Colloidal gold--low density lipoprotein conjugates as membrane receptor probes.

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Abstract
A method was developed for conjugating low density lipoproteins (LDL) with colloidal Au. Conjugation, complete after 1 min, occurs by electrostatic adsorption of the LDL to the negatively charged Au particle. Each conjugate consists of approximately 8 biologically active LDL molecules clustered around a central 19-mm Au granule. Acidic (pH 4), alkaline (pH 9) or high ionic (600 milliosmolar NaCl) environments do not dissociate the conjugate. Colloidal Au is an electron-dense, nondegradable marker that is easily identified within the cell and serves as a valuable probe for studying receptor binding and endocytosis. By using a modified method of ruthenium red staining, the LDL molecules of the conjugate can be directly visualized when they are bound to the cell surface receptor. Receptor binding (4.degree. C) of the conjugate by cultured human fibroblasts reveals that the Au granule is positioned 18-21 nm from the coated pit region of the membrane. This distance, similar to the diameter of LDL, suggests concomitant internalization of the receptor during vesicular endocytosis and early lysosomal incorporation (10 min at 37.degree. C). Continued internalization (30-60 min at 37.degree. C) results in the formation of free pools of Au within the lysosome.

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