The light microscope is an invaluable tool for cell biology and it has been improved over time to meet different optical imaging demands.1
For example, phase contrast microscopy has been developed to image a low-contrast objects, such as live cells which can not be seen clearly using a bright-field microscope. The phase contrast microscopy uses an annular phase ring that transforms small phase differences to amplitude differences which enables live cell imaging. Polarization microscopy utilizes polarized light to observe ordered molecules such as spindle fibers and actin filament bundles in living cells. Differential interference contrast (DIC) microscopy is yet another technique that uses dual-beam interference optics for applications demanding high resolution and contrast for inspection of cultured cells.1
Few efforts have focused on scaling down the size of a light microscope, but because of the development of cell-based lab-on-a-chip applications, this is now an important goal. In biology, many cellular processes, including embryonic development, wound healing and disease progression occur over extended periods of time. Hence, it is beneficial to continuously monitor the state of cells during these processes. The current practice for monitoring of cells in culture requires users to remove the cells from the incubator and inspect them under a microscope. This process not only introduces an added disturbance to cells in culture, but it is also prone to microbial contamination during cell handling. Furthermore, for many lab-on-a-chip applications, this requires detachment of the system from syringe pumps and exposure to numerous subsequent deleterious effects, such as the introduction of bubbles.2
One possible remedy for observation of cells during culture is to build an incubator around a conventional microscope, an approach which results in bulky and expensive systems. The availability of a miniaturized microscope that can be placed inside an incubator will alleviate most of these problems.
Recently, with the availability of low-cost, compact and high-performance image sensors, such as charge coupled devices (CCDs) and complementary metal oxide semiconductors (CMOS), numerous compact lens-free imaging systems have been reported. In a lens-free imaging system, the diffracted or holographic image of objects are recorded directly onto an image sensor and the real image of the object is usually reconstructed numerically.3–8
Since lens-free imaging systems do not require any bulky lens systems, they are cost-effective, light-weight and portable. Due to these attractive features, lens-free imaging technology is finding novel applications in biomedical sciences. For example, by using a lens-free imaging system, an ultra wide-field cell detection system has been developed by analyzing the diffraction signature of different cells.5,6,9
Also, a lens-free cardiotoxicity screening system was recently reported to monitor the beating rates and beat-to-beat variations of cardiomyocytes induced by different drugs in real-time.10
Furthermore, by using a holographic technique with an improved numerical algorithm, a lens-free imaging system with sub-pixel resolution was introduced,4,7,11,12
and a lens-free optical tomographic microscope has been developed for three-dimensional (3D) microscopy applications.13,14
Despite its compact size, a lens-free imaging system has drawbacks for in situ
cell monitoring because the object has to be placed close to the image sensor (~100 μm), which is less than the thickness (~2 mm) of the most common cell culture substrates (e.g.
, flasks, multi-well plates and Petri dishes). This problem can be resolved using a multi-angle illumination scheme, however, the distance between the object and the image sensor has to be less than 1.1 mm.15
Recently, an ePetri dish has been reported, which uses an image sensor as the cell culture substrate.16
The ePetri dish has been applied for long-term observation of cellular behavior, but it still needs numerical reconstruction to obtain images of cells.
Several miniaturized microscopes have been commercialized. Examples include IncuCyte™ (Essen BioScience, Inc.) and LumaScope (Etaluma, Inc.). These microscopes can image cellular behavior inside an incubator but are still expensive and hard to construct by the user. In this study, we used a different approach to develop an inexpensive microscope for in situ live cell monitoring by modifying a commercial webcam. This mini-microscope is constructed by combining a compact CMOS imaging sensor with a simple lens that enables objects to be observed without numerical reconstruction. The developed mini-microscope is easy-to-fabricate, highly cost-effective and compact. Due to its compactness, the mini-microscope can be easily placed and used inside a conventional incubator. To demonstrate the versatility of the mini-microscope, we used this instrument to carry out long-term observations of mouse 3T3 fibroblast cells that were undergoing migration in a scratch assay; we also successfully monitored and recorded the formation of mouse embryonic stem cell (ESC) aggregates in vitro. In addition, we observed the generation of droplets in a microfluidic device in real-time using the mini-microscope.