cortical Neurogenesis
Article Snippet: Sections were rinsed in PBS 0.1 M, pH 7.4, incubated overnight in antibody buffer containing 2% serum, 0.1% Triton X, and 0.2% gelatin diluted in PBS. .. Sections were rinsed, incubated with biotinylated secondary antibodies (1:100, Jackson Laboratories, West Grove, PA) for 1 hour at room temperature (RT). .. Sections were stained with avidin-biotin complex (Vector, Burlingame, CA) for 1 hour at RT, rinsed, and placed in 0.04% DAB (Sigma) with 0.001% H2 O2 for 2–5 minutes.

Plasmid Preparation:

Article Title: Brachyury proteins regulate target genes through modular binding sites in a cooperative fashion
Article Snippet: .. Biotinylated secondary antibodies (goat, Jackson Laboratories) and the Vectastain ABC Elite enhancement kit (Vector) were used. ..

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    The Jackson Laboratory anti mouse bdnf secondary antibody
    Representative photomicrographs showing immunohistochemistry for Arc (A, B), <t>c-fos</t> (C, D) and <t>BDNF</t> (E, F) from the AC of group reared males. A, C, and E are from rats that were not exposed to a novel conspecific and B, D, and F are from rats that were
    Anti Mouse Bdnf Secondary Antibody, supplied by The Jackson Laboratory, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti mouse bdnf secondary antibody/product/The Jackson Laboratory
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    86
    The Jackson Laboratory biotinylated goat anti rabbit secondary antibody
    RGS3 proteins can be overexpressed in chick sensory neurons. ( a ) Schematic diagram of the primary structure of RGS3s and RGS3ss splice variants. Marked are EF-like (black), RGS (gray), and CaM-binding (cross-hatched) domains. RGS3ss lacks the first 125 aa of RGS3s, including the EF-like region. ( b ) Frozen sections of intact dorsal root ganglia from 12-day-old embryos, 10.5 days after infection with either empty retroviral vector (“virus control”) or retroviral vectors carrying HA-tagged RGS3s (“RGS3s”). Sections were labeled with anti-rabbit HA primary antiserum followed by <t>biotinylated</t> goat anti-rabbit secondary antiserum and FITC-conjugated streptavidin. The low-intensity staining of cells infected with empty virus represents background fluorescence (comparable with sections labeled with secondary antiserum in the absence of primary antiserum). (Calibration bar: 20 μm.) ( c ) Representative families of superimposed calcium currents evoked by 35-ms step depolarizations to potentials between -50 and 50 mV in wild-type DRG neurons ( Left ) or cells infected with viral vector alone ( Right ). (Calibration: 2 nA, 10 ms.) ( d ) Current–voltage relationships (means ± SEMs) for calcium currents recorded from uninfected DRG neurons ( Left , circles, n = 18), vector alone-infected neurons (triangles, n = 10), or neurons expressing recombinant RGS3s ( Right , diamonds, n = 12) or RGS3ss ( Right , squares, n = 12).
    Biotinylated Goat Anti Rabbit Secondary Antibody, supplied by The Jackson Laboratory, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/biotinylated goat anti rabbit secondary antibody/product/The Jackson Laboratory
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    biotinylated goat anti rabbit secondary antibody - by Bioz Stars, 2021-03
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    86
    The Jackson Laboratory biotinylated goat anti human secondary ab
    Design and characterization of HCDR3-grafted Ubq fusion proteins. ( A ) A ribbon representation of Ubq (1UBQ.pdb [106] ) is shown at bottom right, colored prismatically from N- (blue) to C-terminus (red); the targeted loop for HCDR3 engraftment is highlighted in gray with red bounding residues. The structures of the three targeted HCDR3 segments in context of their parent Abs are shown above in a licorice-stick representation, colored by atom type (oxygen: red; nitrogen: blue; carbons of buried residues: marine blue; and carbons of exposed residues: gray). Residue exposure was determined with the program GetArea [107] , with greater than 30% solvent accessibility considered “exposed”, based on the structural context in the corresponding crystal structures (2F5: 3IDG.pdb [108] ; b12: 3RU8.pdb [104] ; 4E10: 2FX7.pdb [9] ). The three transferred sequences (2F5: AHRRGP T T LFGVPIARG PVNAMDVW; b12: ARVG PYSWDDSP QYNYYMDVW; 4E10: AREGTT GWGWLGKP IGAFAHW; exposed residues bolded ), were characterized as “hydrophobic” with the Sigma-Aldrich PEPscreen Library Design Tool (Φ 2F5 = 0.52; Φ b12 = 0.51; Φ 4E10 = 0.56). Key residues and Cα-Cα distances (Å) are indicated. The inset shows a schematic representation of the design process. Molecular images were generated with MacPyMOL [109] . ( B ) SEC analyses of the solution properties of the purified recombinant HCDR3-grafted Ubq fusion proteins are shown, confirming monodispersivity. ( C ) Corrected SPR responses (duplicate runs) of HCDR3-grafted Ubq fusion proteins to HIV SF162 gp140 protein amine-coupled to biosensor chips; analyte concentrations are indicated. While detectable, these responses are consistent with K D values > > 10 µM and are thus too weak to quantify reliably. ( D ) SPR responses (duplicate runs) of Annexin V and HCDR3-grafted Ubq fusion protein analytes to liposomes incorporating <t>biotinylated</t> lipids captured on streptavidin-coated biosensor chips are shown, with liposome compositions indicated above each frame. HCDR3-grafted Ubq fusion protein responses are colored as indicated, but none showed detectable binding on any liposome composition.
    Biotinylated Goat Anti Human Secondary Ab, supplied by The Jackson Laboratory, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/biotinylated goat anti human secondary ab/product/The Jackson Laboratory
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    biotinylated goat anti human secondary ab - by Bioz Stars, 2021-03
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    86
    The Jackson Laboratory biotinylated secondary antibodies
    Effect of WNK2 and WNK3 on the protein level and surface expression of NKCC1 and KCC4. Western blot analysis of the total and <t>biotinylated</t> fraction of proteins extracted from oocytes injected with NKCC1 ( A ) or KCC4 ( B ) cRNA in the absence or presence of WNK2 or WNK3 cRNA. C and D depict the results of the functional expression performed the same day using oocytes from the same batch for NKCC1 or KCC4, respectively, expressed as bumetanide-sensitive ( C ) or chloride-dependent ( D ) 86 Rb + uptake.
    Biotinylated Secondary Antibodies, supplied by The Jackson Laboratory, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/biotinylated secondary antibodies/product/The Jackson Laboratory
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    biotinylated secondary antibodies - by Bioz Stars, 2021-03
    86/100 stars
      Buy from Supplier

    Image Search Results


    Representative photomicrographs showing immunohistochemistry for Arc (A, B), c-fos (C, D) and BDNF (E, F) from the AC of group reared males. A, C, and E are from rats that were not exposed to a novel conspecific and B, D, and F are from rats that were

    Journal: Physiology & behavior

    Article Title: Isolation rearing attenuates social interaction-induced expression of immediate early gene protein products in the medial prefrontal cortex of male and female rats

    doi: 10.1016/j.physbeh.2012.09.002

    Figure Lengend Snippet: Representative photomicrographs showing immunohistochemistry for Arc (A, B), c-fos (C, D) and BDNF (E, F) from the AC of group reared males. A, C, and E are from rats that were not exposed to a novel conspecific and B, D, and F are from rats that were

    Article Snippet: Sections were then incubated in biotinylated goat anti-rabbit (Arc, c-fos) or anti-mouse (BDNF) secondary antibody (1:200, Jackson Labs) for 2 h, followed by incubation in avidin biotin complex (ABC kit, Vector Laboratories) for 2 h. Sections were washed 3 times in 0.1 M PB, then immunoreactivity was visualized with 3,3′-diaminobenzidine (DAB substrate kit, Vector Laboratories) and nickel ammonium sulphate as chromogens.

    Techniques: Immunohistochemistry

    Stereological estimates of cells positive for Arc (A), c-fos (B), and BDNF (C) immunoreactivity in the mPFC of male and female rats exposed to either group or isolation rearing. Rats then were subjected to either a 15 minute exposure to a novel same-sex

    Journal: Physiology & behavior

    Article Title: Isolation rearing attenuates social interaction-induced expression of immediate early gene protein products in the medial prefrontal cortex of male and female rats

    doi: 10.1016/j.physbeh.2012.09.002

    Figure Lengend Snippet: Stereological estimates of cells positive for Arc (A), c-fos (B), and BDNF (C) immunoreactivity in the mPFC of male and female rats exposed to either group or isolation rearing. Rats then were subjected to either a 15 minute exposure to a novel same-sex

    Article Snippet: Sections were then incubated in biotinylated goat anti-rabbit (Arc, c-fos) or anti-mouse (BDNF) secondary antibody (1:200, Jackson Labs) for 2 h, followed by incubation in avidin biotin complex (ABC kit, Vector Laboratories) for 2 h. Sections were washed 3 times in 0.1 M PB, then immunoreactivity was visualized with 3,3′-diaminobenzidine (DAB substrate kit, Vector Laboratories) and nickel ammonium sulphate as chromogens.

    Techniques: Isolation

    RGS3 proteins can be overexpressed in chick sensory neurons. ( a ) Schematic diagram of the primary structure of RGS3s and RGS3ss splice variants. Marked are EF-like (black), RGS (gray), and CaM-binding (cross-hatched) domains. RGS3ss lacks the first 125 aa of RGS3s, including the EF-like region. ( b ) Frozen sections of intact dorsal root ganglia from 12-day-old embryos, 10.5 days after infection with either empty retroviral vector (“virus control”) or retroviral vectors carrying HA-tagged RGS3s (“RGS3s”). Sections were labeled with anti-rabbit HA primary antiserum followed by biotinylated goat anti-rabbit secondary antiserum and FITC-conjugated streptavidin. The low-intensity staining of cells infected with empty virus represents background fluorescence (comparable with sections labeled with secondary antiserum in the absence of primary antiserum). (Calibration bar: 20 μm.) ( c ) Representative families of superimposed calcium currents evoked by 35-ms step depolarizations to potentials between -50 and 50 mV in wild-type DRG neurons ( Left ) or cells infected with viral vector alone ( Right ). (Calibration: 2 nA, 10 ms.) ( d ) Current–voltage relationships (means ± SEMs) for calcium currents recorded from uninfected DRG neurons ( Left , circles, n = 18), vector alone-infected neurons (triangles, n = 10), or neurons expressing recombinant RGS3s ( Right , diamonds, n = 12) or RGS3ss ( Right , squares, n = 12).

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: RGS3 mediates a calcium-dependent termination of G protein signaling in sensory neurons

    doi: 10.1073/pnas.1231837100

    Figure Lengend Snippet: RGS3 proteins can be overexpressed in chick sensory neurons. ( a ) Schematic diagram of the primary structure of RGS3s and RGS3ss splice variants. Marked are EF-like (black), RGS (gray), and CaM-binding (cross-hatched) domains. RGS3ss lacks the first 125 aa of RGS3s, including the EF-like region. ( b ) Frozen sections of intact dorsal root ganglia from 12-day-old embryos, 10.5 days after infection with either empty retroviral vector (“virus control”) or retroviral vectors carrying HA-tagged RGS3s (“RGS3s”). Sections were labeled with anti-rabbit HA primary antiserum followed by biotinylated goat anti-rabbit secondary antiserum and FITC-conjugated streptavidin. The low-intensity staining of cells infected with empty virus represents background fluorescence (comparable with sections labeled with secondary antiserum in the absence of primary antiserum). (Calibration bar: 20 μm.) ( c ) Representative families of superimposed calcium currents evoked by 35-ms step depolarizations to potentials between -50 and 50 mV in wild-type DRG neurons ( Left ) or cells infected with viral vector alone ( Right ). (Calibration: 2 nA, 10 ms.) ( d ) Current–voltage relationships (means ± SEMs) for calcium currents recorded from uninfected DRG neurons ( Left , circles, n = 18), vector alone-infected neurons (triangles, n = 10), or neurons expressing recombinant RGS3s ( Right , diamonds, n = 12) or RGS3ss ( Right , squares, n = 12).

    Article Snippet: For immunocytochemical detection of infected cells, sections were first incubated 30 min with 0.25% gelatin (in PBS) to block nonspecific binding, followed by 1 h with rabbit anti-HA primary antiserum (1:500 dilution), 1 h with a biotinylated goat anti-rabbit secondary antibody (1:150 dilution, The Jackson Laboratory), and 40 min with FITC-conjugated streptavidin (1:1,000, The Jackson Laboratory).

    Techniques: Chick Chorioallantoic Membrane Assay, Binding Assay, Infection, Plasmid Preparation, Labeling, Staining, Fluorescence, Mass Spectrometry, Expressing, Recombinant

    Design and characterization of HCDR3-grafted Ubq fusion proteins. ( A ) A ribbon representation of Ubq (1UBQ.pdb [106] ) is shown at bottom right, colored prismatically from N- (blue) to C-terminus (red); the targeted loop for HCDR3 engraftment is highlighted in gray with red bounding residues. The structures of the three targeted HCDR3 segments in context of their parent Abs are shown above in a licorice-stick representation, colored by atom type (oxygen: red; nitrogen: blue; carbons of buried residues: marine blue; and carbons of exposed residues: gray). Residue exposure was determined with the program GetArea [107] , with greater than 30% solvent accessibility considered “exposed”, based on the structural context in the corresponding crystal structures (2F5: 3IDG.pdb [108] ; b12: 3RU8.pdb [104] ; 4E10: 2FX7.pdb [9] ). The three transferred sequences (2F5: AHRRGP T T LFGVPIARG PVNAMDVW; b12: ARVG PYSWDDSP QYNYYMDVW; 4E10: AREGTT GWGWLGKP IGAFAHW; exposed residues bolded ), were characterized as “hydrophobic” with the Sigma-Aldrich PEPscreen Library Design Tool (Φ 2F5 = 0.52; Φ b12 = 0.51; Φ 4E10 = 0.56). Key residues and Cα-Cα distances (Å) are indicated. The inset shows a schematic representation of the design process. Molecular images were generated with MacPyMOL [109] . ( B ) SEC analyses of the solution properties of the purified recombinant HCDR3-grafted Ubq fusion proteins are shown, confirming monodispersivity. ( C ) Corrected SPR responses (duplicate runs) of HCDR3-grafted Ubq fusion proteins to HIV SF162 gp140 protein amine-coupled to biosensor chips; analyte concentrations are indicated. While detectable, these responses are consistent with K D values > > 10 µM and are thus too weak to quantify reliably. ( D ) SPR responses (duplicate runs) of Annexin V and HCDR3-grafted Ubq fusion protein analytes to liposomes incorporating biotinylated lipids captured on streptavidin-coated biosensor chips are shown, with liposome compositions indicated above each frame. HCDR3-grafted Ubq fusion protein responses are colored as indicated, but none showed detectable binding on any liposome composition.

    Journal: PLoS Pathogens

    Article Title: Autoreactivity and Exceptional CDR Plasticity (but Not Unusual Polyspecificity) Hinder Elicitation of the Anti-HIV Antibody 4E10

    doi: 10.1371/journal.ppat.1003639

    Figure Lengend Snippet: Design and characterization of HCDR3-grafted Ubq fusion proteins. ( A ) A ribbon representation of Ubq (1UBQ.pdb [106] ) is shown at bottom right, colored prismatically from N- (blue) to C-terminus (red); the targeted loop for HCDR3 engraftment is highlighted in gray with red bounding residues. The structures of the three targeted HCDR3 segments in context of their parent Abs are shown above in a licorice-stick representation, colored by atom type (oxygen: red; nitrogen: blue; carbons of buried residues: marine blue; and carbons of exposed residues: gray). Residue exposure was determined with the program GetArea [107] , with greater than 30% solvent accessibility considered “exposed”, based on the structural context in the corresponding crystal structures (2F5: 3IDG.pdb [108] ; b12: 3RU8.pdb [104] ; 4E10: 2FX7.pdb [9] ). The three transferred sequences (2F5: AHRRGP T T LFGVPIARG PVNAMDVW; b12: ARVG PYSWDDSP QYNYYMDVW; 4E10: AREGTT GWGWLGKP IGAFAHW; exposed residues bolded ), were characterized as “hydrophobic” with the Sigma-Aldrich PEPscreen Library Design Tool (Φ 2F5 = 0.52; Φ b12 = 0.51; Φ 4E10 = 0.56). Key residues and Cα-Cα distances (Å) are indicated. The inset shows a schematic representation of the design process. Molecular images were generated with MacPyMOL [109] . ( B ) SEC analyses of the solution properties of the purified recombinant HCDR3-grafted Ubq fusion proteins are shown, confirming monodispersivity. ( C ) Corrected SPR responses (duplicate runs) of HCDR3-grafted Ubq fusion proteins to HIV SF162 gp140 protein amine-coupled to biosensor chips; analyte concentrations are indicated. While detectable, these responses are consistent with K D values > > 10 µM and are thus too weak to quantify reliably. ( D ) SPR responses (duplicate runs) of Annexin V and HCDR3-grafted Ubq fusion protein analytes to liposomes incorporating biotinylated lipids captured on streptavidin-coated biosensor chips are shown, with liposome compositions indicated above each frame. HCDR3-grafted Ubq fusion protein responses are colored as indicated, but none showed detectable binding on any liposome composition.

    Article Snippet: The Fab of a biotinylated goat anti-human secondary Ab (Jackson Laboratory 109-067-993) was applied for 30 minutes followed by incubation with streptavidin-HRP (Jackson Laboratory 016-030-084) at a dilution of 1∶2000 for 30 minutes.

    Techniques: Generated, Size-exclusion Chromatography, Purification, Recombinant, SPR Assay, Binding Assay

    SPR analyses of 4E10/liposome interactions. Corrected SPR responses are shown (response units: RU; duplicate runs) of Annexin V (black), αCL polyclonal Ab (orange), 4E10 IgG (red), 2F5 IgG (blue), and b12 IgG (green) analytes binding to liposomes incorporating biotinylated lipids captured on streptavidin-coated biosensor chips. All analyte concentrations were 300 nM. The Annexin V analyte buffer included 2.5 mM CaCl 2 . Liposome compositions are indicated above each frame.

    Journal: PLoS Pathogens

    Article Title: Autoreactivity and Exceptional CDR Plasticity (but Not Unusual Polyspecificity) Hinder Elicitation of the Anti-HIV Antibody 4E10

    doi: 10.1371/journal.ppat.1003639

    Figure Lengend Snippet: SPR analyses of 4E10/liposome interactions. Corrected SPR responses are shown (response units: RU; duplicate runs) of Annexin V (black), αCL polyclonal Ab (orange), 4E10 IgG (red), 2F5 IgG (blue), and b12 IgG (green) analytes binding to liposomes incorporating biotinylated lipids captured on streptavidin-coated biosensor chips. All analyte concentrations were 300 nM. The Annexin V analyte buffer included 2.5 mM CaCl 2 . Liposome compositions are indicated above each frame.

    Article Snippet: The Fab of a biotinylated goat anti-human secondary Ab (Jackson Laboratory 109-067-993) was applied for 30 minutes followed by incubation with streptavidin-HRP (Jackson Laboratory 016-030-084) at a dilution of 1∶2000 for 30 minutes.

    Techniques: SPR Assay, Binding Assay

    Analysis of the effects of salt concentration, mutation and overall cassette charge on 4E10/liposome interactions. Corrected SPR responses are shown for Annexin V or 4E10 IgG analytes (300 nM; duplicate runs) to liposomes incorporating biotinylated lipids captured on streptavidin-coated biosensor chips; liposome compositions are indicated above each frame. ( A ) SPR responses of wild-type 4E10 IgG analytes at different salt concentrations are plotted. ( B ) SPR responses of Annexin V (300 nM), wild-type 4E10 IgG (300 nM) or the 4E10 [G(L50)E] mutant IgG (400 nM) are plotted. A higher concentration of 4E10 mutant IgG was used with the expectation that binding might be significantly reduced. Since this did not occur, mutant IgG responses appear elevated due to the concentration differential. ( C ) The net charge at neutral pH of the Fv cassettes of the anti-HIV Abs with structures currently available through the PDB [103] was calculated with the structure-based algorithm PDB2PQ [51] – [54] . Two Abs, PG16 and NIH45-46, were excluded because their structures included modified amino acids that could not be accommodated by PDB2PQ. Fvs are plotted with their names, with assigned PDB accession codes in parentheses. Ab labels are colored by the locale of their epitopes on Env, as indicated; 4E10 is also bolded.

    Journal: PLoS Pathogens

    Article Title: Autoreactivity and Exceptional CDR Plasticity (but Not Unusual Polyspecificity) Hinder Elicitation of the Anti-HIV Antibody 4E10

    doi: 10.1371/journal.ppat.1003639

    Figure Lengend Snippet: Analysis of the effects of salt concentration, mutation and overall cassette charge on 4E10/liposome interactions. Corrected SPR responses are shown for Annexin V or 4E10 IgG analytes (300 nM; duplicate runs) to liposomes incorporating biotinylated lipids captured on streptavidin-coated biosensor chips; liposome compositions are indicated above each frame. ( A ) SPR responses of wild-type 4E10 IgG analytes at different salt concentrations are plotted. ( B ) SPR responses of Annexin V (300 nM), wild-type 4E10 IgG (300 nM) or the 4E10 [G(L50)E] mutant IgG (400 nM) are plotted. A higher concentration of 4E10 mutant IgG was used with the expectation that binding might be significantly reduced. Since this did not occur, mutant IgG responses appear elevated due to the concentration differential. ( C ) The net charge at neutral pH of the Fv cassettes of the anti-HIV Abs with structures currently available through the PDB [103] was calculated with the structure-based algorithm PDB2PQ [51] – [54] . Two Abs, PG16 and NIH45-46, were excluded because their structures included modified amino acids that could not be accommodated by PDB2PQ. Fvs are plotted with their names, with assigned PDB accession codes in parentheses. Ab labels are colored by the locale of their epitopes on Env, as indicated; 4E10 is also bolded.

    Article Snippet: The Fab of a biotinylated goat anti-human secondary Ab (Jackson Laboratory 109-067-993) was applied for 30 minutes followed by incubation with streptavidin-HRP (Jackson Laboratory 016-030-084) at a dilution of 1∶2000 for 30 minutes.

    Techniques: Concentration Assay, Mutagenesis, SPR Assay, Binding Assay, Modification

    Effect of WNK2 and WNK3 on the protein level and surface expression of NKCC1 and KCC4. Western blot analysis of the total and biotinylated fraction of proteins extracted from oocytes injected with NKCC1 ( A ) or KCC4 ( B ) cRNA in the absence or presence of WNK2 or WNK3 cRNA. C and D depict the results of the functional expression performed the same day using oocytes from the same batch for NKCC1 or KCC4, respectively, expressed as bumetanide-sensitive ( C ) or chloride-dependent ( D ) 86 Rb + uptake.

    Journal: The Journal of Biological Chemistry

    Article Title: WNK2 Kinase Is a Novel Regulator of Essential Neuronal Cation-Chloride Cotransporters *

    doi: 10.1074/jbc.M111.222893

    Figure Lengend Snippet: Effect of WNK2 and WNK3 on the protein level and surface expression of NKCC1 and KCC4. Western blot analysis of the total and biotinylated fraction of proteins extracted from oocytes injected with NKCC1 ( A ) or KCC4 ( B ) cRNA in the absence or presence of WNK2 or WNK3 cRNA. C and D depict the results of the functional expression performed the same day using oocytes from the same batch for NKCC1 or KCC4, respectively, expressed as bumetanide-sensitive ( C ) or chloride-dependent ( D ) 86 Rb + uptake.

    Article Snippet: Following washes with PBS, they were incubated with biotinylated secondary antibodies (Jackson Laboratories, 1:250 dilution) for 2 h at room temperature, washed in PBS, and then incubated with Vectastain ABC Elite solution (Vector Laboratories, Burlingame, CA) for 2 h. Sections were developed using 0.05% DAB (pH 7.4), 0.2% glucose, 0.01% nickel ammonium sulfate, 0.04% ammonium chloride, and 8 mg/ml glucose oxidase, and then rinsed, mounted onto glass slides, allowed to dry, dehydrated, and coverslipped.

    Techniques: Expressing, Western Blot, Injection, Functional Assay