Technical Reference #1724

1724.

The C type natriuretic peptide receptor tethers AHNAK1 at the plasma membrane to potentiate arachidonic acid-induced calcium mobilization Abdel A. Alli; William R. Gower; Jr., University of South Florida, American Journal of Physiology: Cell Physiology, 297(1724), (2009)
Link To Paper

Keywords:
3T3-L1; smooth muscle cells; rat gastric muscosa cell 1; phospholipase C

Materials & Methods:
Reagents and antibodies. Glass-bottom culture dishes were purchased from MatTek (Ashland MA). All other cell culture plastic ware was purchased from Nalge Nunc (Rochester NY). Precast gels (7.5% resolving with 4% stacking) were purchased from Bio-Rad (Hercules CA). Protein G agarose was purchased from Upstate Biotechnologies (Lake Placid NY). Mammalian protein extraction reagent (M-PER) Halt protease and phosphatase inhibitor Super Signal West Pico chemiluminescent substrate and BCA reagents were purchased from Pierce (Rockford IL). Fluo-3 AM dye was purchased from Molecular Probes (Eugene OR). All other chemicals were purchased from Sigma (St. Louis MO) unless otherwise noted. G I antibody was kindly provided by Dr. Dave Manning (University of Pennsylvania Philadelphia PA); NPR-A and NPR-B polyclonal antibodies were kindly provided by Dr. David L. Garbers (University of Texas Southwestern Dallas TX); polyclonal AHNAK antibody was kindly provided by Dr. Hannelore Haase (Max Delbru¨ck Center Berlin Germany); monoclonal AHNAK antibody was purchased from Abnova (Taipei City Taiwan); -actin antibody was purchased from Sigma; and PLC 3 PLC 1 PLC 2 GAPDH Lamin A/C and peroxisome proliferator-activated receptor- (PPAR ) antibodies were purchased from Cell Signaling Technology (Danvers MA). Anti-NPR-C antibody (JAH84) was from our laboratory. Hybond-C extra nitrocellulose membranes and Hyperfilm ECL were purchased from Amersham Biosciences (Piscataway NJ).

Microscopic Technique
Confocal Fluorescence Microscopy

Cell Type(s)
3T3-L1; HeLa cells; AoSMC cells; RGM1 cells