Qproteome Mitochondria Isolation Kit

For efficient isolation of high-purity mitochondria preparations

Features

  • High-purity, intact mitochondria
  • Highly reproducible using a standardized procedure
  • Convenient, fast, and easy-to-use kit format
Qproteome Mitochondria Isolation Kit

Cat. No. / ID: 37612

Buffers and reagents for 12 high-purity mitochondrial preparations
The Qproteome Mitochondria Isolation Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.

Product Details

The Qproteome Mitochondria Isolation Kit provides fully active mitochondrial preparations of very high purity in a fast, reproducible, and standardized procedure that uses a standard bench-top centrifuge.

Performance

In contrast to a differential centrifugation method from the literature, analysis of cell-compartment marker proteins shows that the Qproteome Mitochondria Isolation Kit provides mitochondrial fractions virtually free of cross-contamination (see figure "Mitochondrial fractions virtually free of cross-contamination"). The virtual absence of cytosolic contaminants greatly facilitates analysis of low-abundance species as shown in 2D-PAGE analyses and the gentle procedure provides a highly enriched fraction containing intact mitochondria (see figure " High purity mitochondria with intact mitochondrial membranes "). Integrity can also be demonstrated by measuring uptake of a fluorescent dye (see figure "More intact mitochondria per milligram protein").
See figures

Principle

One of the major problems facing researchers working with mitochondria is efficiently separating mitochondrial and cytosolic fractions. Traditional preparation methods are tedious, time-consuming, and require access to an ultracentrifuge. The Qproteome Mitochondria Isolation Kit provides fully active mitochondrial preparations of very high purity in a fast, reproducible, and standardized procedure that uses a standard bench-top centrifuge.

Applications

Mitochondria are the subject of intense study in a number of fields, including apoptosis, proteomics, biomedical research, and drug discovery. The Qproteome Mitochondria Isolation Kit delivers high-purity mitochondria whose proteins retain their native conformation and full biological activity — making them highly suited for apoptosis studies, 2D-PAGE analysis, and other biochemical assays.

Supporting data and figures

Specifications

FeaturesSpecifications
applicationsSDS-PAGE, mass spectrometry, activity assays
samplesize5 x 10e6 - 2 x 10e7 cells, 20 mg - 60 mg
forglycoproteinswhichtypeofglycoproteinsn.d
bindingcapacityyield20–80 µg
speciesEukaryotes
fractionsisolatedOne fraction
startmaterialCells, Tissue

FAQ

Can mitochondria separated with the Qproteome Mitochondria Isolation Kit be used for downstream DNA isolation?

Even though we have not tested this, we assume that both the QIAamp DNA Micro Kit and the AllPrep DNA/RNA Mini Kit will work to isolate DNA from mitochondria separated with the Qproteome Mitochondria Isolation Kit.

 

FAQ ID -1188
Which Qproteome or Protein Fractionation Kits from QIAGEN are compatible with tissue samples?

The Qproteome Cell Compartment Kit, the Qproteome Nuclear Protein Kit, the Qproteome Mitochondria Isolation Kit, and the PhosphoProtein Purification Kit are compatible with tissue samples. The respective protocol can be found in the respective handbook.

The Qproteome Mammalian Protein Prep Kit is also compatible with tissue, there is the QIAGEN Supplementary Protocol: Purification of protein from animal tissues using the Qproteome Mammalian Protein Prep Kit and the TissueRuptor™ available. For all these protocols the TissueRuptor is used.

 

 

 

FAQ ID -755
Can the nuclei and cell debris fraction generated in the QProteome Mitochondria isolation procedure be used for further fractionation?

We have not tested this application. We would suggest to split the sample, and perform different fractionation procedures separately on the two sample portions.

If splitting the sample is not feasible, then it should theoretically be possible to use the pellet generated at step 7 of the QProteome Mitochondria Isolation Kit protocol (containing cell debris, nuclei and unbroken cells) as starting material for the Qproteome Nuclear Protein Kit.

Please note that this application will have to be tested and optimized by the enduser.

 

FAQ ID -1185
Is it possible to collect intact mitochondria before or after the density gradient centrifugation step of the Qproteome Mitochondria Isolation protocol?

Highly pure and intact mitochondria can be collected after the density gradient centrifugation at step 11b of the Qproteome Mitochondria Isolation protocol.

The fraction collected at step 9 of the protocol, prior to the cushion centrifugation, contains intact as well as broken mitochondria, and may also comprise structures of the ER, Golgi and plasma membranes.

FAQ ID -1187
How many cells can be used with the Qproteome Mitochondria Isolation Kit, and how much mitochondrial protein can be expected?

The Qproteome Mitochondria Isolation Kit can be used with 5 x 106 to 2 x 107 cells. Depending on the cell line, the yield from a single fractionation procedure is 20–80 µg protein from 5 x 106 cells.

Protein can be isolated from the mitochondria using, e.g. the QProteome Mammalian Protein Prep Kit, or home-made lysis buffers.

 

 

FAQ ID -1189
Do you have a kit for protein isolation from mitochondria prepared with your Qproteome Mitochondria Isolation Kit?

Yes, you can use the Qproteome Mammalian Protein Prep Kit to isolate proteins from mitochondria separated with the Qproteome Mitochondria Isolation Kit.

 

FAQ ID -1190
Do I need an ultracentrifuge for the density gradient centrifugation step in the Qproteome Mitochondria Isolation protocol?
No, an ultracentrifuge is not required for the Qproteome Mitochondria Isolation Kit. All centrifugation steps are performed in a standard table-top centrifuge.
FAQ ID -1186
What starting material can be used with the Qproteome Mitochondria Isolation Kit?

The Qproteome Mitochondria Isolation Kit has been developed for cultured mammalian cells. We have tested HeLa, Cos7, NIH, HEK 293 and SH-SY5Y cell lines.

We have not tested animal tissue, yeast, or plant cells. Animal tissue might work if a method for cell disruption that leaves the mitochondria intact can be found. Unfortunately, we do not have any protocol recommendations for these starting materials.

FAQ ID -1184