Abstract
Objective
To evaluate the effect of ambient room temperature on equipment typically used in in vitro fertilization (IVF).
Design
We set the control temperature of the room to 20 °C (+/−0.3) and used CIMScan probes to record temperatures of the following equipment: six microscope heating stages, four incubators, five slide warmers and three heating blocks. We then increased the room temperature to 26 °C (+/−0.3) or decreased it to 17 °C (+/−0.3) and monitored the same equipment again. We wanted to determine what role, if any, changing room temperature has on equipment temperature fluctuation.
Results
There was a direct relationship between room temperature and equipment temperature stability. When room temperature increased or decreased, equipment temperature reacted in a corresponding manner. Statistical differences between equipment were found when the room temperature changed. What is also noteworthy is that temperature of equipment responded within 5 min to a change in room temperature.
Conclusions
Clearly, it is necessary to be aware of the affect of room temperature on equipment when performing assisted reproductive procedures. Room and equipment temperatures should be monitored faithfully and adjusted as frequently as needed, so that consistent culture conditions can be maintained. If more stringent temperature control can be achieved, human assisted reproduction success rates may improve.
Keywords: Assisted reproductive technology, ART, Room temperature, Equipment stability
Introduction
Temperature is an essential component of cellular physiology and a critical aspect of embryo culture [1]. It is well documented in the literature that temperature stability is necessary during in vitro fertilization (IVF) [2, 3]. In many species such as rabbit [4], bovine [5, 6], mouse [3, 7], rat [8, 9] and human [2, 3, 10–12] in vitro exposure to anomalous temperatures affects gamete competency, embryo development, and offspring survival.
Because temperature can influence in vivo and in vitro reproduction, temperature control is vital in assisted reproduction to ensure that no gametes or embryos are exposed to extreme temperature fluctuations. Stringent temperature control is essential not only during embryo culture, but also during manipulation of gametes and embryos [2, 3]. Incubators, microscope heating stages, slide warmers, and heating blocks must all be calibrated and monitored regularly during use [2, 3] to avoid unnecessary exposure of gametes and embryos to detrimental conditions.
While IVF laboratories are more capable of strict control over equipment temperature, room temperature is not as manageable. Most laboratories’ room temperatures fluctuate between 20 and 25 °C [13]. Room temperature and ambient temperatures are directly affected by the heating and cooling system. Due to air flow patterns, hot and cold spots occur in the laboratory [13]. These hot and cold spots, therefore, affect equipment temperature as these airflow patterns will change during heating and cooling cycles. While many scientists believe surrounding air affects equipment temperature [14–18], the literature is lacking to support this theory.
The aim of this study is twofold. First, we demonstrate the effect that room temperature has on laboratory equipment, and second, we reveal how quickly a change in room temperature can effect laboratory equipment. This narrowly focused study pertaining to a specific aspect of ART labs will prompt other labs to access their own laboratory environments.
Materials and methods
Background
This was an observational study that was conducted in the assisted reproduction laboratory at Greenville Memorial Hospital, located in Greenville, South Carolina from July 5 to July 26, 2011. Because patient data were not utilized, no Internal Review Board was required.
Mechanism of action for CIMScan
We used CIMScan (CIMTechniques, Inc., Beaufort SC) technology to monitor temperature changes in equipment. CIMScan worked via wireless temperature probes attached to each piece of monitored equipment. The probes sent measurements to a monitoring station. These data were sent to a server where it was stored and compared against user-assigned alarm limits. If an alarm was detected, users were immediately notified of the condition. Data were also delivered to the user program where values were displayed.
Validation of CIMScan
To ensure accuracy, CIMScan was validated with the use of two manufacturer calibrated NIST Greisinger GMH 3230 digital thermometers (NIST; Greisinger Electronics, Germany). Before the study began, temperature was recorded for each piece of equipment. The two thermometers measured room temperature and a randomly selected piece of equipment to ensure that both thermometers agreed within +/−0.1 °C. The average difference between the two thermometers at room temperature was 0.03 °C. The average difference between the two thermometers when measuring equipment was 0.04 °C. The p-value associated with room temperature was P = 0.15 and the p-value associated with equipment was P = 0.13. From this evaluation it was agreed that the two thermometers were reading similarly and could be used to validate CIMScan.
So as not to interfere with equipment use, CIMScan probes were attached towards the edges of work surfaces of the equipment. Thirty measurements were taken for each piece of equipment at a specific point on its work surface using the NIST thermometers. Once the values were averaged, they were compared to the values reported by CIMScan. A technician re-calibrated the CIMScan probes to match the average temperature of the work surface area of each piece of equipment. After adjustments were made, another 30 temperatures were taken with the NIST thermometers and these were averaged (data not shown), to verify that CIMScan probes and NIST thermometers all agreed within +/−1.0 °C.
Because heating elements operate in a sine wave fashion [3], it was decided that +/−1.0 °C was the closest that could be achieved at any particular point in time. This value was selected because the NIST thermometers may be recording the temperature at any point along the sine wave and may not represent an average temperature over a 5 min interval as did CIMScan.
As a result of the comparison between the NIST thermometers and CIMScan, we decided agreement was comparable and the rest of the measurements were made using CIMScan only. Equipment included in the study is listed in Table 1.
Table 1.
Manufacturer/model number of equipment used in an in vitro fertilization laboratory
| Equipment | Manufacturer/model number |
|---|---|
| Heating block 3 | Labline Multiblock/2052 |
| Heating block 5 | Barnstead International Modular Block/2003 |
| Slide warmer 1 | Precision Scientific Inc./66632 |
| Slide warmer 2 | Labline/26020 |
| Slide warmer 3 | Labline/26005 |
| Slide warmer 4 | Labline/26020 |
| Slide warmer 5 | Labline/26020 |
| Dry incubator 2 | Boekel Scientific/132000 |
| Incubator 1 | Thermo Electron Corp, Forma series II/3140 |
| Incubator 11 | Thermo Electron Corp, Forma series II/3110 |
| Incubator 12 | Thermo Electron Corp, Forma series II/3110 |
| Scope 1 | Nikon/A0 stereomicroscope |
| Scope 2 | Nikon/SMZ 1550 |
| Scope 3 | Nikon/Diaphot-MD |
| Scope 4 | Nikon/Diaphot-MD |
| Scope 5 | Nikon/SMZ 1550 |
| Scope 6 | Eclipse/TES TE2000-S |
Experiment I: effect of room temperature changes on equipment temperatures
The control temperature for the room was set at 20 °C (+/−0.3 °C). The room temperature was increased to 26 °C (+/−0.3 °C) or decreased to 17 °C (+/−0.3 °C) and equipment temperature was recorded. The room temperature ran for three consecutive days at each temperature. Shields were placed over air vents to divert direct airflow away from equipment. When the room temperature was stable, the temperature of each piece of equipment was recorded every 5 min over a 12 h period for a total of 129 measurements. The temperature variance (for each individual piece of equipment) and the mean temperature variance (comparison between the different types of equipment) were determined at the three different room temperatures (17 °C vs. 20 °C and 20 °C vs. 26 °C).
Experiment II: effect of sudden room temperature changes on equipment temperatures
To determine how quickly equipment can respond to room temperature, a sudden temperature increase that occurred during the investigatory period was accessed to observe the effect that a rapid increase or decrease in room temperature had on equipment.
Experiment III: comparison of the temperature stability of analog and digital microscope stage warmers
A comparison of analog and digital microscope stage warmers was performed to determine if one type was more temperature stable than the other. Three hundred and eighty seven temperature measurements for three analog and 387 temperature measurements for three digital stage warmers were taken at room temperatures of 17 °C and 26 °C. The results for each type of stage warmer at each room temperature were averaged and compared.
Statistical analysis
We attempted to take sufficient temperatures for each piece of equipment with the hopes of reducing variability and bias. There were no changes to the study outcomes once the trial commenced. No data were lost. All statistical analyses of the data were performed using a Student’s paired t test. Statistical significance was defined as P < 0.05.
Results
Experiment I: effect of gradual room temperature changes on equipment temperatures
20 °C versus 26 °C
Equipment temperature was compared at two different room temperatures to investigate the affect of room temperature on equipment stability (Table 2). Measurements from identical models of microscope stages and slide warmers were pooled for analysis. When comparing equipment temperature at 20 °C and 26 °C, there were statistically significant temperature differences between all microscope stages, slide warmers, heating blocks and two of the three incubators, regardless of model. The only piece of equipment that maintained the set temperature was a non-humidified Forma incubator.
Table 2.
Effect of temperature on equipment at 20 °C vs. 26 °C. One hundred and twenty-nine measurements were taken for each piece of equipment. Measurements from identical pieces of equipment were pooled for analysis
| Equipment | 20 °C | 26 °C | Paired difference | P value |
|---|---|---|---|---|
| Scope 1 | 34.30 ± 0.02 | 36.16 ± 0.07 | 1.86 ± 0.07 | <0.0001 |
| Scope 6 | 35.00 ± 0.07 | 37.05 ± 0.16 | 2.05 ± 0.17 | <0.0001 |
| Scope 3, 4 | 35.15 ± 1.13 | 36.90 ± 0.21 | 1.75 ± 1.32 | <0.0001 |
| Scope 2, 5 | 33.08 ± 0.79 | 35.02 ± 0.36 | 1.94 ± 0.46 | <0.0001 |
| Slide warmer 1 | 34.38 ± 0.16 | 36.38 ± 0.08 | 3.39 ± 0.20 | <0.0001 |
| Slide warmer 3 | 32.74 ± 0.36 | 35.89 ± 0.47 | 3.14 ± 0.55 | <0.0001 |
| Slide warmer 2, 4, 5 | 32.82 ± 0.29 | 35.76 ± 0.39 | 2.94 ± 0.48 | <0.0001 |
| Heating block 3 | 34.70 ± 0.93 | 35.19 ± 0.90 | 0.49 ± 1.21 | <0.0001 |
| Heating block 5 | 37.12 ± 0.08 | 37.67 ± 0.05 | 0.55 ± 0.09 | <0.0001 |
| Dry incubator 2 | 36.79 ± 0.17 | 37.7 ± 0.16 | 0.88 ± 0.27 | <0.0001 |
| Incubator 1 | 37.0 ± 0.08 | 37.0 ± 0 | 0.008 ± .09 | 0.32 |
| Incubator 11 | 36.60 ± 0.02 | 37.0 ± 0.009 | 0.10 ± 0.009 | <0.0001 |
| Incubator 12 | 36.4 ± 0 | 36.6 ± 0 | 0.20 ± 0.0 | <0.0001 |
17 °C versus 20 °C
There was a direct relationship between room temperature and equipment temperature when comparing equipment temperatures at 17 °C and 20 °C. Again, for all microscope stages, slide warmers, heating blocks and incubators, a statistically significant temperature difference was observed at a room temperature of 17 °C versus 20 °C (Table 3).
Table 3.
Effect of temperature on equipment at 17 °C vs. 20 °C. One hundred and twenty-nine measurements were taken for each piece of equipment. Measurements from identical pieces of equipment were pooled for analysis
| Equipment | 17 °C | 20 °C | Paired difference | P value |
|---|---|---|---|---|
| Scope 1 | 33.14 ± 0.08 | 34.3 ± 0.02 | 1.15 ± 0.08 | <0.0001 |
| Scope 6 | 33.69 ± 0.09 | 35.0 ± 0.07 | 1.30 ± 0.11 | <0.0001 |
| Scope 3, 4 | 32.60 ± 0.43 | 35.10 ± 1.13 | 2.50 ± 0.77 | <0.0001 |
| Scope 2, 5 | 30.05 ± 0.09 | 33.08 ± 0.79 | 2.50 ± 0.73 | <0.0001 |
| Slide warmer 1 | 28.01 ± 0.17 | 30.45 ± 0.08 | 2.44 ± 0.18 | <0.0001 |
| Slide warmer 3 | 30.54 ± 0.42 | 32.74 ± 0.36 | 2.20 ± 0.53 | <0.0001 |
| Slide warmer 2, 4, 5 | 30.32 ± 0.25 | 32.82 ± 0.29 | 2.50 ± 0.39 | <0.0001 |
| Heating block 3 | 34.15 ± 0.87 | 34.70 ± 0.93 | 0.55 ± 1.32 | <0.0001 |
| Heating block 5 | 36.71 ± 0.03 | 37.12 ± 0.03 | 0.41 ± 0.05 | <0.0001 |
| Dry incubator 2 | 36.27 ± 0.12 | 36.78 ± 0.12 | 0.50 ± 0.18 | <0.0001 |
| Incubator 1 | 36.90 ± 0.01 | 37.00 ± 0 | 0.10 ± 0.01 | <0.0001 |
| Incubator 11 | 36.50 ± 0 | 36.60 ± 0 | 0.10 ± 0 | <0.0001 |
| Incubator 12 | 36.40 ± 0 | 36.30 ± 0 | 0.10 ± 0 | <0.0001 |
Experiment II: effect of sudden room temperature changes on equipment temperatures
Equipment temperature responded within 5 min to a sudden change in room temperature (Fig. 1). Data demonstrated a direct relationship between a sudden upward spike in room temperature and an increase in equipment temperature.
Fig. 1.
Direct effect of a sudden upward spike in room temperature on equipment. Room temperature is shown in red and microscope stage temperature is shown in blue
Experiment III: comparison of the temperature stability of analog and digital microscope stage warmers
A comparison between digital and analog equipment was performed. Three analog and three digital microscope stage warmers were compared at 17 °C and 26 °C. Analog microscope stage warmer temperatures ranged from 30.3 °C to 33.4 °C at 17 °C and 36.1 °C to 37.4 °C at 26 °C. Digital microscope stage warmer temperatures ranged from 30.3 °C to 33.9 °C at 17 °C and 34.2 °C to 37.3 °C at 26 °C. Both analog and digital microscope stage warmers were similar in their ability to maintain temperature at 17 °C (P = 0.35). A statistical difference in their ability to maintain set temperature was observed at 26 °C (P < 0.0001).
Discussion
To the authors’ knowledge, this is the first time that a study clearly demonstrates a direct relationship between room temperature and laboratory equipment temperature. As room temperature increased, the temperature of heating blocks, microscope stage warmers, slide warmers and incubators increased regardless of make or model (P < 0.0001). The same equipment decreased in temperature as room temperature decreased (P < 0.0001). Microscope stage warmers and slide warmers had the greatest temperature fluctuation compared to heating blocks and incubators. Heating blocks and incubators did have some temperature fluctuation; however, these slight temperature fluctuations most likely do not affect gametes and embryos. This study also demonstrates that equipment temperature reacts rapidly to sudden changes in room temperature. If room temperature spikes, equipment temperature will show the same increase within 5 min.
Since digital heating elements are a more recent technology, there may be a bias that digital equipment maintains temperature more consistently than analog equipment; however, this was not demonstrated in this study. When comparing digital and analog microscope stage warmers, there was no statistical difference between the two at 17 °C. However, when the room temperature increased to 26 °C, there was a significant temperature difference between the digital and analog stage warmers. Analog stage warmers were able to hold closer to the set point at both temperatures.
Though there is limited clinical correlation in this study, there is an implication that changes in lab equipment temperature may have an effect on gamete and embryo development. Equipment tested in our laboratory is certainly not an exhaustive list of equipment used to maintain temperature of gametes and embryos during ART procedures. Other examples include different types of test tube warmers used during retrievals [14], IVF workstations [18], and heated microscope objectives [11, 15]. Each laboratory must evaluate their own equipment and optimize temperature control.
For ethical reasons, it is not possible to assess the effect of changes in room temperature on human embryos and gametes. However, with the use of animal models, future studies include assessing changes in embryos and gametes as room temperature changes.
The concept of room temperature fluctuation and its effect on equipment temperature needs to be taken into consideration when constructing an ART laboratory. Such a facility should not be placed so as to have the walls be part of the exterior of a building. A facility should not reside on the top floor of a building where outside temperatures that change with the seasons of the year have the potential of altering the inside room temperatures. The use of windows in the walls of an ART laboratory should be assessed carefully due to the transfer of heat and cold through glass.
In a field where temperature is critical, it is troublesome to think that our equipment is quite unstable. During the performance of assisted reproduction procedures, there are many instances when gametes and embryos can be exposed to temperature fluctuations (i.e., oocyte retrieval and embryo transfer, incubator door openings, changes in location from incubator to microscope stage). These fluctuations are magnified if the room temperature is not constant. Perhaps room temperature is one of the culprits in the inefficiency of human assisted reproduction.
Conclusion
Clearly, it is necessary to be aware of the affect of room temperature on equipment when performing assisted reproduction procedures. Room and equipment temperatures should be monitored faithfully and adjusted as frequently as needed, so that consistent culture conditions can be maintained. If more stringent temperature control can be achieved, human assisted reproduction success rates may improve.
Acknowledgments
The authors' would like to acknowledge H. Lee Higdon III, Ph.D. for his statistical assistance.
Conflicts of interest
None of the authors have a conflict of interest.
All funding for this research came from within the department.
Footnotes
This research was submitted to the Faculty of Eastern Virginia Medical School in partial fulfillment of the requirement for the degree of Master of Science in Biomedical Sciences-Clinical Embryology and Andrology. Norfolk, Virginia May 2012.
Capsule Because modest changes in ambient air can affect surrounding equipment, it is necessary to be aware of room temperature when performing assisted reproductive procedures.
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