2024年5月26日发(作者：秘叶舞)
Product Information
Revised: 03–March–2005
MitoSOX™ Red mitochondrial superoxide indicator *for live-cell
imaging* (M36008)
Quick Facts
Storage upon receipt:
•
≤
–20ºC
• Protect from light
• Desiccate
• Avoid freeze-thaw cycles
Ex/Em:
510/580 nm
Note:
To preserve the activity of the MitoSOX™ Red
reagent, minimize exposure to air.
线粒体超氧化物作为氧化磷酸化的副产物产生。 在另外紧密耦合的电子传输链
中，大约1-3％的线粒体氧消耗不完全减少; 这些“泄漏”电子可以与分子氧快
速相互作用以形成超氧化物阴离子，线粒体中的主要活性氧（ROS）.1-4增加细
胞超氧化物产生已经涉及心血管疾病，包括高血压，动脉粥样硬化和糖尿病 -
相关的血管损伤，5-7以及神经变性疾病如帕金森氏病，阿尔茨海默氏病和肌萎
缩性侧索硬化（ALS）
Introduction
Mitochondrial superoxide is generated as a byproduct of oxi-
dative phosphorylation. In an otherwise tightly coupled electron
transport chain, approximately 1–3% of mitochondrial oxygen
consumed is incompletely reduced; those “leaky” electrons can
quickly interact with molecular oxygen to form superoxide anion,
the predominant reactive oxygen species (ROS) in mitochon-
dria.
1–4
Increases in cellular superoxide production have been
implicated in cardiovascular diseases, including hypertension,
atherosclerosis, and diabetes-associated vascular injuries,
5–7
as well as in neurodegenerative diseases such as Parkinsonʼs,
Alzheimerʼs, and amyotrophic lateral sclerosis (ALS).
8–12
The assumption that mitochondria serve as the major intracel-
lular source of ROS has been based largely on experiments with
isolated mitochondria rather than direct measurements in living
cells. MitoSOX
™
Red mitochondrial superoxide indicator is a
novel fluorogenic dye for highly selective detection of super-
oxide (Figure 1) in the mitochondria of live cells. MitoSOX™
Red reagent is live-cell permeant and is rapidly and selec-
tively targeted to the mitochondria. Once in the mitochondria,
MitoSOX™ Red reagent is oxidized by superoxide and exhibits
red fluorescence. MitoSOX™ Red reagent is readily oxidized by
superoxide but not by other ROS- or reactive nitrogen species
(RNS)–generating systems, and oxidation of the probe is pre-
vented by superoxide dismutase. The oxidation product becomes
highly fluorescent upon binding to nucleic acids.
This reagent may enable researchers to distinguish artifacts
of isolated mitochondrial preparations from direct measurements
of superoxide generated in the mitochondria of live cells. It may
also provide a valuable tool in the discovery of agents that modu-
late oxidative stress in various pathologies.
副产品
Figure 1. Selectivity of the MitoSOX™ Red mitochondrial superoxide indicator. Cell-
free systems were used to generate a variety of reactive oxygen species (ROS) and
reactive nitrogen species (RNS); each oxidant was then added to a separate 10 µM
solution of MitoSOX™ Red reagent and incubated at 37˚C for 10 minutes. Excess DNA
was added (unless otherwise noted) and the samples were incubated for an additional
15 minutes at 37˚C before fluorescence was measured. The Griess nitrite determina-
tion kit (for nitric oxide, peroxynitrite, and nitrite standards only; gray bars) and
dihydrorhodamine 123 (DHR 123; white bars) were employed as positive controls for
oxidant generation. Superoxide dismutase (SOD), a superoxide scavenger, was used as
a negative control for superoxide. The results show that the MitoSOX™ Red probe (black
bars) is readily oxidized by superoxide but not by the other oxidants.
Materials
Contents
MitoSOX™ Red mitochondrial superoxide indicator (MW = 759)
is supplied in 10 vials, each containing 50 μg.
Storage and Handling
Upon receipt, the MitoSOX™ Red reagent should be stored
upright, desiccated, and protected from light at ≤–20˚C. Avoid
freeze-thaw cycles. MitoSOX™ Red reagent is packaged with
an oxygen scavenging pouch that will extend the shelf life of the
product. After removing vials for use, seal the remaining vials
into the pouch to preserve the activity of the reagent. Vials
should be allowed to warm to room temperature before opening.
When stored properly, components should be stable for at least
6 months. Note: MitoSOX™ Red reagent is a derivative of ethid-
ium bromide and should be treated with the appropriate caution.
MP 36008 MitoSOX ™ Red mitochondrial superoxide indicator
Spectral Characteristics
MitoSOX™ Red mitochondrial superoxide indicator has exci-
tation/emission maxima of approximately 510/580 nm.
protocols should be used as a starting point, and optimal labeling
conditions should be determined empirically.
1.1 Prepare 5 μM MitoSOX™ reagent working solution.
Dilute the 5 mM MitoSOX™ reagent stock solution (prepared
above) in HBSS/Ca/Mg or suitable buffer to make a 5 μM
MitoSOX™ reagent working solution.
Note: The concentration of the MitoSOX™ reagent working so-
lution should not exceed 5 μM. Concentrations exceeding 5 μM
can produce cytotoxic effects, including altered mitochondrial
morphology and redistribution of fluorescence to nuclei and the
cytosol.
1.2 Load cells. Apply 1.0–2.0 mL of 5 μM MitoSOX™ reagent
working solution (prepared in step 1.2) to cover cells adhering
to coverslip(s). Incubate cells for 10 minutes at 37˚C, protected
from light.
1.3 Wash cells. Wash cells gently three times with warm buffer.
1.4 Prepare cells for viewing. Stain cells with counterstains as
desired and mount in warm buffer for imaging.
复染
Materials Recommended but Not Provided
Hankʼs balanced salt solution with calcium and magnesium
(HBSS/Ca/Mg, available from Gibco (14025-092)).
Experimental Protocol
Reagent Preparation
Prepare 5 mM MitoSOX™ reagent stock solution. Dissolve
the contents (50 μg) of one vial of MitoSOX™ mitochondrial
superoxide indicator (Component A) in 13 μL of dimethylsulfox-
ide (DMSO) to make a 5 mM MitoSOX™ reagent stock solution.
Labeling Live Eukaryotic Cells
This protocol was developed using live bovine pulmonary
epithelial (BPAE) cells, MRC5 human lung fibroblasts, and
mouse 3T3 fibroblasts adhering to coverslips, but can be adapted
for use with other cell types. Recommendations for experimental
牛肺上皮
References
1. J Cell Mol Med 6, 175 (2002); 2. J Biol Chem 279, 4127 (2004); 3. J Neurochem 80, 780 (2002); 4. Bioorganic and Medicinal Chemistry 10, 3013
(2002); 5. Am J Physiol Heart Circ Physiol 284, H605 (2003); 6. Bioorganic and Medicinal Chemistry 10, 3013 (2002); 7. J Neurochem 89, 1283
(2004); 8. Trends Neurosci 24, S15 (2001); 9. J Cell Mol Med 6, 175 (2002); 10. J Neurosci 18, 923 (1998); 11. Neurochem Res 28, 1563 (2003);
12. IUBMB Life 55, 329 (2003).
Prod uct List
Current pric es may be ob tained from our Web site or from our Customer Service Department.
Cat #
M36008
Product Name Unit Size
MitoSOX™ Red mitochondrial superoxide indicator *for live-cell imaging* ................................................................................................... 10 x 50 µg
MitoSOX ™ Red mitochondrial superoxide indicator 2
Contact Information
Further information on Molecular Probes products, including product bibliographies, is available from your local distributor or directly from Molecular Probes. Customers in
Europe, Africa and the Middle East should contact our office in Paisley, United Kingdom. All others should contact our Technical As sis tance Department in Eugene, Oregon.
Please visit our Web site —
— for the most up-to-date information.
Molecular Probes, Inc.
29851 Willow Creek Road, Eugene, OR 97402
Phone: (541) 465-8300 • Fax: (541) 335-0504
Invitrogen European Headquarters
Invitrogen, Ltd.
3 Fountain Drive
Inchinnan Business Park
Paisley PA4 9RF, UK
Phone: +44 (0) 141 814 6100 • Fax: +44 (0) 141 814 6260
Email: euroinfo@
Technical Services: eurotech@
Customer Service:
6:00 am to 4:30 pm (Pacific Time)
Phone: (541) 335-0338 • Fax: (541) 335-0305 • order@
Toll-Free Ordering for USA:
Order Phone: (800) 438-2209 • Order Fax: (800) 438-0228
Technical Assistance:
8:00 am to 4:00 pm (Pacific Time)
Phone: (541) 335-0353 • Toll-Free (800) 438-2209
Fax: (541) 335-0238 • tech@
Molecular Probes products are high-quality reagents and materials intended for research pur pos es only. These products must be used by, or directl y under the
super vision of, a tech nically qual i fied individual experienced in handling potentially hazardous chemicals. Please read the Material Safety Data Sheet pro vid ed for
each prod uct; other regulatory considerations may apply.
Several Molecular Probes products and product applications are covered by U.S. and foreign patents and patents pending. Our prod ucts are not available for resale or
oth er commercial uses without a specifi c agreement from Molecular Probes, Inc. We welcome inquiries about licensing the use of our dyes, trade marks or technolo-
is tered with the U.S. Patent and Trade mark Offi . Please submit inquiries by e-mail to busdev@. All names con tain ing the des ig na tion
®
are reg
Copyright 2005, Molecular Probes, Inc. All rights reserved. This information is subject to change without notice.
MitoSOX ™ Red mitochondrial superoxide indicator 3
2024年5月26日发(作者：秘叶舞)
Product Information
Revised: 03–March–2005
MitoSOX™ Red mitochondrial superoxide indicator *for live-cell
imaging* (M36008)
Quick Facts
Storage upon receipt:
•
≤
–20ºC
• Protect from light
• Desiccate
• Avoid freeze-thaw cycles
Ex/Em:
510/580 nm
Note:
To preserve the activity of the MitoSOX™ Red
reagent, minimize exposure to air.
线粒体超氧化物作为氧化磷酸化的副产物产生。 在另外紧密耦合的电子传输链
中，大约1-3％的线粒体氧消耗不完全减少; 这些“泄漏”电子可以与分子氧快
速相互作用以形成超氧化物阴离子，线粒体中的主要活性氧（ROS）.1-4增加细
胞超氧化物产生已经涉及心血管疾病，包括高血压，动脉粥样硬化和糖尿病 -
相关的血管损伤，5-7以及神经变性疾病如帕金森氏病，阿尔茨海默氏病和肌萎
缩性侧索硬化（ALS）
Introduction
Mitochondrial superoxide is generated as a byproduct of oxi-
dative phosphorylation. In an otherwise tightly coupled electron
transport chain, approximately 1–3% of mitochondrial oxygen
consumed is incompletely reduced; those “leaky” electrons can
quickly interact with molecular oxygen to form superoxide anion,
the predominant reactive oxygen species (ROS) in mitochon-
dria.
1–4
Increases in cellular superoxide production have been
implicated in cardiovascular diseases, including hypertension,
atherosclerosis, and diabetes-associated vascular injuries,
5–7
as well as in neurodegenerative diseases such as Parkinsonʼs,
Alzheimerʼs, and amyotrophic lateral sclerosis (ALS).
8–12
The assumption that mitochondria serve as the major intracel-
lular source of ROS has been based largely on experiments with
isolated mitochondria rather than direct measurements in living
cells. MitoSOX
™
Red mitochondrial superoxide indicator is a
novel fluorogenic dye for highly selective detection of super-
oxide (Figure 1) in the mitochondria of live cells. MitoSOX™
Red reagent is live-cell permeant and is rapidly and selec-
tively targeted to the mitochondria. Once in the mitochondria,
MitoSOX™ Red reagent is oxidized by superoxide and exhibits
red fluorescence. MitoSOX™ Red reagent is readily oxidized by
superoxide but not by other ROS- or reactive nitrogen species
(RNS)–generating systems, and oxidation of the probe is pre-
vented by superoxide dismutase. The oxidation product becomes
highly fluorescent upon binding to nucleic acids.
This reagent may enable researchers to distinguish artifacts
of isolated mitochondrial preparations from direct measurements
of superoxide generated in the mitochondria of live cells. It may
also provide a valuable tool in the discovery of agents that modu-
late oxidative stress in various pathologies.
副产品
Figure 1. Selectivity of the MitoSOX™ Red mitochondrial superoxide indicator. Cell-
free systems were used to generate a variety of reactive oxygen species (ROS) and
reactive nitrogen species (RNS); each oxidant was then added to a separate 10 µM
solution of MitoSOX™ Red reagent and incubated at 37˚C for 10 minutes. Excess DNA
was added (unless otherwise noted) and the samples were incubated for an additional
15 minutes at 37˚C before fluorescence was measured. The Griess nitrite determina-
tion kit (for nitric oxide, peroxynitrite, and nitrite standards only; gray bars) and
dihydrorhodamine 123 (DHR 123; white bars) were employed as positive controls for
oxidant generation. Superoxide dismutase (SOD), a superoxide scavenger, was used as
a negative control for superoxide. The results show that the MitoSOX™ Red probe (black
bars) is readily oxidized by superoxide but not by the other oxidants.
Materials
Contents
MitoSOX™ Red mitochondrial superoxide indicator (MW = 759)
is supplied in 10 vials, each containing 50 μg.
Storage and Handling
Upon receipt, the MitoSOX™ Red reagent should be stored
upright, desiccated, and protected from light at ≤–20˚C. Avoid
freeze-thaw cycles. MitoSOX™ Red reagent is packaged with
an oxygen scavenging pouch that will extend the shelf life of the
product. After removing vials for use, seal the remaining vials
into the pouch to preserve the activity of the reagent. Vials
should be allowed to warm to room temperature before opening.
When stored properly, components should be stable for at least
6 months. Note: MitoSOX™ Red reagent is a derivative of ethid-
ium bromide and should be treated with the appropriate caution.
MP 36008 MitoSOX ™ Red mitochondrial superoxide indicator
Spectral Characteristics
MitoSOX™ Red mitochondrial superoxide indicator has exci-
tation/emission maxima of approximately 510/580 nm.
protocols should be used as a starting point, and optimal labeling
conditions should be determined empirically.
1.1 Prepare 5 μM MitoSOX™ reagent working solution.
Dilute the 5 mM MitoSOX™ reagent stock solution (prepared
above) in HBSS/Ca/Mg or suitable buffer to make a 5 μM
MitoSOX™ reagent working solution.
Note: The concentration of the MitoSOX™ reagent working so-
lution should not exceed 5 μM. Concentrations exceeding 5 μM
can produce cytotoxic effects, including altered mitochondrial
morphology and redistribution of fluorescence to nuclei and the
cytosol.
1.2 Load cells. Apply 1.0–2.0 mL of 5 μM MitoSOX™ reagent
working solution (prepared in step 1.2) to cover cells adhering
to coverslip(s). Incubate cells for 10 minutes at 37˚C, protected
from light.
1.3 Wash cells. Wash cells gently three times with warm buffer.
1.4 Prepare cells for viewing. Stain cells with counterstains as
desired and mount in warm buffer for imaging.
复染
Materials Recommended but Not Provided
Hankʼs balanced salt solution with calcium and magnesium
(HBSS/Ca/Mg, available from Gibco (14025-092)).
Experimental Protocol
Reagent Preparation
Prepare 5 mM MitoSOX™ reagent stock solution. Dissolve
the contents (50 μg) of one vial of MitoSOX™ mitochondrial
superoxide indicator (Component A) in 13 μL of dimethylsulfox-
ide (DMSO) to make a 5 mM MitoSOX™ reagent stock solution.
Labeling Live Eukaryotic Cells
This protocol was developed using live bovine pulmonary
epithelial (BPAE) cells, MRC5 human lung fibroblasts, and
mouse 3T3 fibroblasts adhering to coverslips, but can be adapted
for use with other cell types. Recommendations for experimental
牛肺上皮
References
1. J Cell Mol Med 6, 175 (2002); 2. J Biol Chem 279, 4127 (2004); 3. J Neurochem 80, 780 (2002); 4. Bioorganic and Medicinal Chemistry 10, 3013
(2002); 5. Am J Physiol Heart Circ Physiol 284, H605 (2003); 6. Bioorganic and Medicinal Chemistry 10, 3013 (2002); 7. J Neurochem 89, 1283
(2004); 8. Trends Neurosci 24, S15 (2001); 9. J Cell Mol Med 6, 175 (2002); 10. J Neurosci 18, 923 (1998); 11. Neurochem Res 28, 1563 (2003);
12. IUBMB Life 55, 329 (2003).
Prod uct List
Current pric es may be ob tained from our Web site or from our Customer Service Department.
Cat #
M36008
Product Name Unit Size
MitoSOX™ Red mitochondrial superoxide indicator *for live-cell imaging* ................................................................................................... 10 x 50 µg
MitoSOX ™ Red mitochondrial superoxide indicator 2
Contact Information
Further information on Molecular Probes products, including product bibliographies, is available from your local distributor or directly from Molecular Probes. Customers in
Europe, Africa and the Middle East should contact our office in Paisley, United Kingdom. All others should contact our Technical As sis tance Department in Eugene, Oregon.
Please visit our Web site —
— for the most up-to-date information.
Molecular Probes, Inc.
29851 Willow Creek Road, Eugene, OR 97402
Phone: (541) 465-8300 • Fax: (541) 335-0504
Invitrogen European Headquarters
Invitrogen, Ltd.
3 Fountain Drive
Inchinnan Business Park
Paisley PA4 9RF, UK
Phone: +44 (0) 141 814 6100 • Fax: +44 (0) 141 814 6260
Email: euroinfo@
Technical Services: eurotech@
Customer Service:
6:00 am to 4:30 pm (Pacific Time)
Phone: (541) 335-0338 • Fax: (541) 335-0305 • order@
Toll-Free Ordering for USA:
Order Phone: (800) 438-2209 • Order Fax: (800) 438-0228
Technical Assistance:
8:00 am to 4:00 pm (Pacific Time)
Phone: (541) 335-0353 • Toll-Free (800) 438-2209
Fax: (541) 335-0238 • tech@
Molecular Probes products are high-quality reagents and materials intended for research pur pos es only. These products must be used by, or directl y under the
super vision of, a tech nically qual i fied individual experienced in handling potentially hazardous chemicals. Please read the Material Safety Data Sheet pro vid ed for
each prod uct; other regulatory considerations may apply.
Several Molecular Probes products and product applications are covered by U.S. and foreign patents and patents pending. Our prod ucts are not available for resale or
oth er commercial uses without a specifi c agreement from Molecular Probes, Inc. We welcome inquiries about licensing the use of our dyes, trade marks or technolo-
is tered with the U.S. Patent and Trade mark Offi . Please submit inquiries by e-mail to busdev@. All names con tain ing the des ig na tion
®
are reg
Copyright 2005, Molecular Probes, Inc. All rights reserved. This information is subject to change without notice.
MitoSOX ™ Red mitochondrial superoxide indicator 3