For quantification of surface receptor expression, cells were trypsinized and brought to single-cell suspension in FACS buffer (1× PBS + 2% FBS). Primary antibody was added at 1 μg/million cells in 100 μL total solution (for mouse cells: rat anti-mouse αv integrin (BD Pharmingen 551380) or rat IgG isotype control (Invitrogen); for human cells: mouse anti-αvβ3 integrin, direct PE conjugate (BioLegend 304406) or mouse IgG κ chain isotype control, direct PE conjugate (BioLegend 400112); for neuropilin-1 staining in all cells, rabbit anti-NRP-1 (Novus Biologicals NBP1-40666) or normal rabbit IgG isotype control (R&D)) and incubated for one hour on ice. For direct fluorophore-conjugated primary antibodies, cells were washed with PBS and resuspended in FACS buffer. Otherwise, after washing the cells 2× in PBS, cells were incubated with secondary fluorescently-tagged antibody (Invitrogen) for 45 minutes and washed 1× in PBS. Cells were analyzed on BD LSR-II or Fortessa HTS flow cytometers. Data were analyzed in FlowJo (TreeStar Software).
Normal rabbit igg isotype control
Manufactured by R&D Systems
Sourced in Japan
Normal rabbit IgG isotype control is a non-specific antibody that is used as a control in immunoassays and other applications where the use of an isotype-matched control is required. It is derived from the serum of normal rabbits and does not recognize any known antigenic target.
Automatically generated - may contain errors
Lab products found in correlation
Direct pe conjugate, by BioLegend (1 mentions)
Rat igg isotype control, by Thermo Fisher Scientific (1 mentions)
Goat anti rabbit igg hrp conjugate, by Thermo Fisher Scientific (1 mentions)
Mouse anti desmin, by Agilent Technologies (1 mentions)
Rabbit anti β actin, by BioLegend (1 mentions)
Mouse anti vimentin, by Agilent Technologies (1 mentions)
2 protocols using normal rabbit igg isotype control
1
Quantifying Surface Receptor Expression
2
Immunocytometry and Western Blotting Antibodies
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The following antibodies (Abs) were used in immunocytometry and immunohistochemistry: rabbit anti-von Willebrand factor (Factor VIII) (1:200, DAKO, Carpinterria, CA, USA), mouse anti-CD31 Ab (1:200, Abcam, Tokyo, Japan), mouse anti-Vimentin (1:200, Dako, CA, USA), mouse anti-Desmin (1:100, DAKO, Carpinterria, CA, USA), rabbit anti-hepatocyte growth factor (HGF) Ab (1:200, Abcam, Tokyo, Japan), rabbit anti-Met Ab (1:200, Abcam, Tokyo, Japan), normal Rabbit IgG isotype control (1:200, R&D Systems, Minneapolis, MN, USA), goat anti-rabbit IgG conjugated with Alexa-488 fluorescent dye (1:200, Thermofisher Scientific, Tokyo, Japan) and goat anti-mouse IgG conjugated with Alexa-594 fluorescent dye (1:200, Thermofisher Scientific, Tokyo, Japan). These antibodies were dissolved in PBS-T.
The following Abs were used in Western blotting: rabbit anti-βactin (1:1000, Biolegend, Tokyo, Japan), rabbit anti-HGF (1:1000, Abcam, Tokyo, Japan) and Goat anti-Rabbit IgG HRP conjugate (1:1000, Thermoscientific, Tokyo, Japan). These antibodies were dissolved with blocking solution (5% non-fat dried milk in PBS containing 0.5% Tween20).
The following Abs were used in Western blotting: rabbit anti-βactin (1:1000, Biolegend, Tokyo, Japan), rabbit anti-HGF (1:1000, Abcam, Tokyo, Japan) and Goat anti-Rabbit IgG HRP conjugate (1:1000, Thermoscientific, Tokyo, Japan). These antibodies were dissolved with blocking solution (5% non-fat dried milk in PBS containing 0.5% Tween20).
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