Probing Mitochondria in Living Cells with Rhodamine 123

Chen, L. B.; Summerhayes, Ian C.; Johnson, Lincoln V.; Walsh, Marcia L.; Bernal, Samuel D.; Lampidis, Théodore J.

doi:10.1101/sqb.1982.046.01.018

Cited by 169 publications

(94 citation statements)

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“…The overall fluorescent signal obtained from a suspension of isolated mitochondria is assumed to be the sum of contributions from dye in the buffer, in mitochondrial matrix, and in the membrane: (8) In Eq. (8) the first two terms in the numerator correspond to volume-weighted contributions from the aqueous-phase compartments.…”

Section: Relationship Between Fluorescence and Transient Membrane Potmentioning

confidence: 99%

“…13 Changes in accumulation produce changes in fluorescence signal, indicating changes in ΔΨ. 7,8,13 Yet the measured dye distribution (concentration in buffer vs. that in mitochondria) can deviate significantly from Nernst equilibrium. The deviation comes from (i) response time needed to reach equilibrium due to the dye's limited permeability in measurements of transient changes, and (ii) substantial partition of the dye into the mitochondrial membranes.…”

Section: Introductionmentioning

confidence: 99%

“…first demonstrated that R123 specifically stained mitochondria in response to mitochondria energization in cultured cells, and those chemical agents that collapse membrane potential prevents uptake of R123 by mitochondria. 7,8,16 Emaus et al 13 extensively characterized R123 as a probe of membrane potential for suspensions of isolated mitochondria. They showed that the R123 fluorescence spectrum shifts to the red in response to mitochondria energization, and that there is an empirical linear relationship between fluorescence intensity change and membrane potential.…”

Section: Introductionmentioning

confidence: 99%

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Mitochondrial Inner Membrane Electrophysiology Assessed by Rhodamine-123 Transport and Fluorescence

et al. 2007

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Rhodamine-123 is widely used to make dynamic measurements of mitochondrial membrane potential both in vitro and in situ. Yet data interpretation is difficult due to a lack of quantitative understanding of how membrane potential and measured fluorescence are related. To develop such understanding, a model for dye transport across the mitochondrial inner membrane and partition into the membrane was developed. The model accounts for experimentally measured dye selfquenching and was integrated into a model of mitochondrial electrophysiology to estimate transients in mitochondrial membrane potential from kinetic fluorescence measurements. Our analysis indicates that (i) R123 fluorescence peaks at concentrations near 50 μM due to selfquenching; (ii) measured fluorescence intensity and membrane potential are related by a nonlinear calibration curve sensitive to certain experimental details, including total concentration of dye and mitochondria in suspensions; and (iii) the time courses of membrane potential and electron transport fluxes following a perturbation (i.e. addition of ADP) significantly differ from observed transients in fluorescence intensity. These findings are consistent with the model predictions that mitochondria display a characteristic time of response to changes in substrate concentration of less than 0.1 s, corresponding to the time scale over which the rate of ATP synthesis changes to meet changes in ADP concentration.

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Section: Relationship Between Fluorescence and Transient Membrane Potmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Mitochondrial Inner Membrane Electrophysiology Assessed by Rhodamine-123 Transport and Fluorescence

et al. 2007

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“…1 bladder epithelial cells, and butyl-(4-hydroxybutyl)nitrosamineinduced mouse bladder transitional cell carcinoma retained a significant level of rhodamine 123 in dye-free medium even after 24 hr. Some of these tumorigenic cell lines (e.g., MB48, MB49, and RBC-1) still retain brightly stained mitochondria 4 days after incubation in dye-free medium, similar to the mitochondria of muscle cells. Most tumorigenic bladder epithelial cell lines tested retained mitochondrial fluorescence in dye-free medium for a prolonged period of time as assessed by fluorescence microscopy ( Table 2).…”

mentioning

confidence: 99%

“…The use of rhodamine and cyanine dyes in localizing mitochondria in living cells by epifluorescence microscopy has been described (1)(2)(3)(4)(5). The remarkable specificity appears to result from the high membrane potential (negative inside) across mitochondria on the one hand and the net positive charge carried by these dyes at physiological pH on the other.…”

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confidence: 99%

Unusual retention of rhodamine 123 by mitochondria in muscle and carcinoma cells.

Summerhayes

Lampidis

Bernal

et al. 1982

Proc. Natl. Acad. Sci. U.S.A.

303

200

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Mitochondria in cardiac muscle cells and myoblast-fused myotubes display unusually long (3-5 days) retention times of rhodamine 123, a mitochondria-specific fluorescent probe, in living cells. Among 50 keratin-positive carcinoma or transformed epithelial cell lines tested, mitochondria with prolonged rhodamine 123 retention are detected in most of the transitional cell carcinoma, adenocarcinoma, and chemical carcinogen-transformed epithelial cell lines and in some squamous cell carcinoma lines but notin any oat cell carcinoma lines. The presence of mitochondria having unusual dye retention may be useful for diagnosis and exploitable for chemotherapy of certain human carcinomas.

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Translocation of active mitochondria during hamster preimplantation embryo development studied by confocal laser scanning microscopy

1996

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Probing Mitochondria in Living Cells with Rhodamine 123

Cited by 169 publications

References 0 publications

Mitochondrial Inner Membrane Electrophysiology Assessed by Rhodamine-123 Transport and Fluorescence

Mitochondrial Inner Membrane Electrophysiology Assessed by Rhodamine-123 Transport and Fluorescence

Unusual retention of rhodamine 123 by mitochondria in muscle and carcinoma cells.

Translocation of active mitochondria during hamster preimplantation embryo development studied by confocal laser scanning microscopy

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