The document is a presentation by Justin Croft on physoxic research. It discusses oxygen terminology, how cells sense oxygen levels via the HIF-1 transcription factor, and options for hypoxia and physoxia research equipment from Scintica Instrumentation. It recommends considering features like partial pressure measurements, ease of use, and whether the system meets short or long term study needs when choosing equipment.

1. Justin Croft, MSc
Product Manager
Scintica Instrumentation
Phone: +1 (519) 914 5495
Jcroft@scintica.com
A Roadmap to Physoxic
Research: Where to Start
and What to Consider

2. Scintica Instrumentation provides sales, service
and support for life science research products in the US,
Canada and Europe.
Centrifuge
Anesthesia Equipment
Hypoxia Chambers
Automatic Colony Counter
Isolated Heart
Isolated Muscle Tissue
Isolated Nerve Tissue
Noninvasive Blood Flow
Noninvasive Blood Pressure
Rodent Surgical Monitoring
Cell Counter
Wire Myography
CO2 Incubator
Oxygen Perfusion
PRECLINICAL RESEARCH
SOLUTIONS
2

3. Justin Croft, MSc
Product Manager
Scintica Instrumentation
Phone: +1 (519) 914 5495
Jcroft@scintica.com
A Roadmap to Physoxic
Research: Where to Start
and What to Consider

4. Audience Poll
4

5. 1. Basics of CO2 incubators
2. Oxygen terminology
3. Cellular oxygen and HIF-1 transcription factor
4. Hypoxia and physoxia research: What to consider and
the options available
Webinar Contents
5

6. • Air e.g. ‘dry air’ by volume percentage
contains
• 78.09% nitrogen (N2)
• 20.95% oxygen (O2)
• 00.93% argon (Ar)
• 00.04% mainly carbon dioxide (CO2)
and trace amount of other gases (Ne,
He, CH4, Kr, H2)
General Basics About Air And O 2
• Cell incubators
• Temperature mostly at 37°C for
mammalian cells
• 5% CO2 used to keep pH
• 70-90% humidity
• No O2 control
6

7. • Touch Screen Controls
• Fan-assisted air circulation for
uniformity and rapid recovery
• High temperature sterilization for easy
cleaning
• Accurate sensors for Measuring CO2
RWD D180-P CO2 Incubator
7

8. • CO2 incubators have lower internal oxygen than room air at
21%
• O2 levels around 18%
• Why?
• 5% CO2
• 70-90% Humidity
• Remaining volume of gas filled with ‘air’
Cell Culture In CO2 Incubators
8

9. • Normoxia not clearly defined though ‘assumed’ as to be the oxygen concentration at
sea level (i.e. 21%)
Oxygen Terminology
9

10. ORGANS AND PHYSOXIA
10

11. • Normoxia not clearly defined though ‘assumed’ as to be the oxygen concentration at
sea level (i.e. 21%)
• Hypoxia is medically defined as: “a deficiency of oxygen reaching the tissues of the
body.”
• In its extreme form, where oxygen is entirely absent, the condition is called anoxia
• Hyperoxia is when cells, tissues and organs are exposed to an excess supply of O2
• Relevance in relation to the tissues/organs in the body >> Physoxia
Oxygen Terminology
11

12. Research fields/subjects related to O2 levels
• Sleep apnea
• Cancer cell research
• Pulmonary research (COPD, asthma)
• Reactive Oxygen Species effects (ROS)
• Muscle physiology (endurance, general metabolism)
• Brain stroke, seizure
• Heart infarct (Ischemia / reperfusion)
• Embryonal development
Why Bother With Oxygen Levels?
Cancer Research UK
12

13. • Breathing pO2160mmHg
• Lung alveolar tissue ~100mmHg pO2
• Arterial blood carrying ~75- 100mmHg pO2
• Venous is between 30-40mmHg pO2
• What are the implications of maintaining cells
in vitro at higher pO2 141mmHg?
Cell / Tissue Exposure/ Access To O 2
Cancer Research UK
13

14. • ‘Normoxia’ ≠ ‘Physoxia’ and likely considered hyperoxic
• (Incubator air 141mmHg/18.9% vs. cells range ~40-100mmHg/ ~5-12% O2)
• How do cells ‘see’ as hypoxia / physoxia / hyperoxia
• Hypoxia Inducible Factor (HIF)-1 is sort of the oxygen sensor of the cell
Physoxia vs. Hypoxia: How Does A Cell Know
14

15. O2
CO2
HypoxiaNormoxia
PHD
pVHL
Ub
HIF1α Glycolytic enzymes, VEGF, EPO
HIF1α
―OHHIF1α HIF1α
HRE
HIF1βHIF1α
O2
CO2
HIF1α
PHD
X
HIF1β
Hypoxia Inducible Factor (HIF)-1α

16. • Matrix and barrier
function genes
• Inflammation
• Increase oxygen delivery
(EPO, Heme)
• Angiogenesis
HIF-1 Co-factor Driven Gene Expression Of:
• Vascular tone
• Reduction of oxygen
consumption
• Promote anaerobic metabolism
• Regulate cell proliferation and
apoptosis
16

17. Bordt et al. 2017
Practical Example in vivo vs. in vitro
• oxygen tension plays an important role in
cellular function
• in vitro environment should more closely
resembling in vivo conditions
• tissue oxygenation in particular brain
regions can vary
(Physoxia)
17

18. • How do I navigate this space?
• Which option is right for my needs?
• What does Scintica offers in this space?
Roadmap into Hypoxia and Physoxia research
18

19. • Look for industry leaders in the area
• Experienced and have seen it all
• Look for features that make your research more
reproducable
• Partial pressure measurements are more percise
• Look for features that make your life easier
• Ease of changing sensors or parts if needed
• Talk to experts that can help you figure it out
How Do I Navigate This Space
19

20. • Short term vs. long term studies?
• Do you need to incubate cells and for how long?
• How many plates do you need to incubate?
• Do you need to work on cells under controlled
oxygen conditions?
• Do you need to image cells, irradiate cells or
measure oxygen content?
• What are your goals?
Which Option Is Right For My Needs?
20

21. • OxyGenie
• PhO2x Box
• CondoCell
• HypoxyLab
• HypoxyLab Add-ons – Microscope, cell oxygen monitoring
21
Hypoxia/Physoxia Options from Scintica

22. • Super compact and efficient
• Portable tool, ideal for specific research
• Ideal for short term hypoxia studies
• Irradiation compatible
OxyGenie
22

23. 23

24. • Fits neatly into CO2 incubator shelving
• Easy navigation software for steady state
incubation or cycling
• Up to 3 boxes can be controlled by one gas
controller
• Best for short term studies
PhO2 x Box
24

25. • Constant Physiological culture
• Filtered, protected and contained environment for
you and your cells
• Utility for O2 and non-O2 culture
• Ideal for short term hypoxia studies or increased
long term storage when coupled with a workstation
CondoCell
25

26. 26

27. • Compact and efficient
• Delivers “true physoxia” using the partial pressure
of oxygen, imitating in vivo oxygen state
• Chamber rapidly equilibrates while consuming
less gas
• HEPA filtration scrubs the air in seconds
HypoxyLab
27

28. Lux 2 microscope
• Small microscope for live cell imaging and
monitoring
CondoCell
• Take cells out of system and keep at set
conditions while imaging.
HypoxyLab add-ons
28

29. • OXYLITE
• Non-oxygen consuming determination
• Oxygen (pO2)
• Temperature monitor
• Glass fiber probe
HypoxyLab add-ons
29

30. 30

31. • CO2 incubators have some inherent issues
• Oxygen should be considered in all cell research looking to provide context for
the in vivo condition
• When choosing a system one should look for a system that meets their current
needs and application goals.
Conclusion
31

32. Justin Croft, MSc
Product Manager
Scintica Instrumentation
Phone: +1 (519) 914 5495
Jcroft@scintica.com
Q&A
SESSION:
To ask a question, click the Q&A Button,
type your question and click send. Any
questions that are not addressed during
the live webinar will be answered
following the event.
Thank you for participating!

33. Audience Poll
33

34. Justin Croft, MSc
Product Manager
Scintica Instrumentation
Phone: +1 (519) 914 5495
Jcroft@scintica.com
Q&A
SESSION:
To ask a question, click the Q&A Button,
type your question and click send. Any
questions that are not addressed during
the live webinar will be answered
following the event.
Thank you for participating!

35. 35

36. minutes
HIF-1α RESPONSE O2 NORMOXIA TO ANOXIA
Jewel et al. 2001
36

37. HIF-1α RESPONSE TO DEOXYGENATING MEDIA
Jewel et al. 2001
37

38. • O2 dissolves less in higher
temperatures
• Work smooth and fast do not
bubble your media
• Device for deoxygenating and
storing prepped media
(HypoxyCOOL)
DE-GASSING/DE-OXYGENATING MEDIA
38

39. • Accelerated oxygen conditioning for tissue culture media
Jewel et al. 2001
HypoxyCOOL
Jewel et al. 2001
39

40. • Progenitor cell trafficking is regulated by hypoxic
gradients through HIF-1 induction of SDF-1
• Ceradini et al. 2014 Nature Medicine
OXYGEN (HIF-1α) RELATED
SCIENTIFIC PUBLICATIONS
• Reactive Oxygen Species Impair the Function of
CD901 Hematopoietic Progenitors Generated
from Human Pluripotent Stem Cells
• Rönn et al. 2016 Stem Cells
40

41. • Defining physiological normoxia for improved
translation of cell physiology to animal models
and humans
• Keeley et al. 2019 Physiol Rev
OXYGEN RELATED SCIENTIFIC PUBLICATIONS
• Physioxic human cell culture improves viability,
metabolism, and mitochondrial morphology while
reducing DNA damage
• Timpano et al. 2019 FASEB
41