Free cholesterol and lipid raft content of normal and NPCD55 cells. (A) Cellular cholesterol was measured in uninfected and HIV-1-infected Normal (filled) and HIV-1-infected NPCD55 (slashed) cells by AmplexRed assay. Cholesterol concentration was normalized to total protein. Error bars represent triplicate experiments. Values are relative to the normal uninfected control which was arbitrarily set to 1. (B) Virus produced from each cell type was concentrated by centrifugation through a 20% sucrose cushion. The cholesterol content of the virus was measured by AmplexRed assay and input was normalized by p24 concentration. (C) Lipid raft fractions were collected for normal and NPCD55 cells and loaded for dot blot analysis. Membranes were probed with cholera toxin B to detect GM1. The samples were normalized by protein concentration and volume. (D) Dot intensities for the blot shown were quantified using ChemiDoc XRS and Quantity One software.