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Difference between revisions of "Cell count and normalization in HRR"

From Bioblast
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  Communicated by [[Gnaiger Erich]] 2020-09-18
  Communicated by [[Gnaiger Erich]] 2020-09-18
== Cell count and cell concentration ==
== Cell count and cell concentration ==


== Cell concentration from cell suspension to experimental chamber ==
== Cell concentration from cell suspension to experimental chamber ==
:::: Table 1. Definition of terms, symbols and units.
:::: {| class="wikitable"
|-
! Term !! Symbol !! = !! Definition !! Unit !! Comment
|-
| Volume of cell suspension σ || ''V''<sub>σ</sub> ||  ||  || mL || The symbol σ is used for 'suspension', distinguished from the symbol s for 'sample'. Frequently the sediment of cells obtained from centrifugation is re-suspended in the experimental medium (''e.g.'' MiR05 plus pyruvate) to obtain ''V''<sub>σ</sub>.
|-
| Cell count in ''V''<sub>σ</sub> || ''N''<sub>ce,σ</sub> || = || ''C''<sub>ce,σ</sub>∙''V''<sub>σ</sub> || mL || The total cell harvest available for respirometry and additional analyses of this sample of cells.
|-
| Cell-count concentration in cell suspension ''V''<sub>σ</sub> || ''C''<sub>ce,σ</sub> || = || ''N''<sub>ce,σ</sub>/''V''<sub>σ</sub> || mL ||
|-
| Subsample from ''V''<sub>σ</sub> for cell counting || ''V''<sub>σ→''N''</sub> ||  ||  || mL || For reliable normalization of respiration, ''V''<sub>σ→N</sub> should be equal or larger than 0.05 mL. Smaller volumes are taken as a subsample for counting, if the aim is merely to define a procedure of dilutions, with final subsamples taken after dilutions for cell-counting.
|-
| Subsample from ''V''<sub>σ</sub> for respirometry ''J'' || ''V''<sub>σ→''J''</sub> || = || ''V''<sub>σ</sub>-''V''<sub>σ→''N''</sub> || mL || .
|-
|  ||  ||  ||  ||  ||
|-
| Experimental chamber volume || ''V'' ||  ||  || mL || Effective aqueous volume in the experimental system (2.0 mL or 0.5 mL)
|-
| Experimental chamber volume + ''V''<sub>Stopper capillary</sub> || ''V''' || = || ''V''+''V''<sub>Stopper capillary</sub> || mL || 2.085 or 0.54 mL
|-
| Experimental chamber volume + ''V''<sub>Stopper capillary</sub> + subsample volume || ''V'''' || = || ''V'''+''V''<sub>subsamples</sub> || mL || Subsamples taken from the O2k chamber prior to closing the chamber.
|-
| Cell-count in experimental chamber || ''N''<sub>ce</sub> || = || ''N''·''U''<sub>ce</sub> || x || 10<sup>6</sup> x = 1 Mx
|-
| Cell-count concentration in experimental chamber || ''C''<sub>ce</sub> || = || ''N''<sub>ce</sub>/''V'' || Mx/mL || 10<sup>6</sup> x/mL = 1 Mx/mL = 10<sup>9</sup> x/L = 1 Gx/L
|-
|  ||  ||  ||  ||  ||
|-
| Volume of respirometric stock ''J'' || ''V''<sub>''J''</sub> || = || ''V''<sub>s→J</sub> + Δ''V''<sub>''J''</sub> || mL ||
|-
|}





Revision as of 20:53, 17 September 2020


high-resolution terminology - matching measurements at high-resolution


Cell count and normalization in HRR

Description

Cell count Nce is the number of cells, expressed in the unit [x] (1 Mx = 106 x). Normalization of respiratory rate by cell count yields oxygen flow IO2 expressed in units [amol·s-1·x-1] (=10-18 mol·s-1·x-1).

Abbreviation: Nce

Reference: BEC 2020.1 doi10.26124bec2020-0001.v1, Gnaiger 2020 MitoPathways

Communicated by Gnaiger Erich 2020-09-18

Cell count and cell concentration

Cell concentration from cell suspension to experimental chamber

Table 1. Definition of terms, symbols and units.
Term Symbol = Definition Unit Comment
Volume of cell suspension σ Vσ mL The symbol σ is used for 'suspension', distinguished from the symbol s for 'sample'. Frequently the sediment of cells obtained from centrifugation is re-suspended in the experimental medium (e.g. MiR05 plus pyruvate) to obtain Vσ.
Cell count in Vσ Nce,σ = Cce,σVσ mL The total cell harvest available for respirometry and additional analyses of this sample of cells.
Cell-count concentration in cell suspension Vσ Cce,σ = Nce,σ/Vσ mL
Subsample from Vσ for cell counting Vσ→N mL For reliable normalization of respiration, Vσ→N should be equal or larger than 0.05 mL. Smaller volumes are taken as a subsample for counting, if the aim is merely to define a procedure of dilutions, with final subsamples taken after dilutions for cell-counting.
Subsample from Vσ for respirometry J Vσ→J = Vσ-Vσ→N mL .
Experimental chamber volume V mL Effective aqueous volume in the experimental system (2.0 mL or 0.5 mL)
Experimental chamber volume + VStopper capillary V' = V+VStopper capillary mL 2.085 or 0.54 mL
Experimental chamber volume + VStopper capillary + subsample volume V'' = V'+Vsubsamples mL Subsamples taken from the O2k chamber prior to closing the chamber.
Cell-count in experimental chamber Nce = N·Uce x 106 x = 1 Mx
Cell-count concentration in experimental chamber Cce = Nce/V Mx/mL 106 x/mL = 1 Mx/mL = 109 x/L = 1 Gx/L
Volume of respirometric stock J VJ = Vs→J + ΔVJ mL


MitoPedia methods: Respirometry 


MitoPedia O2k and high-resolution respirometry: DatLab, O2k-Respirometry, O2k-FluoRespirometry