WHO WAS THE CUSTOMER?
This study was done by a group at a university, in a department of biomedicine.
WHAT IS THEIR AREA OF INTEREST?
CD4+ T cells (lymphocytes) and the bioenergy of their mitochondria.
WHAT DID THEY USE THE SCI-TIVE FOR?
Oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) were assessed under hypoxic conditions using the Seahorse XFe 96 metabolic extracellular flux analyzer placed inside the SCI-tive.
WHY WAS THIS IMPORTANT?
They wanted to understand what are the metabolic basis that enable T cell immune surveillance of normoxic and hypoxic tissue sites.
Effector memory (EM) CD4+ T cells recirculate between normoxic blood and hypoxic tissues to screen for their cognate antigens. Mitochondria are the key ATP-producing site of eukaryotic cells. How mitochondria of these cells that shuttle between normoxia and hypoxia can maintain bioenergetic efficiency and stably uphold antiapoptotic features is unknown. They wanted to resolve how mitochondria of these cells can maintain membrane potential and tonic mROS production throughout substantial cyclic changes in oxygen availability as this must pose a nonbioenergetic metabolic challenge to EM CD4+ T cells.
WHAT WAS THEIR STUDY MATERIAL?
CD4+ T cells (lymphocytes) and Jurkat cells (Jurkat cells are an immortalized T lymphocyte cell line that was originally obtained from the peripheral blood of a boy with T cell leukemia).
WHAT WAS THE EXPERIMENTAL SETUP?
For extracellular matrix flux analysis they assessed OCR and ECAR of CD4+ T cells under hypoxic conditions (1% O2) using the Seahorse XFe 96 metabolic extracellular flux analyzer placed in a SCI-tive). They also equilibrated unbuffered medium to hypoxia during 6 h and layered onto sorted CD4+ T cell subsets plated as described above.
They also assessed the total ATP use of sorted CD4+ T cell populations. Cells were incubated overnight at 37˚C in 21% O2/5% CO2 or 1% O2/5% CO2.
WHAT DID THE INVIVO ENABLE THEM TO DO THEY COULD NOT HAVE DONE OTHERWISE?
The Seahorse analyzer measures mitochondrial respiration and glycolysis as well as ATP production rate of live cells – cells that normally reside in low oxygen conditions which Seahorse cannot reproduce. Performing experiments in ambient oxygen conditions skewes the natural behaviour of the cells and produces irreproducible data. Only by placing the analyzer in a SCI-tive they could perform this experiment accurately and in in vivo -like oxygen atmosphere.
WHAT OTHER BAKER RUSKINN PRODUCTS THEY COULD BENEFIT FROM AND WHY?
RecO2ver – For the normoxic experiments they could in the future use Baker Ruskinn RecO2ver CO2 incubator
CondoCell – to ensure contamination free cell culture in an incubator or for taking the cells out of the SCI-tive for imaging with maintaining the workstation atmosphere.
OxyGenie – they could use OxyGenie to image the cells live in high resolution under low oxygen by initially growing cells inside the SCI-tive in the OneWell sample cups that can be sealed and thus retain the atmosphere from the SCI-tive. For longer imaging experiments they could use the OxyGenie platform to support the O2 atmosphere for longer imaging experiments