the role of ambient air in preimplantation toxicology and its impact on clinical outcomes in ivf,...
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![Page 1: The Role of Ambient Air in Preimplantation Toxicology and its Impact on Clinical Outcomes in IVF, Micahel Magee & Kathryn Worrilow PhD](https://reader031.vdocuments.mx/reader031/viewer/2022011722/58701b1d1a28ab7f428b68a5/html5/thumbnails/1.jpg)
The Role of Ambient Air in Preimplantation Toxicology and its
Impact on Clinical Outcomes in In Vitro Fertilization (IVF)
Southwest Embryology Summit
January 4, 2016
Michael P. Magee Kathryn C. Worrilow, Ph.D.
LifeAire Systems
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Our Objectives Today • Discuss the specific role of ambient air
quality in successful preimplantation toxicology and embryogenesis.
• Discuss the common sources of airborne threats to the laboratory environment - there are as many within the IVF laboratory as there are in the outside source air serving your laboratory and clinical space - which airborne pathogens are critical and which are less important to the process.
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Our Objectives Today • Discuss 3-3-3-4! • Evaluate current mechanisms of
remediation and their effectiveness • Recognize what can you do - what
questions can you ask – what solutions exist to assure that your lab design, HVAC system, upstream equipment and protocols are supporting an optimal and consistent laboratory environment?
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3 – 3 – 3 – 4 • 3 categories of airborne pathogens
• 3 sources of air • 3 categories of filtration physics
• 4 THM – which led to the identification of a problem and
ultimately a solution
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v Outside air serving the HVAC system
v Recirculated air within the space to be protected
v Air provided by the HVAC system
Three Sources of Air
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Outside Influences Outside of Your Control
• Road resurfacing • Rooftop resurfacing
• Construction • Idling engines, exhaust
• Waste management, restaurant, generator exhaust direction
• Accidents, tire fires • Seasonal pollutants
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The IVF Laboratory: Common Constituents of Recirculated Air
v Tissue cultureware § Styrene § Toluene § Acetone § 2-butanone
v Isopropanol, acetone v Equipment, material off-gassing v HVAC / refrigerants / compressed gasses v CO2, LN2, tri-gas tanks v Personnel bioburden
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HVAC System-Specific Organisms
v Pathogens – viruses, bacteria, fungi v Allergens – bacteria, mold v Toxins – endotoxins, mycotoxins
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Operation within an ISO 5 cleanroom - thus removing the variable of air –
• ISO 5 design incorporating optimal air flow and air dynamics
• Dedicated AHU and sealed plenum • 30 ACH, + pressure, live monitoring
• Laminar flow • > 300 lbs of carbon, > 150 lbs of KMnO4 (Cohen,
Gilligan and Hall) • ULPA/UVC final filtration (Boone and Higdon)
• Air flow over critical points of process • Cleanroom SOPs followed
• Quarterly certification
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How effective was the IVF laboratory cleanroom and HVAC air filtration system
as designed?
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+ be
ta/C
linic
al P
regn
ancy
Rat
e (%
+fhb
)
Testing Quarters (TQ) Worrilow et al, 2002, 2008 Worrilow, 2013, 2015
52.4% 54.5%
16.0%
50.0%
37.5%
33.3%
41.6%
13.3%
29.4%
64.7%
55.5%
52.6%
0%
10%
20%
30%
40%
50%
60%
70%
TQ1 TQ2 TQ3 TQ4 TQ5 TQ6 TQ7 TQ8 TQ9 TQ10 TQ11 TQ12
Analysis of CPR, TVOC and Biological Loading within the IVF Laboratory
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What did the study of preimplantation toxicology, human embryogenesis, ambient air quality and clinical outcomes tell us? The
study clearly delineated and defined the problem of the variable
of ambient air and the optimal culture environment, and the
specific role that they played in our processes…..it also led to a data
driven solution….
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THM #1: IVF does not require the traditional cleanroom environment.
The traditional cleanroom focuses on nonviable particulates, NOT the level of VOCs and viable particulates that must be maintained to optimize the in vitro culture environment for the human
embryo
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THM #2: The impact of subtle levels of VOCs on our process and clinical
outcomes
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Analysis of CPR & TVOC Loading within the IVF Laboratory
Clin
ical
Pre
gnan
cy R
ate
(%)
Testing Quarters (TQ)
2.7 2.7 2.2 2.2
16.0% 16.0%
37.5% 37.5% 33.3% 33.3% 41.6% 41.6%
13.3% 13.3%
29.4% 29.4% 41.6%
2.7 2.2
16.0%
37.5% 33.3% 41.6%
13.3%
29.4%
0.0
0.5
1.0
1.5
2.0
2.5
0.0
0.5
1.0
1.5
2.0
2.5
TVOC
Levels (Parts Per B
illion)
51.6%
TQ12 0 10
20
30 40 50 60 70 80 90
100
TQ11 TQ10 TQ9 TQ8 TQ7 TQ6 TQ5 TQ4 TQ3 TQ2 TQ1
54.5% 50.0% 52.4%
0.1 0.1 0.1 0.1 0.1
64.7% 55.5%
The presence of ppb VOCs severely compromised clinical outcomes
Worrilow et al, 2002, 2008 Worrilow, 2013, 2015
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FAQ: Can’t we simply add more carbon and
KMnO4 to fix the problem?
No.
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VOCs Common to the IVF Laboratory
§ Polar VOCs § Nonpolar VOCs
§ Low MW hydrocarbons § High MW hydrocarbons
§ Fungal VOCs § Microbial VOCs § Biogenetic VOCs
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Filtration Physics: Mechanisms of VOC Remediation
• Physical adsorption • Chemisorption
• Oxidation • Persulfate oxidation
• Thermal oxidation • Photo-Fenton oxidation
• Ultraviolet photocatalytic oxidation (UVPCO) • Ultraviolet UVV wavelength
• Molecular sieve • Transition metal impregnation
• Fixed bed adsorption • Surface, contact condensation
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“I culture under oil, use tabletop incubators,
time-lapse imaging, etc. VOCs cannot enter my
media or affect my embryos.”
We wish this were true.
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If oil soluble, VOCs can easily enter the oil overlay and if water
soluble, the media. Once present in the media or oil, the VOCs are a permanent resident in the culture environment, and thus a permanent threat to the
embryo.
Partition Coefficients
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THM #3: The impact of subtle levels of viable
particulates/biologicals on our process and clinical
outcomes
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The Impact of Subtle Levels of Airborne Biological Pathogens on
Clinical Pregnancy Rates +
beta
/Clin
ical
Pre
gnan
cy R
ate
(% +
fhb)
(n=54 testing quarters (TQ)) 0
10
20
30
40 50 60 70 80 90
+ beta
53.3% 45.6% 47.1%
17.6%
+ beta CPR
CPR
Loss of power to the ballast boxes supporting the UV lights in our HVAC system coincided with an increase in our clinical loss or
miscarriage rate
Increased biologicals within laboratory air
Worrilow et al, 2002, 2008 Worrilow, 2013, 2015
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Filtration Physics: UV v Critical to successful human
embryogenesis and clinical outcome is the intensity, lamp coordinates, level and longevity of the UV output.
v Selection of the right UV source and assurance of maintenance SOP is paramount.
v Not all UV are equal!
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THM #4: What HEPA filtration does and does NOT do for our culture
environment
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We create the perfect environment for growth…..
IVF laboratory room temperature, humidity and
HEPA/ULPA filter substrate = proliferation of bacterial and viral spores,
mold and biologicals
HEPA/ULPA Final Filter Above IVF Laboratory
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Filtration Physics: Viable Particulates UV, HEPA and ULPA
v HEPA and ULPA filtration are designed to remove or capture particles greater than 0.3 microns in diameter at a 99.97 – 99.99, 99.997 – 99.999% effectiveness rating, respectively.
v HEPA and ULPA filtration remediate by “capture.”
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Confidential Confidential
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v If air bypass or filter leakage are present and/or if inadequate maintenance and change-outs, the capture rate can drop to 99.94 – 99.95%.
v Such decrease in efficiency can allow, in the course of a single day, 7.2M particles to penetrate the HEPA filter and contaminate the IVF laboratory and clinical procedure rooms.
v Effective log kill of viable particulates, bacterial, fungal and viral spores demands the use of the proper UV wavelength, intensity, lamp coordinates, level and longevity of the output.
Filtration Physics: Viable Particulates UV, HEPA and ULPA
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+ be
ta/C
linic
al P
regn
ancy
Rat
e (%
+fhb
)
Testing Quarters (TQ) Worrilow et al, 2002, 2008 Worrilow, 2013, 2015
52.4% 54.5%
16.0%
50.0%
37.5%
33.3%
41.6%
13.3%
29.4%
64.7%
55.5%
52.6%
0%
10%
20%
30%
40%
50%
60%
70%
TQ1 TQ2 TQ3 TQ4 TQ5 TQ6 TQ7 TQ8 TQ9 TQ10 TQ11 TQ12
Data Collected Over the 10 Year Study Led to the Identification of the Problem
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Identification of the Problem Led to a Comprehensive
Understanding ~ Understanding the source of and impact of subtle levels of VOCs and
airborne biologicals on our processes and defining the optimal in vitro culture environment led to a comprehensive
understanding of the critical nature of the environment that we provide ~
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The absence of a solution led to the genesis of the
LifeAire Systems technology.
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v System designed to comprehensively KILL and remove all
airborne pathogens: chemical, biological and particulate
ON A SINGLE PASS and produce NO byproducts.
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✔
✔
✔
✔ ✔✔
✔
✔
✔
✔✔
✔✔✔ ✔
✔
✔✔
✔
✔
✔
• UCSF Medical Center • Stanford University Medical Center
• Northwestern University Medical Center • University of Connecticut Health Center
• University of Iowa Hospital
✔
✔
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Variation in Protocols ü Controlled O2
ü Tabletop incubators ü Upright CO2 incubators ü Continuous culture ü Sequential culture
ü ICSI, PICSI, PGD, PGS ü Day 5/6 vitrification
ü IVF clinical and laboratory staff
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ü Control of ambient air
Shared Variable
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Third Party Validation: Alpha Environmental
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Fertility and Sterility, Vol. 102; Issue 3 Suppl, p. E91.
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Fertility and Sterility, Vol. 102; Issue 3 Suppl, p. E91.
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In addition to air purification…..know what you have in your HVAC
system, pop the ceiling tiles and ask questions….lots of
questions……
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Critical Components: HVAC System
• Location of air intake • Dedicated air handling unit (AHU) • Water and gas pipelines • Upstream filters • Chiller • Dedicated humidity system • Proper dehumidification, cooling and
heating equipment • Comprehensive air purification system
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Quality Control Initiatives Towards the Optimal Environment
• Air quality component, testing and management
• Air quality over critical points of process, energy efficiency
• Air testing, comprehensive TAB • Materials selected for laboratory and clinical
space • Operational considerations to protect
environment • Clinical and laboratory staff and work flow
towards the optimal environment
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When HVAC Solutions Are Proposed: Ask for Data! Proof of Technology! • Do they understand the environment that you
are protecting, the vulnerability of the human embryo?
• Can they assure consistency and performance of your environment?
• Do they understand the requirements needed by your upstream HVAC equipment?
• Is the upstream HVAC equipment meeting your criteria?
• How will maintenance and SOP protocols be followed to assure consistency?
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In Summary v The air serving our in vitro culture environment
is dynamic in nature and is influenced by the outside source air, materials used in the laboratory, laboratory protocols, staff, the HVAC design relative to your critical points of process, and all associated HVAC upstream equipment.
v Subtle levels of airborne VOC and biological contaminants can be impactful to successful human embryogenesis and clinical outcomes – proper mechanisms of filtration physics must be used to comprehensively remediate, remove and control the airborne pathogens.
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v Using ultraviolet genomic modeling, mathematical modeling and a proprietary engineered molecular media, the LifeAire System comprehensively captures and inactivates all chemical and biological contaminants.
v It is critical to control the quality of the ambient air serving your in vitro culture environment to optimize successful preimplantation embryogenesis and provide improved levels of patient care.
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Success is in the air.