Setting up a light microscopy core facility: Facility design

The successful operation of a light microscopy core facility depends also on the initial setup of its infrastructure. This article covers the aspects of location selection and room planning and what environmental factors need to be considered. These include light, temperature, vibrations as well as the basic installations needed for microscope operation.


INTRODUCTION
One of the most important aspects in the creation of an efficiently operating light microscopy core facility is the selection of a suitable location and its technical adaptation for operation.The circumstances under which this takes place can vary significantly, ranging from adapting existing rooms in an already operating institute over the planning and building of facility rooms in a new building to the planning and execution of a building specifically for imaging technologies as is the case for the recently opened EMBL Imaging Centre in Heidelberg, Germany.The following article aims to cover fundamental requirements that underly all scenarios as well as highlight additional considerations that are linked to more specific circumstances.While it draws on existing installations and experiences it aims to present the general concepts that go into the design of a core facility instead of specific examples.The aim is to provide an overview of the building considerations.These are also covered in very instructive articles on facility setup and operation 1-3 but are considered together with other equally important aspects of operation.The complexity of the issues involved merits a closer look as attempted here.
The article will cover all aspects that could ideally be considered in the setup of an imaging facility including the considerations for the location of the facility in the building.In many cases, some or or even most of these aspects may not be decidable by the person in charge of the facility.While it remains important to present the process of core facility setup in its entirety, the article will conclude with strategies that can be applied in more limited decision spaces.

INITIAL CONSIDERATIONS
Current state-of-the-art light microscopy systems range from relatively robust and compact widefield systems to multiple laser-based systems for nonlinear microscopy that can fill a whole room.The associated requirements vary accordingly, but while a robust system may not need a fully adapted room, an advanced system cannot operate without it.In planning a facility, the rooms should be intended as future-proof and ideally be built to a standard that supports future advanced operations.The level of planned future requirement should be identified during initial planning and hold for a number of years to avoid costly refurbishments early on in operation.
The choices of location and distribution of a core facility will inevitably (and understandably) be affected by many other considerations of equal importance.It is however important to communicate to management and direction clearly the requirements for such a facility and to push for their maximal consideration in the decision-making process to ensure successful operation in the future.A very important point to make here is that short-sighted economisation during installation will lead to future costs during operation due to operational incidents, instrument downtime and repair, thus leading to loss of revenue and increased operational cost that will accumulate with every year of operation.

SELECTING THE LOCATION
The quality of any form of microscopy data is easily affected by external disruptions and the problem only increases with the level of resolution and sensitivity of the microscopy method.Disruptions acceptable for quick checking of cell confluency on a cell culture microscope are unacceptable for confocal microscopes and much less for superresolution microscopy methods.Accordingly, a microscopy facility should be in a well isolated part of the institute and protected from:

Vibrations
Constant as well as sporadic vibrations will severely affect microscopy.Specific vibration considerations will still be covered in a separate section, but a microscopy facility should be planned in an area that is well supported by the building's load-carrying structures or on a solid foundation.If it is possible to choose a specific floor in the building, levels close to the ground floor, the ground floor itself or basement levels are preferable.While this makes sense at the level of the building itself, such considerations may still be over-ruled by the building's location and environment.If basement and ground levels are exposed to the risk of flooding by extreme weather events they are not good choices for placing highly sensitive equipment.Proximity to some shorelines (scenic as they may be) may also cause disturbances in the form of surfcrash or colliding ice floes.Depending on the quality of the building's static planning and construction, any floor may be suitable for operation, but this should be checked beforehand and be part of any planning.External factors like passing train tracks or subway lines, motorways etc. also need to be considered in the location selection.Another requirement is to avoid proximity to technical installations with disruptive machinery, for example compressors, pumps or elevators.

Exposure and temperature changes
Building areas that are directly exposed to the sun undergo significant changes in temperature every day.Temperature stability in instrument rooms would under such circumstances require a large amount of environmental control and the associated energy cost, which is not needed if windowless, internal-facing rooms are used as instrument rooms and outward facing rooms as office and lab space.As a building will be exposed to different outside temperatures throughout the year, higher amounts of active compensation and more dynamic regulation will be needed in external rooms than in internal rooms that transition more gradually and to lower extremes.Building areas that are not directly exposed to the sun (i.e.north facing for the northern and south facing on the southern hemisphere) may work as well with good external insulation, unless they are facing strongly reflecting surfaces on neighbouring buildings or in internal courtyards (which may additionally trap heat).

Water installations
One of the most common incidents in a building is water damage.This can be devastating for highly sensitive equipment like microscopes.Especially in lower building levels the risk of water leakage accumulates with the number of floors above.The passage of water in a building during an incident cannot fully be controlled or foreseen, but the location for a microscopy unit should be chosen away from major water conduits or containers (e.g.kitchen equipment) on floors above and should be part of the location planning.This does not mean that microscope rooms cannot have water installations (sinks) where needed for operation, but building drain pipes and reservoirs should ideally be far away.

Electromagnetic fields
For highly sensitive instruments strong electromagnetic fields in the proximity may cause disruptions in the form of background and noise in the measurements.While many commercial microscope systems will be sufficiently shielded, more open custom systems and especially electron microscopes may be affected and the location should be chosen away from possibly disruptive electrical installations or the rooms checked for possibly problematic field strengths.

Accessibility
High-end microscopes are big.And heavy!Large optical table tops easily weigh hundreds of kilograms.While few buildings will allow for the horizontal transport of a fully assembled optical table through all door frames and elevators, care must be taken that all microscope components can be brought to their final station as easily and efficiently as possible and navigate all corners and door frames in the microscope room and in the corridors leading to it.Narrow door frames need to be avoided in the planning.While the access door for daily use does not need to be overly wide (office doors are around 90 cm), it should be in a frame where an opposing hinged element can be swung out to allow a wider passage inside the frame (e.g. to a total width of 140 cm).The inside width of the door frame is often not identical to the real passage width as the hinges are located inside it and it may not be possible to fully swing the doors to the side, especially if the doors open to the inside of the room.Given the size of a fully packaged microscope system, space to temporarily place the crates and also to unpack the system should be considered in the corridors.In case of temporary placements (e.g.equipment demos), storage space for crates needs to be considered.Leaving boxes in the corridor is often not allowed for building safety reasons and possibly degrades the impression of the facility, especially in case of external visits.
As electron microscopy (EM) has even more stringent requirements on the installations, planning a joint space together with the EM core facility can be very helpful leverage in getting a suitable location and technical adaptation.

CONTROLLING THE ROOM ENVIRONMENT
Once the future microscopy unit has a defined location, the creation of a microscopy core facility requires specific adaptations of the instrument rooms for a range of environmental influences that will be covered in the following sections.

Light
In light microscopy, as already stated in its name, the information is conveyed by light.Light in a microscopy room that is not part of the microscopic data generation and that enters into the detection system contributes background and degrades the measurement and at worst makes the measurement unusable.The control of light sources is therefore essential for the design of a microscope room.
Beyond the light switch (or preferably a dimming system) for room lights, this actually means an efficient separation of areas that need light or absence of light at different times and the ability to regulate them accordingly.It is also important to consider that darkness is very problematic for most human operations and actually constitutes a safety hazard, especially when moving around in a poorly lit room.Since a light-sealed room presents its own set of problems, the following should be considered: Absolute darkness is rarely needed.The microscope room is not the detector.It generally contains a computer screen that illuminates the compartment with ≤300 Nits of illumination even during acquisition.While a high numerical aperture objective collects light over large angles and is therefore prone to catch room light, these angles are normally already substantially blocked at the level of the microscope by for example a condenser cover and more and more frequently by microscope encasements that are completely light intransparent or can be covered in addition.In case of confocal pinholes, even significant amounts of stray room light will be rejected by the same principle that creates the optical section itself.It makes sense to work at low light levels for widefield fluorescence imaging where the full collecting power of the objective is used and for nondescanned detection in multiphoton imaging, but the main light blocking mechanisms are these days closer to the microscope itself.Even single molecule detection does not require darkness in the microscope room as much as efficient light protection along the microscope detection path.This can be provided by curtains or blinds around the microscope table or by a microscope encasement (which additionally provides environmental stabilisation against drifts caused by the air conditioning).A benefit of a low light environment for fluorescence microscopy is that it facilitates direct observation as the eye will not be affected by room light at the periphery of vision while observing dim fluorescent structures through the oculars.This is however becoming less and less relevant as the real sample detection has become digital.The historical reason for dark microscope rooms, the direct observation of dim signals by the dark-adapted eye of a human observer, has passed for present core facility operation and the eyes of an observer in a room with a computer screen are not dark-adapted.A reasonable level of desk illumination will actually be beneficial to avoid exposing the eye to the glaring screen illumination through overly dilated pupils.
Rooms can be adapted for light microscopy by removing or covering reflective surfaces (e.g.metallic wall panels) and installing confinements that allow to darken the area and separate the microscope and its control desk from adjacent lit areas.In many facilities, the walls of these compartments are additionally painted black to reduce illuminated surfaces that could be detected by the microscope optics.
The room separations can be brick walls (meaning a permanent room plan that cannot be adapted), movable dry walls or light shielding curtains (possibly made of flame retardant materials if required by building regulations) or a combination thereof, for example a room wall with dry walls on two sides and a curtain on the fourth side serving as an entrance.Importantly, light is easily blocked so the separations can be very light and the choice should be informed by additional considerations.These can be noise shielding (e.g.people working and discussing in nearby areas) and whether the walls of the compartment need to be able to carry installations (e.g.equipment shelves).If a larger room is subdivided into compartments, ventilation and temperature control becomes an issue that will be covered in depth in a separate section.It should however be pointed out that a strongly light isolated microscope cubicle may quickly heat up due to the equipment and operator inside it and not have the required ventilation.
Every microscope room (or subdivision in case of larger rooms with multiple instruments) needs its own light control so that it can be fully switched off (or dimmed) during operation and can be complemented by a desk light on the computer table.It should on the other hand also be capable to provide sufficient light for service interventions and maintenance operations, ideally without affecting other microscopes in the area.In case an accidental activation of the room light (e.g. by cleaning personnel) would be highly disruptive to the conducted experiments a permanent deactivation of the main room lights could be considered, but in that case local lighting sources should be easily reachable and clearly detectable due to the issues of navigating in darkened environments.

Environmental control
A powerful and active room temperature stabilisation is needed to efficiently accommodate complex instruments like advanced light microscopes that generate heat during operation.A good temperature control solution is probably the biggest technical challenge that needs to be considered in the creation of a microscope facility and a free choice may not be possible as this is an integral part of the technical infrastructure of a building and will often not be fully adaptable.
In the planning stage, it is very important to accurately determine the future heat load of the installations and clearly communicate this as well as the temperature tolerance that needs to be maintained.This should be clearly documented for future reference and (unfortunately) be reiterated whenever possible as there may be continued economic pressure on investment cost and a tendency to re-specify both lab as well as equipment rooms to more economic office standards.Again, it needs to be stressed here that such a short-term benefit is far outweighed by the long-term negative consequences during operation.These include the misalignment of optical components in the instruments by temperature changes/fluctuations which lead to increased downtime and more service interventions as well as possibly permanent damage to expensive system components in case of overheating.Aside from the instrumentation, most non-mammalian model systems (animals as well as plants) do not tolerate temperatures significantly above 20 • C. While microscopy cooling systems exist, these are extra installation costs, consume energy and will contribute to the overall heat load of the room as any cooling takes place by heat exchange with the room.
To calculate the heat load (or internal heat gain) of the microscopy unit one can refer to the technical specification and the room requirements of those instruments that have already been decided on.For systems that are not yet specified, a standard confocal microscope with gas lasers may be a good choice as stand-in and would amount to approx.3.2 kW maximum heat load.A separate value should be used for more complex multiphoton systems where at least 7.5 kW maximum heat load (for up to two multiphoton lasers) should be considered.Persons foreseen to be present in the rooms will add approximately 120 W each.All foreseen equipment in the rooms (including not directly microscopy related items like fridges and incubators) should also enter into the planning to make sure that the installations will not be overwhelmed.Margins should be added as retrofitting more capacity may not be easily done.
A target room temperature of 22 • C is a good value for microscopy rooms as it is well matched to the operation of the optical and electronic components of modern microscope system and provides air at a temperature that can efficiently transport heat away from the components when blown over by cooling fans.The temperature regulation width should be specified as maximally 1 • C above and below the set temperature as fluctuations will severely affect the focus stability, alignment and performance of optical equipment.For some commercial superresolution microscopes an even tighter regulation (±0.5 • C) is specified which should be taken into consideration and be discussed with the instrument provider.
A further important consideration is the fully autonomous control of the temperature regulation for all instrument rooms.The environmental control of The connections between the microscope and the components pass through specifically placed openings in the wall (midlevel and floor level).F: Comparison of the temperature stability of a high-end microscope system (Abberior MINFLUX) in the location shown in A (initial location) and its current placing in a dedicated room (shown in D).The upper graph plots the temperature fluctuations in the initial location over a duration of more than a month.Weekends and nights inside the workweek (lower temperatures) can be clearly distinguished from the work days (higher temperatures).In the lower graph, a selected week from the initial location is compared with a representative week in the new location where the temperature never deviates more than 0.5 • C from the set value of 22 • C. a building will run with different settings in office and out-of-office hours to save energy.It has to be made clear from the beginning that microscopy instruments frequently run outside of regular hours and on weekends and that there are expensive components (e.g.multiphoton lasers) that will run and generate heat continuously.These would be easily damaged by an override of temperature control.
Different solutions exist for room temperature control and their suitability varies.A very common solution for smaller rooms is the installation of a wall-mounted air-conditioning unit (Figure 1A), normally close to the ceiling.While such a unit can be powerful enough to maintain the overall temperature of a room, it is a very localised source of cooling and will inevitably create a strong temperature gradient inside the room as well as strong air currents that will make it hard to maintain stability for sensitive equipment.
A more efficient solution that is more adapted for larger rooms are ceiling-based air-conditioning systems (Figure 1B).These can either consist of a central A/C unit that is connected to the building air supply and then regulates the room 4 or a direct connection to the (preregulated) building air supply and extraction that works separately from an internal A/C unit in the same room.Air inlets and extractors can be placed to efficiently cover the whole room and more evenly distribute the temperature regulation.Two parameters need to be distinguished in the room climate control: -External room air exchange (ca.8× room volume/hr) provides the room ventilation.
-Internal room air turnover through the A/C unit provides the temperature control and will vary according to heat load to maintain the set temperature.
A major difference exists between laminar and turbulent air flow solutions (Figure 1B and C).Laminar air flow signifies that the air is moved continuously and uniform in direction and velocity from the outlet and is minimally disruptive as it streams through the room.Low impulse large surface textile-based ductwork on the room ceiling, air socks, 4,5 achieve the same effect.Turbulent outlets blow the air either slightly downward or are very often directed along the ceiling and can be strongly felt depending on the position in the room.In case of strong perceived air currents at a system, baffles at the outlet can be used to redirect air flow away from the microscope system.Instabilities and drift on sensitive systems caused by airflow can be further alleviated by encasing the microscope stands, which is an additional benefit of microscope incubation boxes even when they are not used for active temperature control.
The temperature stability of a microscopy room can be further improved by isolating the system elements that generate significant amounts of heat (e.g.gas lasers, chillers).This can be done by connecting them directly to the air extraction system so that they do not heat up the surrounding area.A very tidy and efficient solution is the creation of service spaces with their own heat extraction on the other side of a microscope room wall so that such elements can placed outside the room itself (Figure 1D  and E).Such spaces can be individual cabinets or centralised corridors for several microscope rooms at once.A major advantage of such an arrangement is that the removal of laser ventilators or chillers from a microscope room also significantly reduces the operating noise in it and improves the working environment while also protecting these components from accidental impacts or spillage.Components should however never be connected to the heat extraction so tightly they cannot ventilate in case of an (in the worst case undetected) extraction failure.If they are placed in a separate room, equipment failures or for example coolant refill warnings could be missed so regular equipment checks in these spaces are needed.
The efficiency of different temperature control solutions is shown in Figure 1F with a ceiling-based laminar flow air-conditioning unit working in conjunction with the pre-regulated building air supply (at 22 • C) and extraction giving a very stable performance.
Building air-conditioning units are often based on fan coil technology that operates with central cooling and possibly even heating water supplies.Ruptures are therefore possible and can be a source of substantial water damage.Whenever possible, the placement of such systems directly in the ceiling of microscope rooms should be avoided.If the units have to be ceiling mounted, they should be placed close to the door so that they are away from the instruments and also easier accessible for maintenance.Air-conditioning units will create and collect water condensation.In case of a blockage this can also lead to water damage.If the installations on the ceiling are covered by a false ceiling, the exit of leaked water can be not only directly underneath the unit but at other points of the room.

Humidity control
As sensitive electronic equipment should not be exposed to high degrees of humidity, active humidity control may be needed in certain climate regions.This means not necessarily a continuously fixed value but a maximum value that should not be exceeded in the microscope rooms.This should be considered in the initial planning of the airconditioning solution and also take into account that past experiences may not apply to future operation due to the effects of climate change as for example warm periods with high humidity levels may increase.Humidity regulation (especially active humidification of dry air) may be limited by local building regulations and also needs to be considered in the context of its energy footprint.A clear understanding of the humidity limits for advanced system operation should be part of the facility planning.
In the case of cryo-microscopy applications (both on the LM and EM side) dehumidification of dedicated spaces will be an inevitable consideration as condensations of crystalline ice on vitreous samples needs to be avoided.

Dust prevention
Dust particles are an inevitable part of our environment, but they are also a problem for complex optical equipment as they can accumulate on sensitive surfaces or affect moving components.Dust intake into microscope rooms therefore needs to be minimised, also because microscope rooms are difficult to clean because of the sensitive equipment and the large amount of electrical and optical fibre cabling.Dust can enter into generally windowless microscope rooms through the door, but also through the ventilation system.Since a high rate of air turnover and exchange is needed in these rooms for temperature control the ventilation system would be a major source of dust and needs to be equipped with highly efficient particle filters.This is also the case for textile-based ductwork where the holes in the air socks need to be protected from clogging.
Norms vary worldwide and have recently been changed in Europe from an EN to a new ISO standard with more realistic criteria (EN 779 to ISO 16890).The EN 779 classification of a combination of an M7 prefilter and F9 fine particle filter (M7 + F9) would hold back dust particles down to a size of 1 µm and provide a good filtration performance for microscope rooms.This would correspond to ISO ePM 1 in the new ISO standard that is based on different particle sizes (10, 2.5 and 1 µm).
In contrast to dust particles down to a size of 1 µm, even smaller particles will not sink down and remain suspended to form aerosols.Removing those through High-Efficiency Particulate Air (HEPA) filters may be relevant in clinical environments and when working with infectious material, but is not generally done for microscope rooms.Planning for optional HEPA filter capability in the initial installation can be considered.
Over-pressurising the microscope rooms helps in keeping dust out and can be considered if feasible.The intake of dust by users entering the microscope rooms can be significantly reduced by adhesive floor mats that retain dust from the soles of the shoes.

Vibrations
Microscopes are vibration-sensitive instruments and therefore can be affected by nearby vibration sources. 6xternal sources are manifold and include vehicular and foot traffic, nearby heavy machinery, building resonances and acoustic noise.They can be permanent and sporadic.
Vibrations are an inevitable occurrence in an active building environment and should already inform the selection of the core facility location as described in a section above.Vibration levels are classified for buildings and rooms by ISO standard 10137 and for more sensitive applications and activities by the vibration criteria (VC) of the Institute of Environmental Sciences and Technology (IEST).They are defined as spectra of root mean square (RMS) velocities over a frequency range of 1 (or 4) to 80 Hz (in 1/3 octave bands, a measure used in acoustic engineering and suited for broadband noise) with decreasing RMS values. 7Even minimal vibrations affect the resolution of highly magnified microscope images and can cause crucial components in the light path to vibrate.Since a microscopy core facility aims to provide high-end technologies the final achievable VCs at the instruments should be between VC-C (<12.5 µm/s) and VC-D (<6,25 µm/s).This can in most buildings only be achieved by using additional vibrational isolators (often floated optical tables) but since the achievable dampening only reduces vibrations by 90% to 99% and varies in performance over the frequency band, the microscope location itself should be as vibration free as possible.This can be optimised by taking into account the considerations for the facility location as stated before.Even locally, placing an instrument close to the load-carrying structures instead of further away can significantly reduce vibrations.Another consideration can be to remove the floor screed from the concrete foundation to place the optical table supports directly on the concrete.Floating screed (as frequently used in living and office spaces) has some dampening qualities but due to its floating properties, it can actually degrade the stability of the foundation of the optical table.The effects of the removal of floating screed are demonstrated in Figure 2.
Since microscopy associated equipment can be a source of vibration (e.g. by ventilators and chillers) moving such equipment away from the microscope table and placing it on its own support 4 can further improve vibrational stability.Properly securing and if needed dampening the cabling and tubing going on the microscope table can also help.While these are not directly room considerations that are the subject of this article they should be made possible by the room planning and the placement of the instrument inside the room.
While planning a facility, the vibrations encountered in designated spaces should be measured thoroughly and under all expected operational conditions.Vibrations can be accurately measured and logged using sensitive dedicated equipment like seismic accelerometers.This is routinely done when checking a room's suitability for transmission electron microscopy but is also useful here, especially for superresolution microscopy, multiobjective instruments like 4 Pi microscopes and for atomic force microscopy.
In addition to vibrations that are transmitted through surface contacts, acoustic noise can also create vibrations and resonances on sensitive microscope systems, which can be quite visible on scanning systems like atomic force microscopes or even confocal microscopes.Just like seismic waves along the surface of a room, sound waves may focus differently in different areas of a room.Even if high ambient noise levels are avoided in the selected microscope spaces, for example the passage of air through ceiling ducts may turn out to create an unwanted resonance that affects the measurement for highly sensitive methods.As such effects are highly individual they will not be covered further here, but can be remedied through a systematic check and optimisation of the affected components and the room.

Electricity supply
The planning of the microscope rooms of a facility needs to include the required number of electricity circuits and number of sockets that are needed to operate all systems in the room.In case of known instrument specifications this will be determined by the technical requirements of the provider, but since the instrument portfolio may change over time the planning should contain sufficient margins.
More generally, at least two separate power circuits are needed for an advanced microscope and its peripherals.
As some critical microscope components (e.g.multiphoton lasers) should be protected from sudden loss of electricity by power grid failure, every microscope location should be provided with an additional circuit connected to an uninterruptible power supply (UPS).Planning for two normal circuits and one UPS circuit should accommodate the requirements of most microscope installations.Each electricity circuit should provide at least three sockets to allow direct connections of all microscope and peripheral system components (Figure 3A).The use of additional extension cords and multiplugs between socket and system should be avoided both for tidiness in the cabling (tripping hazard) as well as for electrical safety reasons (possible overload by 'daisy-chaining').Provisions for three phase electricity circuits (Figure 3B) should be available as needed or be adaptable in the future.These are the standard requirement for some confocal microscopes and need to be available if such systems are being installed.The sockets should be mounted at an easily accessible height above desk level so that they can be easily reached and connected to the backside of a microscope system placed along a wall.Horizontally mounted guide tunnels that allow the flexible positioning of sockets along them are a very good solution that can be routed along all sides of the microscope room that will accommodate microscopes.In case of more complex instruments (e.g.multiphoton systems) that need to be accessible from all sides, wall-connected power cables are an obstruction and the supply should come from the ceiling.
If multiple circuits (normal and UPS) are installed for every microscope in the core facility, the number of circuits quickly becomes significant and clear identification of every single circuit and its associated sockets is needed.Equally important is efficient access to the circuit's fuse and surge protection.Especially the surge protection may at times be triggered.This could of course be an indication of a serious electrical problem in the equipment but may frequently have been caused by a combination of peak power draws by different components (e.g. at start-up) that was not sufficiently dampened to leave the surge protection untouched.The preferable solution is the direct installation of the fuse next to the respective sockets so that it can be immediately reset.Otherwise the well-labelled fuse cabinet should be close by and accessible.An electrical accident is of course a very serious issue and the access to electric circuitry needs to be done fully in-line with all safety regulations of the workplace.In my personal experience, spurious surges are however frequent enough that the placement of and access to the fuses should be thoroughly discussed during planning.Since maintenance interventions on the building power need to be done from time to time a good access to the fuses allows to efficiently prepare the microscope room for such interventions.In case of frequent triggering of the surge protection during start-up, an inrush-current limiter placed between the system and the circuit may alleviate the problem.UPS circuits will not solve the issue as they need to be surge protected as well and will trigger just the same.

Network connectivity
Research microscopes generate huge amounts of data that are either transiently stored on the system computer and then transferred to permanent server storage or even directly written onto the network server.Both solutions require high-speed connections to the institute network that need to be considered already in facility planning.These can be placed inside the same guide tunnels that house the electricity lines (Figure 3A).Given the ever-increasing data volumes, the connections should be specified for at least 10Gbit/s transfer rates.If possible fibre-optic connections should be installed as well (Figure 3A).These are not yet standard for some of the microscope systems, but can transfer data with sufficient speed to the match the acquisition rates of current sCMOS cameras.
To really achieve efficient data transfer from the microscopy facility to the server, the planning discussions have to go beyond the physical installations in the microscope rooms but have to identify all possible bottlenecks between the microscope computer and the server (e.g. the speeds and capacities of the installed ethernet switches) and how many connections can be provided to the microscope rooms.

Gas supply
Since most advanced microscope systems will be placed on floating optical tables that isolate them from residual vibrations and the movement of the operators, these systems will need pressurised air at pressures of up to 6 bar (max.value, normally much less).This is ideally provided by wall feeds next to every system that are connected to the air tables through a pressure regulator with a display gauge.Since the weight and load of the table determines its floating pressure, each table needs its own regulation.
The pressurised air for a table can also be locally provided by a small compressor that activates to re-pressurise when needed instead of a wall feed, but a building supply with wall-mounted outlets at every microscope station is the most efficient solution.
In addition to pressurised air, every microscope station should be equipped with a wall-mounted CO 2 feed with a pressure regulator and gauge to support microscope incubation (Figure 3C).Additional gases are optional.An additional nitrogen (N 2 ) supply would allow to execute experiments at reduced oxygen levels and should be considered in the planning.
Be it for pressurised air or any of the other gases, gas bottles are not a good alternative due to their limited reservoir and the safety requirements for placing and securing highly pressurised containers in the area.
A specific form of gas handling would be the use of liquid nitrogen for cryogenic microscopy.This will not be centrally supplied but brought to the room and stored in highly isolated vacuum flasks and vessels (dewars).These cannot be sealed from the room atmosphere to avoid pressure build-up by evaporation.Steady evaporation in a poorly ventilated room or spilling and evaporation on room temperature surfaces could decrease the air oxygen level in the room and constitute a suffocation hazard.Rooms that are used for liquid nitrogen operation therefore require an oxygen detector that would warn about low oxygen levels.

Maintenance access
Complex equipment like advanced microscope systems need to be serviced in regular intervals and will additionally require service interventions in the case of malfunctions.The planning of the microscope rooms should allow efficient service access to any microscope system while ideally allowing the operation on other systems to continue.Key points here are space management so that all components can be accessed or opened, light management that provides sufficient illumination for the service activities while allowing low light conditions on neighbouring systems and laser safety as the light path may be open during service interventions.

Room planning
The size of the microscope rooms will determine the applicable solutions and distribution of the microscopes and how to arrange the supply lines.Any building space represents a substantial cost commitment space needs to be used as efficiently as possible.The operational footprint of many facility microscopes (not including MP systems and tables) can be approximated as a rectangle of roughly 3 × 2 m (approx.6 m 2 ) that comprise the microscope table, lasers and electronics, the computer desk and the operator space.It can be easily accommodated in a single room of almost any size and a reasonably sized room (around 16 m 2 ) should be able to accommodate two systems.The remaining space would be taken up by the door space and the passage space to the systems that should be spacious enough to not disturb other operators.Closer consideration will immediately reveal additional aspects that are relevant for operation: -The door will most likely open onto a fully lit corridor.Is light protection needed for the microscopes to avoid disruption by persons entering and leaving?-Can training sessions be performed without too much disruption of the other operator?May the presence of a supervisor that wants to discuss the results directly at the microscope interfere with a training session?
Larger rooms come with their own possibilities and limitations.Both fully open 4 as well as subdivided concepts are possible here (example in Figure 4).Open spaces should avoid disruption of operations at adjacent systems while subdivision may allow a closer placement while still limiting disturbances.Subdivision should on the other hand not impede the airflow and temperature control of the rooms.Since the footprint of a frontally operated system is not square but oblong, elongated rooms that allow placement of systems on opposing room sides with an access corridor in between can be used more efficiently than square rooms with a lot of interior depth that is not usable for instrument placement which could be considered dead space.This interior space can however be made accessible by a subdivision that extends from one wall into the room and thus creates two elongated spaces that can be filled with back to back oriented systems, similar to office cubicles in a larger office space.This subdivision could be complete (floor to ceiling) in case the air flow is not affected or incomplete for mainly visual separation allowing free air circulation.In all cases, the number of placeable systems is inevitably smaller than the room size divided by the average operational footprint of the systems.At least 30% need to be added for the room entrance space as well as access routes.Approaching this number will lead to problems in installation, maintenance and possibly operation.Even with subdivisions all room concepts need to maintain clear escape routes in case of any emergency.
In case the installation of a more complex system that requires all around access is planned for the future, subdivision planning can already take this into account to avoid disruptive refurbishments at a later point.
An example of a room design with a combination of room walls, sub-partitions and curtains and its possible evolutions are shown in Figure 4.
Sliding doors instead of swinging doors have two advantages to be considered: They require less space as an opening door will not swing through additional space or impact with an object in its arc trajectory.The door displacement during opening and closing will also be less disruptive as it creates less turbulence than a swinging door that will fan air as it moves along its arc.
In most cases, the decision between larger multisystem rooms or individual microscope rooms is made early on when deciding on the location of the facility inside the building and taking into account the architectural layout of that area.In case the distribution of load-bearing elements still allows a more flexible planning at this point (e.g.subdividing into several rooms with dry walls, here are some considerations for choosing larger or smaller rooms: -Access control is sometimes needed (in case of sensitive equipment or confidential installations/experiments) and is easier implemented in a small room concept.-Larger rooms use available space more efficiently as for example curtains between systems allow easier access to the sides and back of systems than solid walls that require a correspondingly larger instrument footprint that takes the access into account.
An additional consideration for core facilities is access to wet lab space.Even if the facility is mainly servicing internal users and these users have access to the spaces of their own labs in the institute, some sample preparation will have to be done on-site.Minimal wet lab capability is therefore needed and can be provided for example by some bench space in the entrance area as illustrated in Figure 4.This can comprise a small incubator for short-term storage, a fridge and other basic equipment.If significant (open) access provision for external users (e.g. in research infrastructures like Euro-Bioimaging) is intended, more complete wet lab facilities are needed so that a small lab space should be included in the planning.The access to cell and tissue culture facilities that allow a whole experiment workflow also needs to be considered in this case.The microscope rooms as well as any preparation spaces need to comply with the biological safety-level specification for the samples used in the facility.Guidelines for laboratory planning exist 8 and are not covered further here.
This article deals mainly with the room planning and setup for a microscopy facility and not with the systems and instruments that it will accommodate.It is however likely that the facility planning and instrument selection phases will overlap and can accordingly inform each other.General examples of the imaging method affecting the planning can be found several times throughout the text, but it is worthwhile to point out here that the microscope providers are a very useful source of information and can provide important input on the instrument requirements that need to be met and of course also on how to meet them.

Noncommercial and very advanced systems
Even though imaging core facilities aim to offer the latest cutting-edge services, these will mainly be offered on commercial systems as these are best suited for user training and operation and for reliable service provision.While the lasers on most of these systems are technically IEC category 3B and therefore unsafe for intra-beam viewing, 9 most confocal microscopes have strongly divergent beams and accessible power levels like less harmful 3R level lasers. 10n case the facility also offers access to noncommercial systems additional considerations need to be taken into account.While commercial systems are generally certified for safe operation (not maintenance!) and the operator is protected by a clear usage protocol as well as physical safeguards like laser interlocks, self-built systems may not have all of these in place and for example have lasers passing freely through open spaces.Rooms that accommodate such systems need to comply with national safety regulations and have for example a switchable 'Laser on' warning light outside their entrances as well as a clear notice of the laser class used in the room and what precautions (e.g.safety goggles) need to be taken when entering.Door activated interlock solutions should be considered and implemented where possible.
The maintenance of open laser beams and especially any maintenance or alignment of multiphoton systems with their powerful and invisible infrared lasers needs to be done in a space that is completely shielded against any accidental exposure of other people and requires a fully closed work area.In case lasers are located in adjacent equipment rooms next to the microscopes (as described in the environmental control section above) such service interventions can be easier executed since the room separation is a given.

Facility building from the bottom up
As stated in the introduction, it was the intention to cover all relevant aspects that go into setting up a facility.Does this have relevance if not all points can be addressed in one's own situation?The author would hope that the answer is 'yes'.This is also meant to be a guideline for future growth.The treatment was aimed to be exhaustive since an incomplete coverage would not serve its purpose.Microscopy core facilities are success stories.They may start out small, but they have a tendency to grow because they are needed.As an overarching treatment it makes sense for the article to start with the overall location and then define rooms and spaces.The points in the article can however also be applied in reverse.A facility will grow in this way.If you have a room, start with the room considerations.From a room to a better room to a suite of rooms to a relocation into an area that is specifically refurbished for microscopy.And possibly into a new building if the institute moves or a new institute is created (and when a location can be chosen).
Approaching facility setup from this growth perspective and starting at the level of the instrument and for almost any room one can arrive at the following prioritisation: 1) Electricity supply: Most research grade light microscopes are complex enough that sudden power cuts can damage sensitive components.In case of electrical instabilities, the first priority is to establish a stable supply by using protected lines (if available) or a local uninterruptible power supply for critical components.2) Temperature control: Advanced microscopes can only operate in a defined temperature range outside of which damage can occur.Room temperature control is therefore needed at a first level to allow microscope operation at all and tight regulation during operation is needed for advanced systems to maintain their performance parameters and provide a reasonable return on the investment into an advanced system.3) Vibration control: Vibrations ultimately affect resolution and accuracy of microscope measurements.They do however only become really relevant for advanced microscopy methods (confocal and similar) where the following rationale can be applied.The acquisition of an advanced microscope is a significant investment.Vibration mitigating measures (antivibration table and/or a relocation to a better room) are a frac-tion of the cost of the instrument.For the institute or funder to have any return on the total investment the measures need to be taken or there will be no return.
If these measures are taken and prioritised accordingly increasingly complex instruments can be successfully offered for service.Successful service provision can lead to growth and the installation of additional systems.With growth and more instruments, light management, room separation and data connectivity become relevant.Heat load increases and A/C strength may need to be adjusted.More advanced methods (e.g. in vivo imaging) will require gas supply.And one by one, the above covered topics may become relevant as a result of systematic and responsible growth.

CONCLUDING REMARKS
The planning of the rooms of a light microscopy core facility needs to cover a wide range of technical and operational considerations.Experience, common sense and the study of existing solutions in other places will allow to design a good overall solution.There are however so many technical fields that need to be covered that the expertise of most microscopy specialists is inevitably (and at least in the case of this author easily) exhausted.The current article can only provide general pointers, caution against insufficient solutions and advocate ample margins of tolerance in the crucial performance parameters.It is essential to communicate closely with the relevant specialists on the building management side and interact as partners in the creation of the facility rooms.They are the ones that need to provide the technical execution and it is the microscopist's job to convincingly explain the need for certain operational parameters.Ideally, they will be your partners and allies in this undertaking, but they will be continuously confronted with conflicting demands and economic pressures.
A close and frequent communication with them is needed as plans will be modified and decisions revisited.Specifications will be communicated down the line and may then be over-ruled by an architect who may not fully appreciate the difference between an office building and a scientific institute.Re-checking the numbers of electricity circuits, network plugs, size of the doorframes as well as any other parameters throughout the planning and building phase is therefore advised.

A C K N O W L E D G E M E N T S
The author acknowledges the Imaging Centre at the European Molecular Biology Laboratory (EMBL IC), generously supported by the Boehringer Ingelheim Foundation.The author would like to thank Clemens Kney from

F I G U R E 1
Temperature control in microscope rooms.A: Wall-mounted air-conditioning unit.B: Ceiling-mounted air conditioning with one pre-regulated air inlet and three laminar flow A/C outlets for cooling in the centre.Round openings: air extractors.An optional air extraction pipe is installed in the ceiling on the right side of the back wall.C: Ceiling-mounted turbulent A/C with central intake and four cooling outlets along the ceiling (blue arrows: air flow directions).D: Highly temperature stabilised microscope room solution with ceiling-mounted laminar flow air conditioning and the heat-generating microscope components placed in a separate compartment with its own heat extraction.E:

F I G U R E 2
Vibrational control for sensitive microscope systems.A: Comparison of vibration measurements with a seismic accelerometer on concrete with floating screed on top and directly on the concrete (cut-outs).The overall vibration level is significantly reduced on the bare concrete.B: Vibration frequency spectra of floating screed (magenta), concrete (green) and on the antivibration table (grey).Blue line: VC-C, green line: VC-D, cyan line: VC-E, grey line: VC-F.

F I G U R E 3
Microscope room installations.A: Wall-mounted supply panel for electricity and network connectivity (middle: fibre-optics, right: network cable plugs).B: Three phase electricity socket and cable mounted in the same supply panel.C: Gas supply lines for pressurised air (right), carbon dioxide (middle) and nitrogen (left) with pressure regulators and gauges.Main flow interrupters are installed upstream and visible at the top of the panel.

F I G U R E 4
Design example of a 70 m 2 microscope room.A: A combination of wall panels and curtains creates five subdivisions (one with an additional separation) for up to six microscope systems.Left: Arrows indicate movable curtains.Middle: Rooms fully opened, arrow indicates a collapsible wall separation to create one continuous space and access for installation.Right: Fully opened configuration.B: Left: Room plan with six microscope systems (grey), two processing stations (green) and bench space (blue).Asterisk: Electricity cabinet containing the fuses of all circuits in the room.Right: Minimally adapted alternative configuration with a big room for a multiphoton laser shared by two microscopes and the same number of overall systems.