Qproteome Mitochondria Isolation Kit

Para el aislamiento eficiente de las preparaciones mitocondriales de alta pureza

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Qproteome Mitochondria Isolation Kit

Cat. No. / ID:  37612

Tampones y reactivos para 12 preparaciones mitocondriales de alta pureza
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El Qproteome Mitochondria Isolation Kit está concebido para su uso en aplicaciones de biología molecular. Este producto no está concebido para el diagnóstico, la prevención ni el tratamiento de enfermedades.

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Features

  • Mitocondrias intactas de gran pureza
  • Altamente reproducible mediante un procedimiento normalizado
  • Formato de kit práctico, rápido y fácil de usar

Product Details

El Qproteome Mitochondria Isolation Kit proporciona preparaciones mitocondriales totalmente activas de muy alta pureza en un procedimiento rápido, reproducible y normalizado que utiliza una centrífuga estándar de sobremesa.

Performance

En contraste con un método de centrifugación diferencial de la bibliografía, el análisis de las proteínas marcadoras del compartimento celular muestra que el Qproteome Mitochondria Isolation Kit proporciona fracciones mitocondriales prácticamente libres de contaminación cruzada (consulte la figura «Fracciones mitocondriales prácticamente sin contaminación cruzada»). La ausencia virtual de contaminantes citosólicos facilita enormemente el análisis de especies de baja abundancia, como se muestra en los análisis 2D-PAGE, y el procedimiento suave proporciona una fracción altamente enriquecida que contiene mitocondrias intactas (consulte la figura « Mitocondrias de alta pureza con membranas mitocondriales intactas »). La integridad también puede demostrarse midiendo la captación de un colorante fluorescente (consulte la figura «Más mitocondrias intactas por miligramo de proteína»).
See figures

Principle

Uno de los mayores problemas a los que se enfrentan los investigadores que trabajan con mitocondrias es separar de forma eficiente las fracciones mitocondriales y citosólicas. Los métodos tradicionales son tediosos, llevan mucho tiempo y requieren acceso a una ultracentrífuga. El Qproteome Mitochondria Isolation Kit proporciona preparaciones mitocondriales totalmente activas de muy alta pureza en un procedimiento rápido, reproducible y normalizado que utiliza una centrífuga estándar de sobremesa.

Applications

Las mitocondrias son objeto de estudio intenso en diversos campos, como la apoptosis, la proteómica, la investigación biomédica y el descubrimiento de fármacos. El Qproteome Mitochondria Isolation Kit proporciona mitocondrias de alta pureza cuyas proteínas conservan su conformación nativa y su actividad biológica completa, lo que hace que sean muy adecuadas para estudios de apoptosis, análisis 2D-PAGE y otros ensayos bioquímicos.

Supporting data and figures

Specifications

FeaturesSpecifications
ApplicationsSDS-PAGE, espectrometría de masa, ensayos de actividad
Sample size5 x 10e6-2 x 10e7 células, 20 mg - 60 mg
For glycoproteins: which type of glycoproteinss.f.
Binding capacity/yield20-80 µg
SpeciesEucariotas
Fractions isolatedUna fracción
Start materialCélulas, Tejido

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