MT-1 MitoMP Detection Kit

Product Description
Mitochondria is an important organelle that uses oxygen to synthesize ATP, producing the necessary energy for living cells to thrive¹⁾. Decreased mitochondrial activity and mitochondrial dysfunction are associated with cancer, aging, and neurodegenerative diseases such as Alzheimer’s and Parkinson’s disease^2,3). Therefore, mitochondrial membrane potential (MMP) has been widely studied as a promising target for mitochondria-related diseases.

Reference
1) K. F. Ferri, et al., J. Exp. Med., 2000, 192, 1081.
2) N. Matsuda, et al., J. Cell Biol., 2010, 189, 211.
3) J. L. Wang, et al., PNAS, 2000, 97, 7124.

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Overcome three limitations of conventional reagents
JC-1 dye, TMRE, and TMRM are widely used to monitor MMP, however, these dyes have some limitations, such as low photostability and poor retention after aldehyde fixation. These limitations result in poor reproducibility of experiments.
Dojindo’s MT-1 MitoMP Detection Kit overcomes these limitations. In addition, the Imaging Buffer included in this kit minimizes background fluorescence and maintains cell vitality while the assay is being performed.

 

① Applicable to fixation after staining
Since mitochondrial membrane potential (MMP) fluctuates according to slight changes of cellular status, extreme care has been needed to obtain repeatable data. In JC-1 dye, which is widely used for measuring the MMP, the fluorescence is lost after a cell has been fixed; therefore, the use of living cells is necessary for the quick measurement of MMP.
In the meantime, the MT-1 dye, which remains unquenched even in the cells fixed with paraformaldehyde after staining, can be used to conduct a highly repeatable experiment.

<Detection Condition>
Ex: 530-560 nm, Em: 570-640 nm
Scale bar: 100 μm
HeLa cells

 

② Allow to monitor mitochondrial membrane potential

　

<Detection Condition>
Ex: 530-560 nm, Em: 570-640 nm
Scale bar: 100 μm
HeLa cells

 

③ High sensitivity for mitochondrial membrane potential
Sometimes it is difficult to detect slight changes of MMP in JC-1 stained mitochondria. In such a case, tetramethylrhodamine ethyl ester (TMRE) was used to monitor MMP. MT-1 can provide equivalent detection sensitivity to TMRE.

<Detection Condition>
Ex: 530-560 nm, Em: 570-640 nm
HeLa cells

 

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Reagent Comparison

 

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Procedure

 

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Experimental Example: Depolarization
HeLa cells were treated with a depolarizing agent, carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP), and changes in mitochondrial membrane potential were observed in a time-lapse imaging with this kit.
As a result, it was confirmed that the mitochondrial membrane potential of the cells treated with FCCP decreased.

 

Experimental Example: Changes in mitochondrial membrane potential in apoptosis-induced cells
Apoptosis was induced by Etoposide to HL60 cells stained with MT-1 in advance, then co-stained with Annexin V-FITC and analyzed by flow cytometry.
Apoptotic progress and MMP change were confirmed through the fluorescence intensity changes of Annexin V-FITC (an increase in the green fluorescence intensity) and MT-1 (a decrease in the red fluorescence intensity), respectively.

<Detection Condition>
Annexin V-FITC (green): Ex. 488 nm / Em. 500 – 560 nm
Mitochondria membrane potential (MT-1, red): Ex 488 nm / Em 564-604 nm