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Qproteome Nuclear Protein Kit

For separation of nuclear and nucleic acid binding proteins

  • Nuclear preparations free of cytosolic proteins
  • Greatly reduced sample complexity
  • Highly reproducible fractionation
  • Includes protocols for tissue proteomics

The Qproteome Nuclear Protein Kit is designed for specific enrichment of nuclear proteins from cultured mammalian cells.

Cat No./ID: 37582
Qproteome Nuclear Protein Kit
€418.00
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For 12 nuclear protein preparations: Buffers, Reagents, Protease Inhibitor Solution, Benzonase
The Qproteome Nuclear Protein Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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Isolation of an active transcription factor.

A biotinylated DNA oligo containing a specific transcription-factor binding sequence was immobilized on a streptavidin-coated 96-well plate. NX1 extraction buffer (Blank) or 10 µg nucleic acid binding protein fraction was added, washed, and detected colorimetrically in an ELISA procedure using a transcription-factor specific antibody. Specificity of binding was demonstrated by addition of a 10x excess of non-biotinylated oligo that was able to displace the transcription factor.

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Nuclear protein fractionation procedure.
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Reproducible, efficient separation of marker proteins.

Three cell lysate preparations were processed in parallel using the Qproteome Nuclear Protein Kit. Fractions were separated by SDS-PAGE. Fraction-specific markers (GAPDH, cytosolic fraction; transcription factor SP1, nucleic acid binding protein fraction [NABP]; and nucleoporin, insoluble fraction) were detected using protein-specific antibodies in a western blotting procedure.

Performance
The Qproteome Nuclear Protein Kit provides highly effective and reproducible separation of nuclear proteins as demonstrated using specific marker proteins (see figure "Reproducible, efficient separation of marker proteins"), which remain functionally active (see figure "Isolation of an active transcription factor").
Principle

The Qproteome Nuclear Protein Kit is designed for specific enrichment of nuclear proteins from cultured mammalian cells. Lysis and centrifugation are used to separate the cytosolic fraction (supernatant) from the cell nuclei (pellet). A high-salt buffer allows dissociation of nuclear binding proteins (such as transcription factors) and their removal by diffusion from the nuclei.

Procedure
Cells are lysed and centrifuged to isolate nuclei. After washing, an extraction buffer is added to the nuclei and nucleic acid binding proteins dissociate from DNA and RNA. This soluble fraction is separated from the nuclear pellet by centrifugation. Proteins remaining in the pellet are solubilized using a second extraction buffer (see figure "Nuclear protein fractionation procedure").
Applications

The Qproteome Nuclear Protein Kit delivers a nucleic acid binding protein fraction suitable for a wide range of activity assays.

Features
Specifications
Applications SDS-PAGE, mass spectrometry
Binding capacity/yield 200–300 µg
For glycoproteins: which type of glycoproteins n.d
Fractions isolated Three fractions
Sample size 5 x 10e6 cells
Species Mammal
Start material Cell cultures
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