Mitochondria MidiMACS Starting Kit, mouse tissue130-097-039
产品说明
The isolation protocol is based on the renowned MACS Technology, which enables fast isolation of high purity and high yield mitochondria.
Detailed procedure
Tissues can be homogenized with the gentleMACS or gentleMACS Octo Dissociator in combination with the Mitochondria Extraction Kit – Tissue. After one-step homogenization of the tissue and lysis of the cell suspension, mitochondria are magnetically labeled with Anti-TOM22 MicroBeads, mouse, which bind to the translocase of the outer mitochondrial membrane 22 protein (TOM22). The sample is loaded onto an LS Column placed in a MidiMACS or QuadroMACS Separator. After washing, only the magnetically labeled mitochondria are retained on the column. The column is removed from the separator and functional mouse mitochondria are eluted from the column.
Downstream applications
After isolation mitochondria can be further subjected to Western blot analysis1,6 or downstream functional assays, including measurement of O2, membrane potential, and ATP4-5,7, or respiratory control4. Growing evidence also suggests that isolated mitochondria are well suited for mitochondrial RNA expression profiling studies2,3.
特点
Gentle: isolation of functional mitochondria
Specific: high purity of isolated mitochondria
Fast: mitochondria isolation in less than 2 hours
相关图片
Integrated tissue dissociation followed by mitochondria isolation.
Enrichment of functional mitochondria. Mitochondria were prepared from various mouse tissues using the Mitochondria Isolation Kit, mouse tissue. The mitochondria preparations from muscle, liver, and brain, as well as the flow-through fractions were analyzed by a coupled enzymatic citrate synthase assay. The data indicate that citrate synthase activity, normalized to the activity per mg protein assayed in the flow-through fraction, is highly enriched in the eluate.
Isolation of high-purity mitochondria from mouse liver. After isolation of mitochondria from mouse liver using the Mitochondria Isolation Kit, mouse tissue, equal amounts of mitochondrial protein lysates were analyzed by Western blotting against TOM20. PEX1, a cytoplasmic protein anchored to peroxisomal membranes, served as a control. The results show an effective enrichment of mitochondria and no contamination of the mitochondria fraction by peroxisomes.