Nucleoplasm, also referred to as karyoplasm/nucleus sap, consists of chromosomes, nucleolus and nuclear matrix. Nucleoplasm plays a central role in genetic information flow and regulation of gene expression. Nuclear protein isolation has traditionally been accomplished by extraction protocol using detergent-containing buffers, which is effective in extracting total nuclear proteins but unable to separate membrane-bound proteins and water-soluble matrix proteins. The detergent-free nuclear matrix protein extraction kit is designed to separate cellular protein into three fractions: cytosolic, nuclear matrix and water-insoluble fractions (mainly envelope and nucleic acid-associated proteins) using a patented spin-column-based technology. The buffers used are detergent and EDTA-free. The whole protocol can be completed in less than 1 hour. The proteins extracted are in their native state and can be used in many downstream applications such as gel retardation, transcription factor analysis, apoptosis and protein trafficking studies.