Method |
A Chinese hamster lung fibroblast cell line (PS120) previously
selected to lack all endogenous NHEs was stably
transfected, using Lipofectin (GIBCO BRL), with human
NHE1 or rabbit NHE2 or NHE3 cDNAs epitope tagged on the
COOH termini with a previously described 11-amino acid
epitope of vesicular stomatitis virus glycoprotein (VSVG) plus
an 8-amino acid spacer (NHE1V, NHE2V, and NHE3V). Researchers then determined the turnover
numbers of NHE1, NHE2, and NHE3 expressed in the
same cell type (PS120) by correlating the Vmax (as
measured by 22Na+ uptake kinetic studies and fluorometry)
with the amount of NHE protein expressed at the
cell surface. |