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Measuring oxygen levels in Caco-2 cultures.

ABSTRACT: Measuring oxygen levels in three different systems of Caco-2 cell culture.Caco-2 cells were cultured in three different systems, using conventional polystyrene 24-well plates, special 24-well gas permeable plates, or on membrane inserts in conventional plates. Optical sensor spots were used to measure dissolved O2 levels in these cultured cells over the course of 6 days under normoxia (143 mmHg) and for 6 hours under hypoxia (7 mmHg). Western blot analysis was used to determine the protein levels of hypoxia-inducible factor 1? (HIF-1?) in the different cultures.All culture systems displayed lower O2 levels over time than expected when cultured under normoxia conditions. On average, O2 levels reached as low as 25 mmHg in 24-well plates but remained at 97 and 117 mmHg in gas permeable plates and membrane inserts, respectively. Under hypoxia, 1 mL cell cultures equilibrated to 7 mmHg O2 within the first 60 minutes and dropped to 0.39 and 0.61 mmHg O2 in 24-well and gas permeable plates, respectively, after the 6-hour incubation period. Cultures in membrane inserts did not equilibrate to 7 mmHg by the end of the 6-hour incubation period, where the lowest O2 measurements reached 23.12 mmHg. Western blots of HIF-1? protein level in the whole cell lysates of the different Caco-2 cultures revealed distinct stabilization of HIF-1? after hypoxic incubation for 1, 2, and 4 hours in 24-well plates as well as gas permeable plates. For membrane inserts, notable HIF-1? was seen after 4 hours of hypoxic incubation.Cellular oxygen depletion was achieved in different hypoxic Caco-2 culture systems. However, different oxygen levels comparing different culture systems indicate that O2 level should be carefully considered in oxygen-dependent experiments.

SUBMITTER: Zeitouni NE

PROVIDER: S-EPMC5045089 | biostudies-other | 2015

REPOSITORIES: biostudies-other

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Measuring oxygen levels in Caco-2 cultures.

Zeitouni Nathalie E NE Fandrey Joachim J Naim Hassan Y HY von Köckritz-Blickwede Maren M

Hypoxia (Auckland, N.Z.) 20151009

<h4>Purpose</h4>Measuring oxygen levels in three different systems of Caco-2 cell culture.<h4>Methods</h4>Caco-2 cells were cultured in three different systems, using conventional polystyrene 24-well plates, special 24-well gas permeable plates, or on membrane inserts in conventional plates. Optical sensor spots were used to measure dissolved O<sub>2</sub> levels in these cultured cells over the course of 6 days under normoxia (143 mmHg) and for 6 hours under hypoxia (7 mmHg). Western blot analy ...[more]

PMID: 27774482

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Project description:BackgroundDuring the COVID-19 pandemic, there have been concerns regarding potential bias in pulse oximetry measurements for people with high levels of skin pigmentation. We systematically reviewed the effects of skin pigmentation on the accuracy of oxygen saturation measurement by pulse oximetry (SpO2) compared with the gold standard SaO2 measured by CO-oximetry.MethodsWe searched Ovid MEDLINE, Ovid Embase, EBSCO CINAHL, ClinicalTrials.gov, and WHO International Clinical Trials Registry Platform (up to December 2021) for studies with SpO2-SaO2 comparisons and measuring the impact of skin pigmentation or ethnicity on pulse oximetry accuracy. We performed meta-analyses for mean bias (the primary outcome in this review) and its standard deviations (SDs) across studies included for each subgroup of skin pigmentation and ethnicity and used these pooled mean biases and SDs to calculate accuracy root-mean-square (Arms) and 95% limits of agreement. The review was registered with the Open Science Framework ( https://osf.io/gm7ty ).ResultsWe included 32 studies (6505 participants): 15 measured skin pigmentation and 22 referred to ethnicity. Compared with standard SaO2 measurement, pulse oximetry probably overestimates oxygen saturation in people with the high level of skin pigmentation (pooled mean bias 1.11%; 95% confidence interval 0.29 to 1.93%) and people described as Black/African American (1.52%; 0.95 to 2.09%) (moderate- and low-certainty evidence). The bias of pulse oximetry measurements for people with other levels of skin pigmentation or those from other ethnic groups is either more uncertain or suggests no overestimation. Whilst the extent of mean bias is small or negligible for all subgroups evaluated, the associated imprecision is unacceptably large (pooled SDs > 1%). When the extent of measurement bias and precision is considered jointly, pulse oximetry measurements for all the subgroups appear acceptably accurate (with Arms < 4%).ConclusionsPulse oximetry may overestimate oxygen saturation in people with high levels of skin pigmentation and people whose ethnicity is reported as Black/African American, compared with SaO2. The extent of overestimation may be small in hospital settings but unknown in community settings. REVIEW PROTOCOL REGISTRATION: https://osf.io/gm7ty.

Dataset Information

Measuring oxygen levels in Caco-2 cultures.

Publications

Measuring oxygen levels in Caco-2 cultures.

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