Labome conducts systematic surveys of reagents and instruments cited in formal publications. Table 1 lists the types of microscopes among the publications. Confocal microscope appears to be the most cited microscope. However, this does not mean that confocal microscope is more prevalent than other types of microscopes. It is likely that publications tend to not indicate/mention a specific microscope when a conventional, routine microscope is used. In addition, our selection of articles is restricted to certain journals (initially mainly the journal Science, and currently PLOS Biology and eLife), thus, any bias in the research areas presented in these journals will be reflected in our survey data. The image acquisition and analysis software programs associated with microscopes are discussed in a separate article.

Certain ambiguity exists for publications with fluorescence microscopes, since they could be either upright, or inverted. The grouping of the microscopes is based on the explicit statement with regards to the type of microscope in the publications. In addition, a citation instance is only categorized in one type of microscope, for example, an inverted fluorescence microscope is categorized as a fluorescence microscope, but not counted as an inverted microscope in our summation.

There are four major suppliers of microscopes, as listed in Table 2. They are ZEISS, Leica Biosystems, Olympus, and Nikon.

Confocal imaging can usually be achieved through a confocal unit mounted on a microscope body, using a pinhole to create the confocality and the optical sectioning, necessary for 3D imaging. Confocality is either achieved with a single pinhole and a raster scan system projecting the image on a point source detector is used or with an array of pinholes and projecting the image on a camera, the latter is the case of the spinning disk [19].

Table 3 lists the significant suppliers of specific confocal microscopes and some of their most cited models. Table 4 lists the technical details of the various confocal microscopes of today and the additional modules-techniques that can be added on them.

ZEISS LSM is the most cited confocal microscope among the surveyed publications. The current model version of ZEISS confocal microscope is LSM 8 family which includes ZEISS LSM 800 and ZEISS LSM 880 models (Figure 1). In the past years, the first generation of LSM confocals, ZEISS LSM 510, was used to address the molecular mechanism underlying the coronin 1-dependent activation of cAMP/PKA signaling [24], to study the controlled calcium signals by PKG via phosphoinositide metabolism [25], to study the role of BMP signaling pathway in setting the circadian period in PDF neurons in an adult brain [26], to investigate the structural composition of caveolar coat complex [27] and the regulatory mechanism of epithelial growth and autophagy in Drosophila larvae [28]. LSM 710 was cited in publications describing the regulatory mechanism of epithelial growth and autophagy in Drosophila larvae [28], the genomic features of germinal cell lineages in Schistosomes [29], the roles of VAL- and TMT-opsins in neuronal photoreception in medaka fish and zebrafish [30], the roles of calmodulin-binding protein Crag in Drosophila photoreceptor cells [31] and the resemblance between the interspecific hybrids and piRNA effector-protein mutants in Drosophila [32]. Maya-Ramos L and Mikawa T acquired multiphoton time-lapse chick embryos images with a Zeiss LSM 7 MP, W Plan-APOCHROMAT 2-photon microscope [33].

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Figure 1. ZEISS LSM 880 with Airyscan. Photo courtesy: ZEISS.

Through the years, new versions of LSM appeared, with the LSM 980 being the most recent. Table 4 shows the different confocal microscopes, of the 4 main constructors, with the microscope characteristics. As we see, for Zeiss LSM, the main advantage is the early implementation of sensitive Gallium arsenide phosphide detectors (GaAsP) that have much higher sensitivity at the 400-650 nm range than traditional photomultipliers PMT. They are not highly sensitive outside of this wavelength range, so they are typically used in combination with classical PMTs.

For spectral imaging, a 32x array of GaAsp detectors enables simultaneous imaging on a wide wavelength range (400-700 nm). Recently, very interesting modules can be implemented to LSM confocal microscopes. This is the Airy scan unit, that allows higher resolution images (down to 140 nm) and a more sensitive imaging by using the 32x detector array. Another very interesting mode is the fast-Airy scan, that enables fast confocal imaging. At the most recent version of LSM microscopes, like the LSM 980, one can achieve scanning speeds of 13 fps at 512x512 pixel image size at normal confocal mode and go up to 34 fps when using fast Airyscan Multiplex mode. This technical innovation facilitates live cell imaging.

Leica develops multiple solutions of confocal microscopes [34]. Table 4 shows the various Leica confocal microscope versions. In terms of applications, Leica TCS SP5 confocal microscope was used to study the mechanism by which Par1b induces asymmetric inheritance of plasma membrane domains in proliferating hepatocytes [35], the mechanism by which the downy mildew effector attenuates salicylic acid-triggered immunity in Arabidopsis [36], the role of the dual leucine zipper kinase signaling pathway in regulation of dendritic and axonal growth [37], the regulatory effect of TRPM5 activation on MUC5AC secretion from human colon goblet cells [38], and the rational drug design for castration-resistant prostate cancer without mutation-based resistance to antiandrogens [39]. SP2 confocal microscope was used to study the mechanism by which extracellular signals synchronize pacemaker neuron clocks [40], the asymmetric inheritance of plasma membrane domains in proliferating hepatocytes [35], the role of autophagy in glycogen breakdown [41], the mechanism of myelin membrane assembly [42], and the resemblance between the interspecific hybrids and piRNA effector-protein mutants in Drosophila [32].

Technically speaking, Leica technology uses Acousto Optical Tunable Filter (AOTF) for laser line selection and intensity control with microsecond precision. An Acousto Optical Beam Splitter (AOBS) replaces fluorescent filters and beam splitters in the light path. It is a spectrally tunable element which is fast, switchable and without any moving parts. In combination with an AOTF, the AOBS offers a more light efficient system. This optical element allows the excitation light to excite the sample and the emission passes straight through the crystal.

The SP5 and SP8 version have a Tandem Scanner which comprises of a conventional scanner for high resolution morphology imaging (8kx8k) and a resonant scanner for fast imaging. Conventional speed ranges from 10-1400 Hz whereby resonant speed is 8000 Hz. For a 512x512 pixel image, the speed can go up to 40 fps. In terms of detection, Leica offers the Hybrid detector (HyDs) solution. A GaAsp PMT photocathode is used for the initial photon detection and the photoelectrons are then detected on an APD sensor (avalanche photodiode) in order to amplify more the signal.

Nowadays, the most recent Leica confocal microscope is the TCS SP8. It can be combined with a white laser source, to offer broad choice of excitation wavelengths. More interestingly, it can be combined with additional modules, to allow super-resolution STED microscopy (for example, [43] ), fast lifetime imaging microscopy (FALCON) or 2-photon microscopy with a spectrally tunable detection system (DIVE). In 2019 an image treatment module was added at the standard SP8 confocal. The LIGHTNING module offers an adaptive treatment process for extraction of hidden information in the image. Up to now most of treatment methods are using a global set of parameters for image clearing.

Olympus confocal microscopes are shown on table 4. In terms of applications, at the very beginning, Olympus FV1000 laser-scanning microscope was used to study the mechanism by which extracellular signals synchronize pacemaker neuron clocks [40], the mechanism of tip link regeneration in auditory hair cells [44], the functional property of the MCU gene [45], the mechanism of vessel growth and pruning in the developing midbrain vasculature in zebrafish [46], and the retrograde synaptic signal in C. elegans [47].

Nowadays, Olympus new confocal FV3000 has even higher technical performances, with the sensitive GaAsP detectors, the fast imaging with the resonant scanners and the TruSpectral detection technology. The TruSpectral detection is based on the patented Volume Phase Hologram (VPH) transmission and an adjustable slit to control light, making the spectral detection highly efficient, enabling users to select the detection wavelength of each individual channel to 2 nm.

Nikon has also developed confocal microscopy solutions. The most recent is the A1R microscope (Table 4). Its main advantage is the new resonant scanner that supports both high speed and high definition imaging, achieving a maximum resolution of 1024 x 1024 pixels at 15 fps. With both 1024 x 1024 pixel resolution and a large field of view, the new resonant scanner delivers higher throughput in various imaging applications. When combined with the Ti2-E inverted microscope body the imaging area of the confocal is 25 mm nearly twice of the conventional 18 mm, enabling the researcher to obtain significantly more data by capturing a wider region in each shot. GaAsp detectors are implemented for detection and diode lasers are used for excitation.

The spinning disk microscope is a confocal microscope that uses an array of pinholes and an array of microlenses that are mounted on spinning wheels that project the pinholes on the whole sample area. The detection is made on a camera, thus the acquisition times are much shorter than the classic confocal raster scan microscope. The low bleaching and phototoxicity make spinning disk microscopes the adequate tools for 3D live cell imaging. The excitation is made with lasers, most commonly for the commercial systems 4 laser lines are used, at 405, 488, 561 and 640 nm. For recent microscopes the laser intensity is modulated directly by the software with fast ON/OFF control and there is no need of AOTF, facilitating the acquisitions. The detection is made on a camera, most commonly either an EMCCD (for weak signal detections) or a sCMOS (for fast imaging and a wide field of view). Dual camera units are almost always proposed by the constructors for simultaneous 2-color imaging.

Table 5 presents the main companies that commercialize spinning disk solutions. There are either companies that commercialize the whole system (spinning disk unit and the microscope body) or smaller companies that provide the spinning disk unit that can be mounted to different microscope bodies. For example, Maya-Ramos L and Mikawa T acquired time-lapse chick embryo images with Nikon Eclipse TE2000-E supported by Hamamatsu ORCA-Flash 2.8 camera and Nikon Eclipse Ti supported by ANDOR iXon camera [33].

The most common spinning disk unit is the CSU-X1, that turns at a speed of 10000 rpm allowing 2000 fps imaging speeds. It is ideal for living cell imaging where we need high acquisition rates. Most often a 6-position filter wheel is provided for the emission.

The next generation spinning disk unit is the CSU-W1. Here the space of the pinholes is wider, something that reduces pinhole crosstalk on the sample, enabling more clear observations and deeper imaging. Most often a 10-position emission filter wheel is provided. The pinhole size can be 25 or 50 μm. The former value is appropriate for low magnification objectives and the latter aperture value for high magnification objectives.

Recently, high resolution modules are provided by many constructors. They are based on optical re-assignement methods, by adding additional microlenses wheel that will improve the image formation. The resolution can be improved by approximately 1.4x and combined with a deconvolution we can achieve resolutions of 100-120 nm along xy and 300 nm along the z axis.

Andor offers the last years improved spinning disk unit solutions (see Table 5 for product details). First are the XD and WD, that are almost identical to the CSU-X1 and CSU-W1 described above. The detection is made on the Andor EMCCD or sCMOS cameras. More recently a new generation of spinning disk units was launched, the Dragonfly series 200 and 500. Here the pinhole size is slightly modified to 40 μm (instead of 50 μm) to fit also at 60x objectives. A Borealis module is also added at the excitation, for homogeneous illumination, quite important for tile scanning experiments. A camera zoom and an illumination zoom are also provided for the Dragonfly 500 module. GPU deconvolution is incorporated, allowing live deconvolution, very interesting for finding the adequate deconvolution parameters. A SRRF image treatment mode is also provided, to increase image contrast and resolution. A mode that can be interesting for some kind of samples, but for other it can create artefacts. Andor Dragonfly spinning disk unit is commercialized on Nikon or Leica microscope bodies. IA Klein et al, for example, performed live cell imaging with the Andor Revolution Spinning Disk Confocal microscope [16].

Nikon offers full solutions of spinning disk. It offers the classical CSU-X1 and CSU-W1 units, mounted on the recent Ti-2 Eclipse microscope body. What is very interesting here is that this microscope offers the wider field of view of all microscope bodies (25 mm) and thus coupled with the CSU-W1 and a sCMOS camera it can offer a really wide area to study of the biological sample without the need of additional optics. The camera proposed by Nikon is most often a CMOS Prime 95B or pco edge camera. There is also the possibility to add the SoRa module on the CSU-W1 unit, for high resolution imaging.

Nikon provides also silicone immersion objectives (25x, 40x, 100x). These objectives are designed for deep tissue observation. Observation depth is negatively impacted by spherical aberration caused by refractive index mismatch. The refractive index of silicone oil (1.40) is close to that of living cells or tissue slices (1.38), minimizing sperical abberations and enabling deep imaging down to tens of micrometers.

Olympus offers also complete spinning disk microscope solutions. They commercialize the SpinSR100 microscope, which is in fact a CSU-W1 unit coupled with the SoRa module. The detection camera can be a sCMOS, most often a Hamamatsu Flash 4. What is interesting here is the remote correction collar unit that is used to adjust the lens position within the objective to correct for spherical aberration caused by refractive index mismatch, resulting in dramatically improved signal, resolution, and contrast. Olympus provides also silicon objectives, that as we saw above minimize spherical abberations and allow deep imaging.

Apart from the main microscope constructors (like Nikon and Olympus) other companies provide only the spinning disk unit that can be mounted on different microscope bodies. Gataca and Cairn companies commercialize the CSU-X1, CSU-W1 units. Recently they provided the Live-SR module, that can be mounted on either the CSU-X1 or the CSU-W1 unit. It is based on optically demodulated structured illumination methods and combined with online processing can offer high resolutions (down to 110 nm lateral).

The Yokogawa company was the first one to commercialize the spinning disk units. Now it provides all type of CSU units that can be mounted on a microscope body, coupled if needed with a SoRa module. The 3i company is also providing the SoRa module, that can be mounted on a CSU-W1 unit.

A fluorescence microscope is an optical based microscope used to study organic or inorganic substances. Fluorescence microscopes are based on the epifluorescence microscopy. Here we divide our research on inverted and upright epifluorescence microscopes.

Table 6 lists the major manufacturers of inverted microscopes among the surveyed articles. Table 7 shows the main inverted microscope models of the 4 main manufacturers and lists their advantages. There is a special column for the focus control system along the z axis. This is an essential tool for live imaging, because due to temperature differences and mechanical drifts, it is very common that the sample moves along the z direction. The focus control system, by most often using an infrared laser, manages to keep the sample in focus.

ZEISS Axiovert 200 microscope was used in studies to understand the mechanism of temperature-responsive virulence change of Histoplasma capsulatum [48], the role of sFLT-1 in the maintenance of the murine avascular photoreceptor layer [49], the mechanism of tip link regeneration in auditory hair cells [44], the molecular architecture of SpoIIIE pump and the mechanism of its recruitment and assembly [50], and the role of hormonal signal amplification during Caenorhabditis elegans development [51].

A more modern version of Zeiss microscope, the ZEISS Axio Imager Z1 microscope was used to investigate the functional property of CK2 kinase in Drosophila [52], the role of hybrid Th1/2 cells in natural immune responses against parasites [53], the epigenetic regulation of gene expression in budding yeast [54], the axial patterning in Caenorhabditis elegans [55], and the functional properties of MORC family ATPases [56].

Nowadays, Zeiss uses the Definite Focus 2 system to keep the sample in focus over time. As excitation sources, Zeiss proposes the Colibri, a 13 LED illumination. Of course, Zeiss microscopes can be coupled with different X-Cite lamps or other type of sources like HBO and mercury lamps. Colibri is a fully software integrated Zeiss solution. The advantage of LED sources instead of X-Cite or the mercury lamps is obvious, as their lifetime is longer and there is no explosion risk. Traditionally, Zeiss provided Apotome solutions to the epifluorescence microscopes. Apotome module improves the spatial resolution of the resulted image. It is based on structured illumination, by using an evenly spaced grid in the aperture plane to serve as a mask through which the specimen is illuminated. However, the acquisitions take more time. It can be a very useful solution for fixed sample observations, especially when there is no confocal microscope nearby. Lately, Zeiss has integrated at the acquisition software a deconvolution solution. Equipped with a strong PC with GPU card, the system is able to give live deconvoluted images, something very interesting when testing various deconvolution parameters on the observed sample.

Nikon TE-2000 inverted microscope and its successor Eclipse Ti microscopes were used to study the regulation of BMP-dependent cardiac contraction by 3-O-sulfotransferase [57], the mechanism of MYRF self-cleavage [58], the mechanism of tip link regeneration in auditory hair cells [44], the structure and formation principles of Vibrio cholerae biofilms [59], bacterial type 6 secretion system [60], the molecular architecture of SpoIIIE pump and the mechanism of its recruitment and assembly [50], the role of radial glial progenitors in stabilizing nascent brain vascular network [61], the role of cMyBP-C during cardiac muscle contraction [62], and the binding of lac repressor [63].

One of the main advantages of Nikon microscopes is their stable z drift compensation system, the so-called Perfect Focus System (PFS). Recently Nikon develops microscopes with a two-deck multi-port design that offers great expandability for various applications. The great field of view (FOV) of the brand new Ti-2 Eclipse microscope of 25 mm is very useful when coupled to sCMOS cameras, as it maximizes the sensor used area without making compromises or using additional lenses.

Olympus IX71 inverted microscope was used to investigate the structural composition of caveolar coat complex [27], the mechanism of Golgi-mediated protein traffic [64], the role of cell adhesion and cortex tension in cell sorting during gastrulation [65], the conformational changes of bacterial RNA polymerase clamp during gene transcription [66], and the structure of CLOCK:BMAL1 transcriptional activator complex [67].

The more ancient versions of Olympus microscopes, Olympus BX51 and BX51W1 microscopes were used to study the roles of Vav2 and Vav3 in skin cancer [68], the roles of VAL- and TMT-opsins in marine neuronal photoreception [30], the effect of green leaf volatiles on moth ovi position [69], the npas4-dependent transcriptional program in CA3 [70], and the induction of Pf sporozoites specific CD8+ cells [71].

Olympus latest microscope design (iX83) offers great body rigidity and a two-deck port design. Olympus simplified or shortened structures from the focusing handle to the revolving nosepiece, thereby minimizing warp in the image traveling section.

Leica DMI 6000 fluorescence microscope and its variation DMI 6000B were cited, for example, in publications investigating the mechanism of myelin membrane assembly [42], the dual roles of dihydrolipoamide acetyltransferase in gene expression and carbon metabolism [72], the spatial coupling of catabolic and anabolic machinery in facilitating the synthesis of secretory proteins [73], or the apoptotic trigger waves [74].

The latest Leica microscope body (DMi8) offers a great stability and a large field of view.

It should be noted that nowadays almost all manufacturers offer deconvolution solutions integrated to their acquisition software. This is quite convenient for the microscope user, as the images can be directly deconvoluted. When coupled to GPU, the deconvolution can be much faster, which is now the case for most of the solutions. Leica went a step further and added an image clearing before or after the deconvolution process combined with an adaptive treatment process for extraction of hidden information in the image. Up to now most of treatment methods are using a global set of parameters for image filtering. The new version of such a microscope is called Leica THUNDER.

Table 8 lists the major manufacturers of upright microscope among the articles Labome has surveyed.

• ZEISS Axioplan 2 microscope

  ZEISS Axioplan 2 microscope was used to investigate the regulatory role of Erf2 in protein palmitoylation and meiotic entry in Schizosaccharomyces pombe [75], the roles of VAL- and TMT-opsins in neuronal photoreception in medaka fish and zebrafish [30], the mechanisms of meiosis I chromosome segregation pattern in germ cell development [76], the role of FtsZ in cytokinesis [77], and pyruvate uptake [78].

• ZEISS Axioskop 2 microscope

  ZEISS Axioskop 2 microscope was used to investigate the role of Etv2 in vascular development [79], the modification of electrical synaptic strength [80], the LR asymmetric tissue morphogenesis [81], the role of Hif-alpha/Notch interaction for crystal cell functions in Drosophila [82], and the expression of immunosuppressive enzyme IL4I1 in B-cell lymphomas and tumor-associated macrophages [83].

More recently, Zeiss upright microscopes are the Axio Imager Z1 or Z2, that can be easily combined with imaging systems like ApoTome2 and laser scanning microscopes (LSM).

Nikon E600 and 808 microscopes were used in histological examination of inner ears [10].

Leica DMR microscope was used to study chicken embryo axis elongation [21], the mechanism of establishing temporal read-outs of gene expression in a fast-developing chordate embryo [84], the molecular mechanism of organismal death in nematodes [85], the involvement of Fam20C in the phosphorylation of extracellular proteins [86], and the evolution of bacteria [87].

During the last decade there is a revolution in microscopy with the techniques of super-resolution microscopy. The inventors of some of these techniques were awarded by the Chemistry Nobel price in 2014. They allow going beyond the diffraction barrier and seeing details that were unknown before. The aim here is not to analyze the super-resolution techniques. We present the main super-resolution microscopy manufacturers with their products and their characteristics. Table 9 shows a synthesis of the existing commercial microscopes.

The SIM technique gives a 2x resolution improvement in the 3 axes, allowing resolutions down to 100 nm at xy and 300 nm at z direction, after a mathematical treatment of the acquired images. Zeiss launched in 2012 the ELYRA microscope, using gratings that create this structured illumination pattern at 4 colors. In 2019 Zeiss presented the new ELYRA 7 microscope, that combines lattice illumination with SIM and thus offers much faster acquisitions (255 fps) allowing SIM imaging in living cells. IA Klein et al, for example, acquired immunofluorescent images of h frozen breast and colon tissues with the Elyra Super-Resolution Microscope at Harvard Center for Biological Imaging for the analysis of biological condensates [16].

Nikon is also fabricating a SIM microscope, the N-SIM, offering fast 5 color imaging.

General Electrics is another traditional SIM manufacturer with the OMX series [88]. The OMX Blaze offers high acquisition speeds for 3D multicolor SIM.

The STED technique is a full optical super-resolution technique where no image treatment is necessary and offers down to 50 nm resolution in xy and down to 80 nm along the z axis for biological samples. There are 2 manufacturers of commercially available systems (Table 9).

Leica first commercialized the STED microscopy, adding a STED module on a SP5 confocal system. Now it offers a 3D STED microscope, mounted on a SP8 confocal, with a time-gated option to gain in depletion power and up to 3 depletion lasers for wide labeling applications.

The other company that commercializes STED microscopes is Abberior Instruments. They propose two main solutions. The STEDYCON, that is a simple STED module that can be added on a microscope body, offering 2D STED imaging with one depletion laser. The other solution is the complete STED microscope, with different versions, like the expert or facility line. Here 3D STED is possible, with up to 3 depletion laser lines, and an optimized method to autoalign the laser beams. Different improvements of the system have shown up, like the DyMIN and RESCue STED systems, using adaptive illumination. They offer fasted imaging with lower STED power, thus more compatible for living cell imaging.

The other big family of super-resolution techniques is the PALM/STORM microscopy. Here the technique of Single Molecule Localization Microscopy (SMLM) is used that allows localization precisions down to 10-20 nm. Depending on the fluorophore that is used, the techniques are divided in PALM microscopy (for photoactivated proteins) and STORM microscopy (for organic dyes). Due to the high localization precision that is achieved, these techniques are very interesting but at the same time they hide many artefacts and special care has to be taken when analyzing the data.

PALM/STORM is the easiest of the super-resolution techniques to mount on a microscope. The trickiest part is the data analysis. Here we present some commercial solutions, with some of them including a data analysis module at their software (Table 9).

Zeiss commercializes the ELYRA P.1 microscope, equipped with 4 laser lines, that is dedicated to PALM and often STORM imaging. A 3D module is available.

Nikon on its side produces the N-STORM microscope, dedicated for STORM imaging (but can also perform PALM), equipped with powerful lasers, dedicated objectives that stand high powers and a 3D module.

Leica STORM microscope is the GSD one, equipped also with a 3D module.

Lately, new start-ups have emerged, providing very interesting solutions. One of them is the Vutara system, commercialized by Brucker, offering an easy 3D module based on a biplane method. Another interesting solution is the microscope of Abbelight called SAFe 180 or 360 (depends if one or 2 cameras are on it), offering a high 3D resolution improvement, higher that the other systems along the z axis.

It worths mentioning that in 2019 Abberior is commercializing a MINIFLUX microsope. This is a brand new technique, combining SMLM and STED, localizing individual switchable fluorophores with an excitation beam that has a doughnut-shape. The technique is shown to give 3 nm resolution along the 3 axes, in fixed and living cells. A very promising technique that can throw light to many biological processes and protein distributions.

The stereo microscope is a binocular microscope that gives most of the times a low-power stereoscopic view of the subject.

It exists more than 30 companies that commercialize nowadays more that 100 stereo microscopes. Table 10 presents the 4 main stereo microscope manufacturers with their main products. Stereo microscopes can be upright or inverted, bench top or compact. Always they offer a bright-filed observation mode, and sometimes there is a fluorescence option, or even a dark field.

The sources can be LED or halogen lamps. The way light illuminates the sample is important, so more often it is provided a LED ring light for coaxial, diffused or transmitted illumination and a halogen fiber optic ring light or dual pipe light (to illuminate from both sides).

Regarding detection, the simple case is the observation with the binoculars, but for recording purposes, a camera can be added, a CCD or even better a sCMOS to offer a wider field of view.

They can also be motorized, to facilitate automatization at the acquisitions. The zoom is an important characteristic and the value that will be used always depends on the kind of sample and degree of detail that the user wants to observe.

Leica stereo microscope was used to investigate the role of a new sphingolipid-TORC1 signaling pathway in postembryonic growth and development of Caenorhabditis elegans [89], the IGF pathway [90], leaf shape formation [91], phloem transport [92], and the formation of actin filaments [93].

Leica offers a wide variety of stereo microscope solutions, from simple ones with only transmission and binocular observation to high end ones, equipped with cameras and high zoom. The Leica M165 FC is an interesting microscope, as it offers a fully apochromatic corrected 16,5:1 zoom optics resolving structures down to 1.1 micron. Leica commercializes the 2x PLAN APO objective, with NA 0.35, the highest for stereo microscopes. The advantage of this objective is the adjustable refractive index, improving observations of samples immersed in water.

ZEISS stereo microscope was used to investigate the role of Etv2 in vascular development [79], the molecular mechanism for feeding [94], the role of cMyBP-C in the process of cardiac muscle contraction [62], the roles of the gene dll in the evolution of sexually dimorphic traits of Rheumatobates rileyi [95], and parallel evolution in ants [96].

The most advanced stereo microscope for biology is the Axio Zoom V16. It is a motorized fluorescence microscope, with a 16x zoom and high numerical aperture objectives (0.25 NA).

Nikon develops a variety of stereo microscopes, from simple ones (like the SMZ800N on the Table 10) to more advanced ones (SMZ25) The most advanced is the SMZ25, that combines macro and micro imaging for easy viewing of single cells to whole organisms. Nikon has developed the Perfect Zoom System that provides high zoom ratios of 25:1, combined with high NA objectives. For both high and low magnifications, SMZ25 has a 25mm filed of view (FOV) allowing biologists to capture a 35 mm dish with uniform brightness.

Olympus develops stereo microscopes. An advanced model is the SZX16, suitable for oblique and brightfield illumination. Its maximum numerical aperture (NA) of 0.3 (objective 2x Plan Apo) enables seeing details in the sample. The zoom ratio of 16,4:1 combined with the long working distances (for the 2x it is at 20mm) enables switching between micro and macro view.

Table 11 lists the major manufacturers of electron microscopes among the articles. It is of interest to note that the developers of cryo-electron microscope (cryo-EM), Drs. Jacques Dubochet, Joachim Frank and Richard Henderson, were awarded Chemistry Noble Prize in 2017, and cryo-EM has been adopted in pharma research [97].

FEI T12 transmission electron microscope was used to investigate the structural composition of caveolar coat complex [27], the periciliary brush between the lung mucus layer and airway epithelia [100], and SNARE proteins [101]. FEI Tecnai G2 Spirit BioTWIN electron microscope was used to study lacteal junction [102].

The last years a new method has appeared in electorn microscopy for 3D imaging. It is the Serial Block Face Scanning Electron Microscopy (SBF-SEM) that allows obtaing serial block face images and 3D data using a SEM. It is consisted of a microtome equipped with a diamond knife inside the chamber of the SEM that cut s slices of <50nm of the sample in between imaging. It is an automated process allowing for large volume acquisitions. FEI is commercializing such a system, the VolumeScope SEM.

JEOl JEM-100CXII electron microscope was used to study the role of Dmxl2 in the infertility associated with a loss of GnRH neurons in mouse [103], and the mechanism by which CaMV perceives aphid vectors [104]. JEOL 1230 electron microscope was used to investigate the relationship between chromatin structure and gene expression [105], and JEM 1400 electron microscope was used to investigate the role of sFLT-1 in the maintenance of the avascular photoreceptor layer in mouse models [49].

Philips electron microscopes were used to investigate the structural composition of caveolar coat complex [27], the promotion of mucosal innate immunity [106], the relationship between pathogen colonization of the intestine and bacterial virulence [107], the existence of microDNAs in mammalian cells [108], the role of ENT3 in lysosome function and macrophage homeostasis [109].

ZEISS electron microscope was used to investigate the role of neurotransmitter-mediated exosome transfer in the interactions between neurons and oligodendrocytes [110], the mechanism of myelin membrane assembly [42], retinal specialization in Gnathonemus petersii [111].

Zeiss commercializes an SBF-SEM microscope, it is the Zeiss Sigma VP 3View system in a Gatan 3View SBF microscope housed in a Gemini SEM column.

Hitachi electron microscope was used to study spider silk formation [112], the mechanism of tip link regeneration in auditory hair cells [44], the evolution of lignin-degrading peroxidases in Agaricomycetes [113], and the roles of the gene dll in the evolution of sexually dimorphic traits of Rheumatobates rileyi [95].

Jost AP and Waters JC published an excellent article about how to prevent errors in using microscopes [114].

Dr. Orestis Faklaris updated this article and added several tables, and was added as an author in October 2019.