Immunoblotting Analysis of Filamin B

MEFs were cultured with or without PMA in serum-starved condition overnight. Flnb^+/+ and Flnb^−/− tumor tissues were homogenized and immunoblotted as described earlier,^{10 (link)} followed by incubation with primary antibody against FLNB (Chemicon International, Inc., Temecula, CA, USA), FLNA (EMD Millipore), Actin (Sigma Aldrich), phosphorylated ERK1/2, total ERK1/2, phosphorylated-PKC-α/β and phosphorylated-PKC-δ/θ (Cell Signaling Technologies, Danvers, MA, USA) and MMP-9 (Nordic BioSite, Taby, Sweden). Membranes were then incubated with an appropriate anti-mouse or anti-rabbit IgG-horseradish peroxide conjugated secondary antibody (Amersham Biosciences, Amersham, UK) and the proteins were visualized by using enhanced chemiluminescence (Amersham Biosciences). The densities of bands were quantified by ImageQuant software (Bio-Rad, Hercules, CA, USA). Actin was used as an internal loading control.

Partial Protocol Preview
This section provides a glimpse into the protocol.
The remaining content is hidden due to licensing restrictions, but the full text is available at the following link: Access Free Full Text.

Bandaru S., Zhou A.X., Rouhi P., Zhang Y., Bergo M.O., Cao Y, & Akyürek L.M. (2014). Targeting filamin B induces tumor growth and metastasis via enhanced activity of matrix metalloproteinase-9 and secretion of VEGF-A. Oncogenesis, 3(9), e119-.

Publication 2014

phosphorylated ERK1/2 total ERK1/2 enhanced chemiluminescence ImageQuant software

Actin Anti igg Antibody Chemiluminescence Erk1 Horseradish Membranes Mmp 9 Mouse Peroxide Pkc α Proteins Rabbit Serum Tissues Tumor

Corresponding Organization :

Other organizations : University of Gothenburg, Karolinska Institutet, Sahlgrenska University Hospital

Top 5 similar protocols

Variable analysis

independent variables

Presence or absence of PMA in serum-starved condition

dependent variables

Levels of FLNB, FLNA, Actin, phosphorylated ERK1/2, total ERK1/2, phosphorylated-PKC-α/β, phosphorylated-PKC-δ/θ, and MMP-9 in Flnb+/+ and Flnb-/- tumor tissues

control variables

Actin as an internal loading control

controls

Flnb+/+ and Flnb-/- tumor tissues as control groups

Annotations

Based on most similar protocols

Etiam vel ipsum. Morbi facilisis vestibulum nisl. Praesent cursus laoreet felis. Integer adipiscing pretium orci. Nulla facilisi. Quisque posuere bibendum purus. Nulla quam mauris, cursus eget, convallis ac, molestie non, enim. Aliquam congue. Quisque sagittis nonummy sapien. Proin molestie sem vitae urna. Maecenas lorem.

As authors may omit details in methods from publication, our AI will look for missing critical information across the 5 most similar protocols.

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!