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Cell Biology International 33 (2009) 755e757 www.elsevier.com/locate/cellbi
Background ELF magnetic fields in incubators: A factor of importance in cell culture work Kjell Hansson Mild a,*, Jonna Wile´n a,b, Mats-Olof Mattsson c,d, Myrtill Simko d,e a
Department of Radiation Sciences, Umea˚ University, SE-901 87 Umea˚, Sweden b University Hospital of Northern Sweden, Umea˚, Sweden c Department of Natural Sciences, O¨rebro university, Sweden d Department of Cell Biology and Biosystems Technology, University of Rostock, Germany e Institute of Technology Assessment, Austrian Academy of Sciences, Vienna, Austria Received 10 November 2008; revised 4 March 2009; accepted 14 April 2009
Abstract Extremely low frequency (ELF) magnetic fields in cell culture incubators have been measured. Values of the order of tens of mT were found which is in sharp contrast to the values found in our normal environment (0.05e0.1 mT). There are numerous examples of biological effects found after exposure to MF at these levels, such as changes in gene expression, blocked cell differentiation, inhibition of the effect of tamoxifen, effects on chick embryo development, etc. We therefore recommend that people working with cell culture incubators check for the background magnetic field and take this into account in performing their experiments, since this could be an unrecognised factor of importance contributing to the variability in the results from work with cell cultures. Ó 2009 International Federation for Cell Biology. Published by Elsevier Ltd. All rights reserved. Keywords: Magnetic flux density; Heating system; Electronic control; Low intensity; Biological effect; Cell culture
1. Introduction All experimental work, including work where cell culture is involved, needs to have environmental conditions monitored and meticulously controlled to enhance the probability that the result can be reproduced by others. Every so often, especially subtle biological responses can be difficult to replicate, even within the same laboratory. It is well known that factors such as the biological material (specific cell types, origin of cell lines and sub-lines, number of passages in culture, etc.), cell culture media and additives (sera, antibiotics, growth factors, etc.), as well as the plastic components that are used, may contribute to variations in experimental outcomes. Also physical factors such as temperature and CO2 variations, as well as mechanical vibration, can contribute to experimental Abbreviations: MF, magnetic field; mT microtesla, 106 T. * Corresponding author. Tel.: þ46 90 7851712. E-mail address:
[email protected] (K.H. Mild).
artefacts. Here we point to an additional factor to consider, namely background MFs in CO2 cell culture incubators. In commercial cell incubators, there is always a background ELF magnetic field present emanating from the wiring of the heating systems, electronic control equipments, fan engines, etc. These MFs can reach rather high levels if special attention has not been paid to this in the construction of the incubators. Occasionally we have seen levels up to tens of mT, although values are typically a few mT. As a comparison, the background flux density in a normal domestic or office environment is around 0.05e0.1 mT. There is also a spatial gradient in the incubator e usually from top to bottom e depending on where the electronics are located and how the heating wires are drawn. Here we report on measurements of fields in a few sample incubators. The reason why we find it important to consider the background MF in ordinary cell culture work is that research studies on low level effects of electromagnetic fields have reported effects at levels equal to or even below those that can
1065-6995/$ - see front matter Ó 2009 International Federation for Cell Biology. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.cellbi.2009.04.004
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K.H. Mild et al. / Cell Biology International 33 (2009) 755e757
be found in incubators. Biological effects, like changes in gene expression, blocked differentiation, effects on chick embryo development, etc, after such exposure have been reported at field levels from a few tenths to tens of mT. In view of the demonstration that the low intensity MF can cause biological responses, it is essential that cell culture experiments are done under carefully controlled conditions and it is necessary that the cells should be kept at as low background MF level as possible.
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Magnetic flux density (µT)
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Magnetic flux density (µT)
Table 1 Measured magnetic flux density (mT) inside incubators.
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Although we have long-term research experience of performing EMF research, including the need to control for background ELF MF, we were surprised when one fan motor of an incubator broke and was replaced. The new fan motor generated fields in excess of 80 mT in its vicinity. This was irrespective of the fact that the new one was an original spare part of a seemingly identical make and model. It is necessary to measure resident MF in the incubator on all shelves. If the values are too high one may need to rewire the heating system or relocate the electronic control equipment (which are possible adjustments to perform). It is also necessary to control other environmental MF sources in the lab. A typical instance is placing an apparatus on top of the incubator. The device may have a transformer inside giving a high leakage field into your incubator. Another typical situation is the location of a laminar flow hood close to the incubator. A number of biological effects in cellular systems have been shown at low levels of MF. For instance, Tokalov and Gutzeit (2004) exposed human myeloid leukemia (HL-60) cells to different flux densities in the range 10e140 mT. Even the lowest level tested (10 mT) resulted in a significant induction of the
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The results of the spot measurements are shown in Table 1. It can be seen that the magnetic field can be quite high in some of the incubators; one value reached 38 mT in one specific spot. In Figs. 1 and 2 the logged values during night time and during one heating cycle in daytime are shown. The heating cycles are of different length during night or daytime, with typical values of 30e40 min during night time and 60 min during daytime. This difference is due to the more frequent opening of the incubator during daytime as compared to night time. Spot MF measurements taken at the ‘‘wrong’’ time (heat cycle off) can give a rather large erroneous value for the flux density.
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4. Discussion
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3. Results
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Fig. 1. The broad band magnetic flux density (mT) in Innova CO70 as a function of time during night. The broad band mean value was 9.3 mT.
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Spot measurements of CO2-incubators of different brands and models, belonging to scientist colleagues, have been made. Measurements were obtained using a handheld instrument (a triaxial ELF microteslameter, MFD-V, SMPS Electronics) predominately in the 20e1000 Hz range. In addition, long-term measurements were performed in a few cases since the heat-regulation of incubators is thermostat controlled, and thus have oneoff cycles of variable lengths. These values may differ considerably from the spot MF measurements. A measurement of background MF in an incubator thus needs to be extended over an appropriately longer time period. We have used an EMDEX II (Enertech Consultants, CA, USA) with a frequency range of 20e800 Hz to log the magnetic field over time in one incubator.
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2. Material and method
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Fig. 2. The broad band magnetic flux density (mT) in Innova Co70 as a function of time for one heating cycle during daytime. The broad band mean value during one heating cycle was 9.5 mT.
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K.H. Mild et al. / Cell Biology International 33 (2009) 755e757
genes HSP70A, HSP70B, and HSP70C. Chen et al. (2000) showed that low frequency magnetic field exposure could partially block the differentiation of Friend erythroleukemia cells, similar to the results obtained when these cells are treated with chemical-tumor promoters. The cells were exposed to 60 Hz MF which resulted in a dose dependent inhibition of differentiation, with maximal inhibition peaking at 40% and 4 mT. Already at 2.5 mT the differentiation was inhibited 20%. Girgert et al. (2005) showed that exposure of a breast cancer cell line, MCF-7, to a 50 Hz MF could inhibit the growth inhibitory effect of tamoxifen in a flux density dependent manner. A maximal response was seen at 1.2 mT. Similar effects by 1.2 mT MF on tamoxifen growth inhibition of MCF-7 cells have also been published by others (Harland and Liburdy, 1997; Blackman et al., 2001; Ishido et al., 2001). Girgert et al. (2008) point out that they have taken the stray magnetic field in incubators into account. They found that the intensity of the MF present in the observed commercial cell culture incubators could be as high as 1.5 mT at the upper level of the incubators, generated by electric control facilities for heating, pumps, and valves inside the incubators. Therefore they performed their exposure experiments in specially designed incubators with negligible stray fields. Berman et al. (1990) reported effects on chick embryo development from a 1 mT pulsed field. Mullins et al. (1999) exposed L929 cells for 4 h to a 60 Hz MF at different flux densities from 0 to 20 mT. The ODC activity ratio exposed to sham is seen to increase with a sigmoidal function where 5 mT causing 50% increase and maximal response (doubling) at 7 mT. The incubators investigated in the present study were all energized with 230 V. We can assume that incubators operating on 110 V would have a rather higher magnetic field since those currently used would then be approximately doubled. This effect of the voltage is for instance seen in the Henhouse project (Berman et al., 1990) where 6 laboratories performed chick embryo experiment with identical incubators. It turned out that the European labs operating at 230 V had
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a background field of 0.5 mT whereas those from the US reported 1 mT. The stray magnetic fields generated by incubators and other laboratory equipments may thus be one factor of importance for the variability in the results of different cell culture work. If the cells sometimes are placed on the upper shelf and another time on the lower, the difference in magnetic field may very well be one of the reasons for the difference in outcome. This attention to detail is normally not exercised in cell culture experiments. We therefore urge people working with cell culture to check for the stray fields and take this into account when performing experiments. References Berman E, Chacon L, House D, Koch BA, Koch WE, Leal J, et al. The effects of a pulsed magnetic field on chick embryos. Bioelectromagnetics 1990; 11:69e187. Blackman CF, Benane SG, House DE. The influence of 1.2 microT, 60 Hz magnetic fields on melatonin and tamoxifen induced inhibition of MCF-7 cell growth. Bioelectromagnetics 2001;22:122e8. Chen G, Upham BL, Sun W, Chang CC, Rothwell EJ, Chen KM, et al. Effect of electromagnetic field exposure on chemically induced differentiation of friend erythroleukemia cells. Environ Health Perspect 2000;108:967e72. Girgert R, Schimming H, Korner W, Grundker C, Hanf V. Induction of tamoxifen resistance in breast cancer cells by ELF electromagnetic fields. 4. Biochem Biophys Res Commun 2005;336(4):1144e9. Girgert R, Gru¨ndker C, Emons G, Hanf V. Electromagnetic fields alter the expression of estrogen receptor cofactors in breast cancer cells. Bioelectromagnetics 2008;29(3):169e76. Harland JD, Liburdy RP. Environmental magnetic fields inhibit the antiproliferative action of tamoxifen and melatonin in a human breast cancer cell line. Bioelectromagnetics 1997;18:555e62. Ishido M, Nitta H, Kabuto M. Magnetic fields (MF) of 50 Hz at 1.2 mT as well as 100 mT cause uncoupling of inhibitory pathways of adenylyl cyclase mediated by melatonin 1a receptor in MF-sensitive MCF-7 cells. Carcinogenesis 2001;22(7):1043e8. Mullins JM, Penafiel LM, Juutilainen J, Litovitz TA. Dose-response of electromagnetic field e enhanced ornithine decarboxylase activity. Bioelectrochem Bioenerg 1999;48:193e9. Tokalov SV, Gutzeit HO. Weak electromagnetic fields (50 Hz) elicit a stress response in human cells. Environ Res 2004;94:145e51.
