Difference between revisions of "FCCP"

Latest revision as of 10:55, 2 August 2021

high-resolution terminology - matching measurements at high-resolution

FCCP

Description

FCCP (Carbonyl cyanide p-trifluoro-methoxyphenyl hydrazone, C₁₀H₅F₃N₄O) is a protonophore or uncoupler: added at uncoupler concentration U_c; c is the optimum uncoupler concentration in titrations to obtain maximum mitochondrial respiration in the noncoupled state of ET capacity.

Abbreviation: FCCP

Reference: Steinlechner-Maran 1996 Am J Physiol Cell Physiol, Gnaiger_2008_POS

CCCP versus FCCP

Bioblast alert 2013(02): We replace FCCP by the less expensive and equally effective CCCP in our experiments with high-resultion respirometry (HRR).

Application in HRR

FCCP: Carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone ( C₁₀H₅F₃N₄O), Sigma-Aldrich: C2920, store at 4 °C, CAS: 370-86-5, M = 254.17 g·mol^-1

Hazard statements: H302, H314, H317, H413; harmful if swallowed, causes severe skin burns and eye damage, may cause an allergic skin reaction, may cause long-lasting harmful effects to aquatic life

Ethanol stock solutions may be stored in 5 mL glass vials (instead of freezing 0.2 mL portions in small vials). Use subsamples during experiments to avoid contamination of the storage solution. Chemicals stored in the fridge or freezer should be allowed to reach room temperature before opening.

Preparation of 1 mM stock solution (dissolved in ethanol abs.) for 2-mL O2k-chamber:

Weigh 1.27 mg of FCCP into a glass vial.
Dissolve in 5 mL ethanol.
Divide into 0.2 mL portions.
Store in glass vials at -20 °C.

O2k manual titrations MiPNet09.12 O2k-Titrations

Titration volume (2-mL O2k-chamber): 1 µL (or 0.5 µL) steps using a 10 µL syringe.
Final concentration: 0.5 µM (or 0.25 µM) steps.

Preparation of 0.25 mM stock solution (dissolved in ethanol abs.) for 0.5-mL O2k-chamber:

Take 100 µL of 1 mM stock solution.
Dilute with 300 µL ethanol.

» O2k manual titrations MiPNet09.12 O2k-Titrations

Titration volume (0.5-mL O2k-chamber): 1 µL (or 0.5 µL) steps using a 10 µL Hamilton syringe.
Final concentration: 0.5 µM (or 0.25 µM) steps.

Frequently, a more diluted FCCP stock is necessary for reducing the concentration steps in manual titrations. In this case prepare more diluted stock solution (ditution 2-10x) from your storage solution. The highest accuracy of uncoupler titrations is achieved by titrations with the TIP2k at high concentrations of the stock solution. ^[1]

Bioblast links: Uncoupling - >>>>>>> - Click on [Expand] or [Collapse] - >>>>>>>

Specific

» Artefacts by single dose uncoupling

» ATP synthase

» CCCP

» Coupling-control protocol

» DNP

» Dyscoupled respiration

» FCCP

» Is respiration uncoupled - noncoupled - dyscoupled?

» Noncoupled respiration: Discussion

» Uncoupler

» Uncoupled respiration - see » Noncoupled respiration

» Uncoupling proteins

» Uncoupling protein 1

» Uncoupler titrations - Optimum uncoupler concentration

Respiratory states and control ratios

» Biochemical coupling efficiency

» Coupling-control state

» Electron-transfer-pathway state

» Electron-transfer pathway

ET capacity

» E-L coupling efficiency

» Flux control efficiency

» Flux control ratio

» LEAK-control ratio

» LEAK respiration

» Noncoupled respiration

» OXPHOS

» OXPHOS capacity; » State 3

» OXPHOS-control ratio, P/E ratio

» Respiratory acceptor control ratio

» ROUTINE-control ratio

» ROUTINE respiration

» ROUTINE state

» State 3u

» State 4

» Uncoupling-control ratio UCR

Gnaiger E et al ― MitoEAGLE Task Group (2020) Mitochondrial physiology. Bioenerg Commun 2020.1. https://doi.org/10.26124/bec:2020-0001.v1
Gnaiger E (2020) Mitochondrial pathways and respiratory control. An introduction to OXPHOS analysis. 5th ed. Bioenerg Commun 2020.2. https://doi.org/10.26124/bec:2020-0002

General (alphabetical order)

» Adenine nucleotide translocase

» Adenylates

» Electron transfer pathway

» Mitochondrial preparations

» mt-membrane potential

» Oxygen flux

» Phosphorylation system

» Proton leak

» Proton slip

» TIP2k

Other keyword lists

» Template:Keywords: Force and membrane potential

↑ Gnaiger E (2008) Polarographic oxygen sensors, the oxygraph and high-resolution respirometry to assess mitochondrial function. In: Mitochondrial dysfunction in drug-induced toxicity (Dykens JA, Will Y, eds) John Wiley:327-52. »Bioblast link«

MitoPedia topics: Uncoupler

[1] Gnaiger E (2008) Polarographic oxygen sensors, the oxygraph and high-resolution respirometry to assess mitochondrial function. In: Mitochondrial dysfunction in drug-induced toxicity (Dykens JA, Will Y, eds) John Wiley:327-52. »Bioblast link«

[1]

@@ Line 9: / Line 9: @@
 == Application in [[HRR]] ==
+{{Chemical_description
+|abbr=FCCP
+|trivial name=Carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone
+|complete name=
+|chem formula=C<sub>10</sub>H<sub>5</sub>F<sub>3</sub>N<sub>4</sub>O
+|molar mass=254.17
+|vendor=Sigma-Aldrich
+|product number=C2920
+|store at=4 °C
+|sensitivity=
+|cas=370-86-5
+|h statements=H302, H314, H317, H413
+|h info=harmful if swallowed, causes severe skin burns and eye damage, may cause an allergic skin reaction, may cause long-lasting harmful effects to aquatic life
+}}<!--:::: '''FCCP''' (Carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone, C<sub>10</sub>H<sub>5</sub>F<sub>3</sub>N<sub>4</sub>O); Sigma C 2920, 10 mg, store at 4 °C: M = 254.17 g·mol<sup>-1</sup>)-->
+:::: Ethanol stock solutions may be stored in 5 mL glass vials (instead of freezing 0.2 mL portions in small vials). Use subsamples during experiments to avoid contamination of the storage solution. Chemicals stored in the fridge or freezer should be allowed to reach room temperature before opening.
+<!--:::: <span style="color:#8B008B"> '''Caution:''' Toxic!</span>-->
-:::: '''FCCP''' (Carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone, C<sub>10</sub>H<sub>5</sub>F<sub>3</sub>N<sub>4</sub>O); Sigma C 2920, 10 mg, store at 4 °C: M = 254.17 g·mol<sup>-1</sup>)
-:::: Ethanol stock solutions may be stored in 5 ml glass vials (instead of freezing 0.2 mL portions in small vials). Use subsamples during experiments to avoid contamination of the storage solution. Chemicals stored in the fridge or freezer should be allowed to reach room temperature before opening.
-:::: <span style="color:#8B008B"> '''Caution:''' Toxic!</span>
+::: '''Preparation of 1 mM stock solution''' (dissolved in ethanol abs.) for '''2-mL O2k-chamber''':
-::: '''Preparation of 1 mM stock solution''' (dissolved in 99.9% ethanol) for '''2-mL O2k-chamber''':
 ::::# Weigh 1.27 mg of FCCP into a glass vial.
-::::# Dissolve in 5 ml ethanol.
+::::# Dissolve in 5 mL ethanol.
-::::# Divide into 0.2 ml portions.
+::::# Divide into 0.2 mL portions.
 ::::# Store in glass vials at -20 °C.
 ::: '''O2k manual titrations'''  [[MiPNet09.12 O2k-Titrations]]
-::::* Titration volume ('''2-mL O2k-chamber'''): 1 µl (or 0.5 µl) steps using a 10 µl syringe.
+::::* Titration volume ('''2-mL O2k-chamber'''): 1 µL (or 0.5 µL) steps using a 10 µL syringe.
 ::::* Final concentration: 0.5 µM (or 0.25 µM) steps.
-:::: '''Preparation of 0.25 mM stock solution''' (dissolved in Ethanol) for '''0.5-mL O2k-chamber''':
+:::: '''Preparation of 0.25 mM stock solution''' (dissolved in ethanol abs.) for '''0.5-mL O2k-chamber''':
 ::::# Take 100 µL of 1 mM stock solution.
@@ Line 39: / Line 51: @@
-:::: Frequently, a more diluted FCCP stock is necessary for reducing the concentration steps in manual titrations. In this case prepare more diluted stocksolution (ditution 2-10x) from your storage solution. The highest accuracy of uncoupler titrations is achieved by titrations with the [[TIP2k]] at high concentrations of the stock solution. <ref> Gnaiger E (2008) Polarographic oxygen sensors, the oxygraph and high-resolution respirometry to assess mitochondrial function. In: Mitochondrial dysfunction in drug-induced toxicity (Dykens JA, Will Y, eds) John Wiley:327-52. [[Gnaiger_2008_POS |»Bioblast link«]]</ref>
+:::: Frequently, a more diluted FCCP stock is necessary for reducing the concentration steps in manual titrations. In this case prepare more diluted stock solution (ditution 2-10x) from your storage solution. The highest accuracy of uncoupler titrations is achieved by titrations with the [[TIP2k]] at high concentrations of the stock solution. <ref> Gnaiger E (2008) Polarographic oxygen sensors, the oxygraph and high-resolution respirometry to assess mitochondrial function. In: Mitochondrial dysfunction in drug-induced toxicity (Dykens JA, Will Y, eds) John Wiley:327-52. [[Gnaiger_2008_POS |»Bioblast link«]]</ref>
 {{Keywords: Uncoupling}}