Minute™ Cytosolic and Nuclear Isolation Kit for Metabolite Analysis (20 Preps)

Manual & Protocol | MSDS

This detergent-free cytosolic and nuclear isolation kit is designed to generate samples for metabolite analysis from cultured cells and tissues. Intact nuclei can be isolated in less than 40 minutes without the use of Dounce homogenizers or detergents. Traditional nuclear isolation methods typically rely on non-ionic detergents, which can interfere with mass spectrometry and may cause leakage of nuclear metabolites.

In this method, cells or tissues are first sensitized with Buffer A and then passed through a proprietary filter in a zigzag path under high-speed centrifugal force. Cells rupture as they pass through the filter, while intact nuclei are released into the flow-through. The nuclei are subsequently separated from the cytosolic fraction by centrifugation, and nuclear metabolites are extracted using methanol or other suitable solvents.

 Fig. 1

Comparison of metabolite profiles of cytosol and nuclear extracts. Score plots of principal component analysis (PCA) of epigenetic metabolites (a custom platform) in the rat liver tissue. CSAA group (blue) represents healthy control, where Fisher-344 male rats were on a choline-sufficient amino acid defined diet (CSAA). In CDAA group (red), rats were on a hepatocellular carcinoma (HCC)-inducing carcinogenic choline-deficient amino acid defined diet (CDAA). In the third CDAA+PTS experimental group (green), rats remained on the HCC-inducing diet that was supplemented with pterostilbene (PTS). The number of animals in each group was n=6. In the cytoplasm, the CDAA group (red) and CDAA+PTS group (green) show a partial overlap of the metabolite profile but retain distinguishable clustering patterns. In the nucleic fraction, the metabolic signature in the CDAA+PTS group (green) is clearly shifted towards the healthy CSAA group (blue). The analysis was performed using the MetaboAnalyst platform. PC1 and PC2 capture maximum variance in the metabolic data, with the percentage of the total variation explained by each component. PC1 explains the most variance, and PC2 the second most variance.

Kit includes: