Attached cells

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Bioblasts - Richard Altmann and MiPArt by Odra Noel
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Attached cells

Description

Many cell types are grown in culter as attached cells, such as endothelial or neuronal cells in a monolayer.

Template NextGen-O2k.jpg
O2k-Open Support


MitoPedia O2k and high-resolution respirometry: O2k-Open Support 

HRR with attached cells

The Oroboros O2k is principally designed to measure respiration in non-adherent, suspended samples, i.e., suspensions of cells or mitochondria, tissue homogenates or permeabilized muscle fibers. However, successful efforts have been made to apply the O2k for measurements on adherent cells. Specifically, the lab of Gregory Brewer developed techniques for high-resolution respirometry with the O2k using attached neuronal cells:


Fragile samples

The rigorous stirring required for high-resolution respirometry causes problems for organisms / samples that are easily damaged by the mechanical force, such as fish eggs or beads used to grow adherent cells.
Presently an innovation is in preparation and will be available as a new product: a stainless steel mesh is used to maintain the sample separate from the stirrer compartment, without interfering with gentle mixing of the aqueous solution between the two compartments. Samples must be large enough not to pass through the 0.1 mm pore size of the mesh.
  • Dias C, Lourenco CF, Barbosa RM, Laranjinha J, Ledo A (2018) Analysis of respiratory capacity in brain tissue preparations: high-resolution respirometry for intact hippocampal slices. Anal Biochem 551:43-50. - »Bioblast link« - Original publication on the tissue holder with brain slizes.

Smaller stirring bars?

  • Question: Would smaller stirring bars help to minimize damage (combined with lower stirring speed)? Would they be available?
  • Answer: we are not sure if smaller stirring bars would help to avoid damage of fragile samples. In the early phase of the development of the O2k many different geometries and sizes of stirring bars were tested. In any case, commercially available Teflon (R) coated stirring bars cannot be used for high-resolution respirometry due to the high oxygen solubility of Teflon and the corresponing unacceptably high O2 background flux. The possibility of reducing the stirring speed can be evaluated by observing the noise and the [[stirrer test |response time] of the signal.


MitoPedia O2k and high-resolution respirometry: O2k-Open Support