Toward Best Practices for Controlling Mammalian Cell Culture Environments

May combine two claims.

Levels of CO2/HCO3− readily diffuse across cell membranes to moderate intracellular pH, act as metabolic inhibitors, may induce complex transcriptional responses, and signal other critical reactions.

“Besides the role of CO2 in affecting medium acid-base chemistry, levels of CO2/HCO3 − readily diffuse across cell membranes to moderate intracellular pH (Gutknecht et al., 1977), act as metabolic inhibitors, and may induce complex transcriptional responses (Cottier et al., 2012; Follonier et al., 2013), and signal other critical reactions (see, Blombach and Takors, 2015).”

• cites Gutknecht et al., 1977 sourceDisputed

  The Gutknecht citation applies to the clause about CO2/HCO3− crossing membranes and moderating intracellular pH, not the later claims about metabolic inhibition or transcription. The Introduction (p. 780) says CO2 gas diffuses across membranes but HCO3− cannot easily do so across most cell membranes; the Results (p. 786) likewise say HCO3− cannot readily cross the membrane. The paper does not establish moderation of intracellular pH. (LLM, conf=0.88)

  Checked by a reasoning model — may be wrong.

  Experimental — accuracy not yet measured
• cites Cottier et al., 2012 sourcePartly supported

  The Discussion (page not provided) reports that CO2 changes carbonic anhydrase expression through a transcription-factor-mediated pathway in yeast, supporting the cited claim about transcriptional responses. The Introduction describes CO2/HCO3− as important to fungal metabolism, but the passages do not support the adjacent claim that they act as metabolic inhibitors. The membrane-diffusion and other-signalling clauses have separate citations in the citing sentence. (LLM, conf=0.82)

  Experimental — accuracy not yet measured
• cites Blombach and Takors, 2015 sourceSupported

May combine two claims.

Cellular growth of three colorectal cell-lines (NCI-H747, DLD1, Caco2) was optimal at pH 7.4, but when medium pH deviated from 7.4 by >0.3 units all three cell lines exhibited reduced rates of proliferation.

“Michl and others (2019) showed that cellular growth of three colorectal cell-lines (NCI-H747, DLD1, Caco2) was optimal at pH 7.4, but when medium pH deviated from 7.4 by > 0.3 units all three cell lines exhibited reduced rates of proliferation.”

• cites Michl et al., 2019 sourcePartly supported

  The Results (Stability of CO2/HCO3− buffering, Fig. 2e; page unavailable) report that growth of NCI-H747, DLD1 and Caco2 was optimal near pH 7.4. They say the effect at pH 7.7 varied, with the strongest growth inhibition in NCI-H747; they do not establish a >0.3-unit threshold or explicitly state that all three lines had reduced proliferation beyond it. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

May combine two claims.

pH-buffering mechanisms such as Na+/H+ antiporters or histone deacetylation consume cellular energy and can alter gene transcription and reduce cellular growth through changes in the acetylation state of chromatin.

“However, such mechanisms (e.g., Na+/H+ antiporters or histone deacetylation) consume cellular energy and can alter gene transcription and reduce cellular growth through changes in the acetylation state of chromatin (Boron and Russell, 1983; Bowen and Levinson, 1984; Boron, 2004).”

• cites Boron and Russell, 1983 sourcePartly supported

  The opening summary (p. 373) reports that the squid axon's pH-regulating transport system requires ATP, supporting an energy requirement for the mechanism studied. The discussion (p. 397) identifies Na+/H+ exchange as a distinct pH-regulating system, but does not establish its energy requirement. The paper does not address histone deacetylation, chromatin acetylation, gene transcription, or cellular growth. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites Bowen and Levinson, 1984 sourceNot checked — no full text of the source
• cites Boron, 2004 sourceNot checked — no full text of the source

May combine two claims.

The effect of unstirred medium layers presumably determines the delivery of nutrients/growth factors and the removal of other metabolic waste products such as lactic acid, which can also act to directly and indirectly moderate environmental variation.

“The effect of unstirred medium layers also presumably determines the delivery of nutrients/growth factors and the removal of other metabolic waste products (e.g., lactic acid), which can also act to directly and indirectly moderate environmental variation (Michl et al., 2019).”

• cites Michl et al., 2019 sourcePartly supported

  Split into 2 atoms — 1 confirmed, 1 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  Experimental — accuracy not yet measured

May combine two claims.

HEPES activated lysosomal transcription factors in macrophages, inhibited the prion protein conversion in neural stem cells and affected their viability and differentiation.

“Briefly, HEPES activated lysosomal transcription factors in macrophages (Tol et al., 2018), inhibited the prion protein conversion in neural stem cells and affected their viability and differentiation (Delmouly et al., 2011).”

• cites Tol et al., 2018 sourceSupported
• cites Delmouly et al., 2011 sourceNot checked — no full text of the source

May combine two claims.

Experiments testing cell-line strains against anti-cancer compounds uncovered disparate drug responses, although the exact sources of instabilities in culture environments that promoted cell-line heterogeneity were not resolved.

“Further experiments testing the cell-line strains against anti-cancer compounds uncovered disparate drug responses, although the exact sources of instabilities in culture environments that promoted cell-line heterogeneity were not resolved (Ben-David et al., 2018).”

• cites Ben-David et al., 2018 sourcePartly supported

  The Functional consequences of genomic variation section reports markedly different responses to anti-cancer compounds across 27 MCF7 strains. The Discussion and Sources of variation sections identify clonal selection under different culture conditions and ongoing genomic instability as contributors to heterogeneity. They do not establish the citing sentence’s additional claim that the exact sources of culture-related instability remained unresolved. No page numbers are available in our copy. (LLM, conf=0.82)

  Experimental — accuracy not yet measured

May combine two claims.

Exogenous buffer approaches can, in some cases, promote unpredictable changes in pH and introduce confounding artifacts.

“such approaches can, in some cases, promote unpredictable changes in pH and introduce confounding artifacts (Michl et al., 2019, see below).”

• cites Michl et al., 2019 sourceSupported

May combine two claims.

The majority of research papers rely on nominal set points and fail to directly verify and report environmental parameters.

“recent assessments show that the majority of research papers rely on nominal set points and fail to directly verify and report environmental parameters (Al-Ani et al., 2018; Michl et al., 2019; Klein et al., 2021b).”

• cites Al-Ani et al., 2018 sourcePartly supported

  The Results and discussion report that 94% of 200 mammalian-cell-culture papers omitted at least one critical oxygenation variable, and none attempted to calculate oxygen levels around the cells. The Introduction also says incubator settings are often assumed to represent oxygen delivered to cells. This supports the claim for oxygenation in the papers studied, but not its broader wording about research papers and environmental parameters generally; the assessment did not establish how many directly measured oxygen levels. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites Michl et al., 2019 sourcePartly supported

  The Discussion reports that only a small percentage of surveyed cell-culture articles provided necessary information about medium pH and buffering; only a third reported incubator CO2. The Results (“Monitoring culture medium pH under incubation”) also says that medium pH is not routinely monitored. This supports the claim for cell-culture pH, but the paper does not establish it for environmental parameters generally. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites Klein et al., 2021bNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Classified as probably not a substantial claim.

The CO2/HCO3− buffer system is the primary physiological buffering system in mammalian fluids.

“Although the CO2/HCO3 − buffer system is indeed the primary physiological buffering system in mammalian fluids (Boron, 2004; Michl et al., 2019)”

• cites Boron, 2004 sourceNot checked — no full text of the source
• cites Michl et al., 2019 sourceSupported

May combine two claims.

Perturbations to dissolved O2 levels in culture medium can induce cells to switch away from oxidative phosphorylation towards anaerobic glycolysis, leading to large accumulations of lactic acid that force medium acidification.

“For instance, perturbations to dissolved O2 levels in culture medium can induce cells to switch away from oxidative phosphorylation towards anaerobic glycolysis (Wolff et al., 1993), leading to large accumulations of lactic acid that force medium acidification (Michl et al., 2019).”

• cites Wolff et al., 1993 sourceNot checked — no full text of the source
• cites Michl et al., 2019 sourceSupported

May combine two claims.

Another post-publication analysis reported that less than half of studies published in Cancer Research and Nature in the third quarter of 2017 described the brand of medium, and only one-tenth declared the medium-buffering regime.

“Another post-publication analysis reported that less than half of studies published in Cancer Research and Nature in the third quarter of 2017 described the brand of medium, and only one-tenth declared the medium-buffering regime (Michl et al., 2019).”

• cites Michl et al., 2019 sourceSupported

May combine two claims.

Deoxygenation can activate the hypoxia-inducible factor (HIF) transcription system, which triggers the expression of most genes responsible for cellular adaptation to varying O2 levels.

“deoxygenation can also activate the hypoxia-inducible factor (HIF) transcription system, which triggers the expression of most genes responsible for cellular adaptation to varying O2 levels (Semenza et al., 1991; Wang et al., 1995; Semenza, 2012).”

• cites Semenza et al., 1991 sourceNot checked — no full text of the source
• cites Wang et al., 1995 sourcePartly supported

  The paper reports that reduced O2 induces HIF-1 and that HIF-1 participates in hypoxia-responsive transcription (opening section; Results and Discussion, page not specified in our copy). It identifies EPO and several glycolytic genes as examples, but does not establish that HIF triggers expression of most genes responsible for cellular adaptation to changing O2 levels; it says HIF-1 may play a general role. (LLM, conf=0.87)

  Experimental — accuracy not yet measured
• cites Semenza, 2012 sourcePartly supported

  The Hypoxia-Inducible Factors section (page not specified) reports that HIF-1 levels rise as oxygen falls and that hundreds of genes show HIF-1-dependent responses to hypoxia. The Introduction calls HIFs master regulators of oxygen homeostasis, but the copy does not establish that they trigger expression of most genes responsible for cellular adaptation. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

May combine two claims.

Supra-physiological osmolarity can directly affect cell membrane tension and volume, but can also moderate how cells respond to other environmental parameters.

“Supra-physiological osmolarity can directly affect cell membrane tension and volume (Pedersen et al., 2013), but can also moderate how cells respond to other environmental parameters (Dezengotita et al., 1998).”

• cites Pedersen et al., 2013 sourcePartly supported

  The 'Physiology of epithelial transport and role of cell volume' section says that altered extracellular osmolarity creates osmotic gradients and discusses associated changes in cell volume. It does not specifically address supra-physiological osmolarity or state that osmolarity directly changes cell membrane tension. The later clause about responses to other environmental parameters is attributed to Dezengotita et al., not to this source. (LLM, conf=0.82)

  Experimental — accuracy not yet measured
• cites Dezengotita et al., 1998 sourceNot checked — no full text of the source

May combine two claims.

In batch culture set-ups, researchers often select culture medium that contains a pH indicator dye such as Phenol Red to guide the renewal of cell medium, but medium color changes assessed "by eye" can lead to undetected pH declines.

“In batch culture set-ups, researchers often select culture medium that contains a pH indicator dye [e.g., Phenol Red (PhR)] to guide the renewal of cell medium, but medium color changes assessed “by eye” can lead to undetected pH declines (Michl et al., 2019).”

• cites Michl et al., 2019 sourcePartly supported

  The Results section, “Monitoring culture medium pH under incubation” (page unavailable), says Phenol Red is routinely included to assess medium acidity and that assessment can be done ‘by eye,’ while absorbance provides a more quantitative reading. It also warns that medium pH is not routinely monitored despite possible acid production. It does not say that visual assessment guides medium renewal in batch cultures or establish that it leads to undetected pH declines. (LLM, conf=0.85)

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Chemostat and perfusion systems were first described in 1950 for use in bacterial cultures and later adopted for mammalian cells in 1961.

“Chemostat and perfusion systems were first described in 1950 for use in bacterial cultures (Novick and Szilard, 1950a; b) and later adopted for mammalian cells in 1961 (Cohen and Eagle, 1961).”

• cites Novick and Szilard, 1950aNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• cites Cohen and Eagle, 1961 sourceSupported

May combine two claims.

Titration of acids and bases (including HCl, NaOH, and NVBs) introduces osmolytes (Na+, Cl−) to cell medium and can result in substantial changes to medium osmolarity by >10%.

“The titration of acids and bases (including HCl, NaOH, and NVBs) introduces osmolytes (Na+, Cl−) to cell medium and can result in substantial changes to medium osmolarity by > 10% (Michl et al., 2019).”

• cites Michl et al., 2019 sourcePartly supported

  The Results section (‘Effect of NVBs on the stability of medium pH’) says that titrating non-volatile buffers with HCl or NaOH introduces Na+ and Cl− and can substantially increase osmolality. Its example estimates an increase of approximately 10%, not greater than 10%, so the stated threshold is not established by that passage. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

May combine two claims.

NVB addition may not fully prevent pH declines in standard batch cultures and may lead to unexpected pH changes when interacting with the HCO3−/CO2 buffering system, although predictable pH levels can be obtained when appropriate protocols are used.

“Recent assessments also shows that NVB addition may not fully prevent pH declines in standard batch cultures and may lead to unexpected pH changes when interacting with the HCO3 −/CO2 buffering system, although predictable pH levels can be obtained when appropriate protocols are used (see, Michl et al., 2019).”

• cites Michl et al., 2019 sourceSupported

Probably not a substantial claim: may not pass S3 (load-bearing).

Acquiring a bioreactor is high in cost.

“E.g., acquirement of a bioreactor is high in cost”

May combine two claims.

The reported median decline in pH was 0.43 units, but in some particularly extreme cases, cell metabolic activity promoted pH reductions that approached one pH unit and dissolved O2 decreased down to 0.95%.

“The reported median decline in pH was 0.43 units, but in some particularly extreme cases, cell metabolic activity promoted pH reductions that approached one pH unit (Eagle, 1971) and dissolved O2 decreased down to 0.95% (Vallejos et al., 2010).”

• cites Eagle, 1971 sourceNot checked — no full text of the source
• cites Vallejos et al., 2010 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

Bioreactor systems, by design, typically require consistent monitoring of the controlled parameters via a feedback loop to achieve the desired control.

“Indeed, these systems, by design, typically require consistent monitoring of the controlled parameters via a feedback loop to achieve the desired control.”

May combine two claims.

In extreme cases, variations in culture conditions may resemble levels consistent with hypercapnia and hypoxia rather than conditions typical of in vivo extracellular fluids, although in vivo environments vary considerably among selected tissues.

“In such extreme cases, variations in culture conditions may resemble levels consistent with hypercapnia and hypoxia rather than conditions typical of in vivo extracellular fluids, although in vivo environments vary considerably among selected tissues (Ast and Mootha, 2019).”

• cites Ast and Mootha, 2019 sourceNot checked — no full text of the source

May combine two claims.

Even when protocols are declared, there is an unfortunate prevalence of papers stating "as previously described by ref. (x)," which often leads to a chain of citations that generate confusion as to the specific procedures, reagents, and materials involved.

“Even when protocols are declared, there is an unfortunate prevalence of papers stating, “as previously described by ref. (x),” which often leads to a chain of citations that generate confusion as to the specific procedures, reagents, and materials involved (Freedman et al., 2015).”

• cites Freedman et al., 2015 sourceNot checked — no full text of the source

May combine two claims.

A recent synthesis examining the lack of monitoring and reporting sub-sampled 688 papers published between 2014 and 2019 and found that most papers reported the medium manufacturer, but only one third reported the type of culture system utilized and 42% reported temperature and CO2 incubator settings.

“A recent synthesis examining this problem sub-sampled 688 papers published between 2014 and 2019 and found that most papers reported the medium manufacturer, but only one third reported the type of culture system utilized and 42% reported temperature and CO2 incubator settings (Klein et al., 2021b).”

• cites Klein et al., 2021bNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

May combine two claims.

Most commercial media contain buffering systems that act only to regulate pH, whereas levels of dissolved O2 and CO2 are regulated by atmosphere re-equilibration.

“Most commercial media contain buffering systems that act only to regulate pH, whereas levels of dissolved O2 and CO2 are regulated by atmosphere re-equilibration.”

May combine two claims.

Standard batch culture systems undergo substantial environmental changes owing to cell metabolic activity, with pH declines reaching 0.9 units, O2 levels declining down to 0.95%, and CO2 values reaching up to 10.45%.

“Standard batch culture systems undergo substantial environmental changes owing to cell metabolic activity (Balin et al., 1976; Naciri et al., 2008; Vallejos et al., 2010; Pradhan et al., 2012; Al-Ani et al., 2018; Michl et al., 2019), with pH declines reaching 0.9 units, O2 levels declining down to 0.95%, and CO2 values reaching up to 10.45% (Klein et al., 2021b).”

• cites Klein et al., 2021bNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

May combine two claims.

Bioreactors provide the best capacity to control environmental conditions, but are most costly in the context of capital investment, maintenance, and operations.

“Bioreactors provide the best capacity to control environmental conditions, but are most costly in the context of capital investment, maintenance, and operations.”

May combine two claims.

Variations in relative humidity and media evaporation result in changes to osmolarity as well as solute and gas concentrations, that in turn, affect diffusion.

“Since variations in both factors result in changes to osmolarity as well as solute and gas concentrations, that in turn, affect diffusion.”

Probably not a substantial claim: may not pass S4 (based). May combine two claims.

Traditional bioreactor systems are ideal for cells culture in suspension, while attached cell monolayers require different solutions.

“While traditional bioreactor systems are ideal for cells culture in suspension, attached cell monolayers require different solutions.”

Probably not a substantial claim: may not pass S2 (checkable), S4 (based). May combine two claims.

Improved characterization and control over environmental conditions in cell cultures will enhance the reliability of experimental findings and the confidence in their translation to clinical applications.

“Ultimately, improved characterization and control over environmental conditions in cell cultures will enhance the reliability of experimental findings and the confidence in their translation to clinical applications”

May combine two claims.

Strengthening reporting requirements and standards will likely place greater emphasis on cell culturing environments and, in turn, likely enhance the reproducibility of cell-based experiments and their relevance to the in vivo environment.

“Strengthening reporting requirements and standards will likely place greater emphasis on cell culturing environments and, in turn, likely enhance the reproducibility of cell-based experiments as well as their relevance to the in vivo environment.”

Probably not a substantial claim: may not pass S2 (checkable).

Considering how niche in vivo environments affect experimental outcomes could increase the robustness of experiments and the likelihood that findings have relevance to focal in vivo compartments.

“By considering how niche in vivo environments affect experimental outcomes, researchers could further increase the robustness of their experiments and increase the likelihood that findings have relevance to focal in vivo compartments (Ast and Mootha, 2019).”

• cites Ast and Mootha, 2019 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S4 (based). May combine two claims.

Many bioreactor systems lack flexibility in the number of biological replicates and the volume of culture media (often larger than that of batch culture) that can be manipulated, which translates into greater time and monetary costs.

“Importantly, many bioreactor systems lack flexibility in the number of biological replicates and the volume of culture media (often larger than that of batch culture) that can be manipulated, which translates into greater time and monetary costs.”

May combine two claims.

O2 levels vary across human tissues and range from 13% O2 in the lung-pulmonary vein to 1-3% O2 in the uterus.

“For instance, O2 levels vary across human tissues and range from 13% O2 in the lung-pulmonary vein to 1–3% O2 in the uterus (Ast and Mootha, 2019).”

• cites Ast and Mootha, 2019 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S2 (checkable), S4 (based).

Reporting initial and final values of key parameters in batch cultures will capture existing environmental variation affecting published findings, ensuring accurate interpretation of the reported results and improved reproducibility.

“This requirement should capture existing environmental variation affecting published findings, ensuring accurate interpretation of the reported results and improved reproducibility.”

Probably not a substantial claim: may not pass S4 (based).

Measuring systems capable of delivering the required precision and accuracy are available for all key environmental parameters.

“Measuring systems capable of delivering the required precision and accuracy are available for all key environmental parameters.”

Probably not a substantial claim: may not pass S2 (checkable).

Transitioning towards advanced culture systems capable of mimicking in vivo conditions requires attention to other chemical and physical factors known to program cell fate, in addition to environmental parameters.

“Transitioning towards advanced culture systems capable of mimicking in vivo conditions not only requires consideration of environmental parameters, but also necessitates attention to other chemical and physical factors known to program cell fate.”

Probably not a substantial claim: may not pass S2 (checkable), S4 (based).

The PRINCE reporting checklist will ensure standardized reporting of cell culture conditions.

“This will ensure much needed standardized reporting of cell culture conditions.”

May combine two claims.

Batch cell cultures are the most popular, inexpensive and scalable culture system, in terms of the possible number of replicates and treatments, with <1% of the published literature utilizing cybernetic bioreactor or chemostat culture systems (2014-2019).

“Indeed, batch cell cultures are the most popular, inexpensive and scalable culture system, in terms of the possible number of replicates and treatments, with <1% of the published literature utilizing cybernetic bioreactor or chemostat culture systems (2014–2019, Klein et al., 2021b).”

• cites Klein et al., 2021bNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

May combine two claims.

Hybridoma cells exposed to elevated CO2 conditions exhibited reduced growth rates when osmolarity was held constant at 361 mOsm kg−1, but cell growth rates further declined by 30% when medium osmolality was 415 mOsm kg−1.

“For instance, hybridoma cells exposed to elevated CO2 conditions exhibited reduced growth rates when osmolarity was held constant at 361 mOsm kg−1, but cell growth rates further declined by 30% when medium osmolality was 415 mOsm kg−1 (Dezengotita et al., 1998).”

• cites Dezengotita et al., 1998 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S4 (based). May combine two claims.

In bioreactor cultures, often only setpoints are reported without actually measuring environmental conditions in the medium, leading to unknown physiological relevance and low reproducibility.

“but often only setpoints are reported without actually measuring environmental conditions in the medium (mean ± SE) leading to unknown physiological relevance and low reproducibility.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

More sophisticated bioreactor culture systems were first introduced in the 1970s to culture mammalian cells and typically involve the automated control of temperature, gas addition (O2, CO2), and/or acids and bases to maintain set targets for temperature, dissolved gases and pH.

“More sophisticated bioreactor culture systems were first introduced in the 1970s to culture mammalian cells (Knazek et al., 1972; Fouron, 1987) and typically involve the automated control of temperature, gas addition (O2, CO2), and/or acids and bases to maintain set targets for temperature, dissolved gases and pH.”

• cites Knazek et al., 1972 sourceNot checked — no full text of the source
• cites Fouron, 1987 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

Batch cultures are typically maintained in incubators that maintain temperature (typically 37°C) and guarantee a CO2 level (typically 5%) in the atmosphere.

“Batch cultures are typically maintained in incubators that maintain temperature (typically 37°C) and guarantee a CO2 level (typically 5%) in the atmosphere.”

May combine two claims.

A reasonable degree of control over environmental conditions is achievable in routine culture systems, but requires consideration of workflow factors that can, directly and indirectly, promote environmental drift.

“A reasonable degree of control over environmental conditions is achievable in routine culture systems, but requires consideration of workflow factors that can, directly and indirectly, promote environmental drift (Figure 2).”

• cites Figure 2Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• AssumptionRests on an assumption: The most common approach to in vitro cell culture is standard batch culture, in which cells are grown either in suspension or as adherent monolayers in standard media within tissue culture flasks.

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Such systems range in cost, from moderately priced sensors for temperature, pH, O2, and salinity to the more expensive sensing equipment required for monitoring dissolved CO2, which often require complex calibration protocols.

“Such systems range in cost, from moderately priced sensors for temperature, pH, O2, and salinity to the more expensive sensing equipment required for monitoring dissolved CO2, which often require complex calibration protocols (Supplementary Table S1).”

• cites Supplementary Table S1Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

May combine two claims.

Environmental stability is most easily achieved in advanced bioreactor culture systems, whereas achieving stability in routine batch culture systems is most challenging, with perfusion systems (and chemostats) providing intermediate solutions.

“Environmental stability is most easily achieved in advanced bioreactor culture systems, whereas achieving stability in routine batch culture systems is most challenging, with perfusion systems (and chemostats) providing intermediate solutions (Table 1).”

• cites Table 1Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• AssumptionRests on an assumption: The most common approach to in vitro cell culture is standard batch culture, in which cells are grown either in suspension or as adherent monolayers in standard media within tissue culture flasks.