GBM cells were plated on Lab-Tek® Chamber Slide coated with laminin (1 μg/cm2). Cells were fixed in 4% paraformaldehyde (pH 7.4) for 30 mins, washed 3 times and blocked with 10% normal donkey serum in PBS for 1 h. Subsequently, fixed cells were incubated with primary antibody at 4 °C overnight. Primary antibodies include: Cofilin 1 (ref: 5175, Cell signaling), Beta1 Sodium Potassium ATPase (ref: ab2873, AbCAM) and NKCC1 (ref: 8351, Cell signaling). After primary antibody, slides where incubated with Alexa Fluor-conjugated 594 and 488 secondary antibodies (1:500, Invitrogen), counter stained with DAPI and mounted using Aquamount (VWR). The cells were imaged using an Olympus IX81 epifluorescence microscope. Co-localization analysis was performed using Image J Colocalization Colormap plugin as previously described (Jaskolski et al., 2005 (link)).
Nkcc1
Manufactured by Cell Signaling Technology
NKCC1 is a protein that functions as a sodium-potassium-chloride cotransporter. It is involved in the movement of these ions across cell membranes, which is important for various physiological processes.
Automatically generated - may contain errors
2 protocols using nkcc1
1
Immunocytochemistry of GBM cells
2
Lung Protein Extraction and Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Nuclear and cytoplasmic proteins were extracted from the lungs using a nuclear/cytosol extraction kit (BioVision, Inc., Mountain View, CA). The blots were incubated with antibodies against phosphorylated nuclear factor-κB (NF-κB) p65, inhibitor of NF-κB alpha (IκB-α), and NKCC1 (Cell Signaling Technology, Danvers, MA) overnight at 4 °C. Enhanced chemiluminescence reagents were used to visualize the bands, that were then exposed to radiography films. The blots were stripped using an anti-TATA antibody or anti-β-actin antibody to ensure equal loading.
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