Journal of Lipid Research, Vol 28, 108-112, Copyright © 1987 by Lipid Research, Inc.
A dot-blot assay for the low density lipoprotein receptor
FM Maggi and AL Catapano
We describe a new method for detecting the interaction of low density
lipoprotein with its receptor using unmodified nitrocellulose as support
for membrane protein. The method is specific and sensitive down to 3
micrograms of membrane protein. Unlabeled LDL, but not HDL, competes with
125I-labeled LDL for binding, and binding is abolished by pretreatment of
the membranes with pronase and is dependent upon the presence of Ca2+.
Furthermore, modification of arginine or lysine residues on LDL abolishes
the lipoprotein interaction with the receptor protein supported on the
nitrocellulose. When the membranes are solubilized with octyl glucoside,
purification steps of the receptor can be directly followed with no
interference of the detergent, therefore eliminating the need for its
removal. The increased expression of LDL receptors on liver membranes from
estradiol-treated rats was also demonstrated. We suggest, therefore, that
this method can be used to detect the presence of LDL receptors on minute
amounts of membrane protein.
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