Editorial Article
Monday June 15, 2009
by Caitlin Smith
Whether you are a professional microscopist or once did an imaging rotation in graduate school—and then resolved never again—your ability to see what’s really happening inside cells is improving at an exciting pace. Experienced microscopists are enthusiastic about the changes being made and the new developments on the horizon. Today’s imaging
systems allow you to look at your samples relatively easily, with a minimum of sample disruption, and a maximum of speed, sensitivity, and resolution.
Minimizing sample disruption
You know what it looks like when your cells are happy. So why take them out of their cozy incubator home to collect data from them? Depending on what kind of microscope you require, Olympus’s new VivaView™ Incubator Fluorescence Microscope may make data collection easier on your cells, giving you better results in the process. VivaView is essentially a microscope contained entirely within an incubator. “Because the system is based on a true incubator, physical stress on the cultures is minimized, so cells enjoy a consistently beneficial environment throughout the experimental process,” says Stuart Shand, associate product manager in the life science group at Olympus America. “Imaging is performed within the incubator, handling up to eight vessels at a time on its precision motorized stage, and allowing maximum compliance with a wide variety of experimental protocols. The researcher has the flexibility to add agonists or inhibitors with minimal disruption to the environment.”
Nikon’s Ti-E Inverted Microscope System with Perfect Focus System (PFS) also has features to keep your cells alive longer and minimize disruptions while performing complex techniques. It is “a powerful live cell imaging system that is essential for top performance in techniques like TIRF, confocal, FRET, photoactivation, and microinjection applications that take advantage of Nikon’s revolutionary CFI60 optics system,” says Stanley Schwartz, vice president for microscopy in the product/marketing division at Nikon Instruments. “Nikon’s PFS, the second place winner of The Scientist’s Top Innovations of 2008 Award, is designed to combat thermal and mechanical focus drift and keep cells alive and in focus during long-term, live-cell experiments.” Schwartz says that the PFS is better than most autofocus systems because it does not photobleach the specimen. “It also has the highest temporal sampling rate at 200 times per second, providing instantaneous perfect focus response,” he says.
Horiba Scientific offers the Raman microscope “as a paradigm-shifting innovation in microscopy that provides molecular bonding information with better than 0.4 um spatial resolution,” according to Fran Adar, worldwide Raman applications manager at HORIBA Jobin Yvon. “Careful attention to engineering specifications has enabled us to produce instruments optimized for spectral resolution (for research) or for automated, ease-of-use (for the analyst). Seeing how the instrumentation has become smaller and easier to use makes pursuing what used to be challenging problems now fun.”
Horiba’s LabRAM line of confocal Raman microscopes includes the ARAMIS for automation, the HR and HR UV-VIS-NIR for high resolution imaging, the INV for inverted fluorescence microscopy, and the IR2 for confocal and FTIR. A confocal Raman microscope incorporates not only a microscope and a Raman spectrometer, but also a spatial filter that controls the volume of sample analyzed down to microns. “What is truly unique about the Raman microscope is its potential to probe cellular organelles under functioning physiological conditions,” says Adar. “Electron micrographs of mitochondria, the Golgi apparatus, and the endoplasmic reticulum are beautiful. But these bodies are dead in the EM. It will be great to follow their chemistry while they are performing their respective functions.”
Creating more microscopists
Adar says that Horiba is “seeing people without any Raman background come to us with a genuine interest in the technology, because they have heard what it can offer, and that it is accessible to the novice.” Indeed, imaging systems that require no expert knowledge but can easily generate reliable high-quality data are attracting attention. According to Schwartz, the future of imaging will include a “shift to basic microscope systems being incorporated into simple-to-use boxed instruments that will be more compact, and less expensive to own and maintain.”
The Olympus FSX100 is ideal for scientists who cannot dedicate all their time (or lab space) to microscopy. This all-in-one system is a fluorescence and brightfield microscope with a small footprint. It is completely contained within its own darkroom, saving researchers the necessity of finding darkroom space or disrupting other lab work (it can be used with the room lights on). “The first thing you'll notice is that the FSX100 doesn't look like a microscope, and doesn't have eyepieces—the system is completely controlled through the computer, and the user follows the intuitive workflow of the software to compose and capture an image,” says Mark Clymer, product manager in the scientific equipment group at Olympus America. “But don't let its appearance fool you. The FSX100 is a sophisticated fluorescence and brightfield microscope; Olympus has just made it easier to use for everyone.”
Clymer notes that the FSX100’s ease-of-use appeals to researchers who wouldn’t normally use microscopy at all. “Traditionally, researchers may rely on resident experts for microscopy, perhaps through a core imaging facility,” says Clymer. “Now, they can capture their own images in minutes, without scheduling appointments with the expert. Even core imaging facilities must balance basic or routine imaging needs of some researchers with the advanced techniques demanded by others. With the FSX100, they can focus on the challenges of advanced imaging, and the FSX100 can handle the routine. The concept is very enabling.”
Motic Instruments recently launched its new BA series of microscopes, designed to make microscopy more affordable for classrooms, labs, or clinical settings where quality cannot be sacrificed. Their BA400, for example, is an expandable upright microscope with optional epi-fluorescence illumination that is rugged but built for comfort over long hours of use.
Closing in on the “Holy Grail”?
Many say that the ever-sought triad in microscopy is having the desired resolution, sensitivity, and speed—simultaneously. “Nikon has done just this by two or three fold over the last two years by introducing the newest confocal microscopy system that actually improves in all three areas of microscopy without sacrificing in other areas,” says Schwartz. Their A1R series of laser point scanning confocal microscopes include Resonant Scanning, which improves speed, and the Virtual Adaptable Aperture System, which improves sensitivity. “That leaves resolution, and that is what the exclusive CFI60 optical system by Nikon provides, together with the highest NA 1.49 and most aberration-corrected 60X and 100X objectives available today,” says Schwartz.
Leica Microsystems’ new TCS SP5 X Supercontinuum Confocal Microscope also offers improvements. Named one of the most innovative life science products of 2008 by The Scientist, the TCS SP5 X uses a white laser that lets you use wavelengths between 470 and 670 nm. This gives researchers the ability to use fluorophores that were previously inaccessible, as well as future fluorophores not yet invented. Multi-color experiments with up to eight simultaneous excitation lines are possible.
Imaging challenges for the future, according to Adar, will include sampling and information extraction. “The time that it takes to record the signal is now dictated by the sample, not the condition of the equipment,” says Adar. “Using multivariate analysis techniques to extract relevant information, we will all have to prove to ourselves and others that the conclusions that we draw from the data are accurate and robust.”