Protein enrichment is essential for biological samples that contain low protein concentrations, especially for proteomic studies that require sufficient quantities for subsequent MS analysis. Traditional precipitation methods, however, are limited in the sample volume and protein concentration required to cause efficient precipitations. We showed that gold nanoparticles (Au-NPs) can be easily applied to concentrate proteins from more than 15 mL of human urine, in which the total protein concentration is less than 1.4 ppm. Moreover, Au-NP-aggregated proteins can be directly applied to gel electrophoresis for Au-NP-protein dissociation followed by free protein separation as well as for the subsequent in-gel digestion and protein identification by mass spectrometry. We compared this method with trichloroacetic acid (TCA) precipitation method, one of the most common precipitation methods, and TCA method showed no enrichment effect for protein samples with large volumes (>2 mL) or with low protein concentrations (4 ppm). Therefore, Au-NP aggregation is not only a simple and efficient method for enriching a broad range of proteins, it is also particularly useful for concentrating proteins from a relatively large volume of dilute biological fluids, under which TCA method is ineffective.
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