Review of Laser Microdissection Systems

Laser microdissection systems allow you to dissect a tiny region of interest—such as a cell cluster, single cell, or nucleus—from sample tissue and isolate it for further study. Common uses for these miniscule dissectates include isolation of DNA, RNA, or protein. Research that depends on spatial information, such as the study of different cell types within tumors, can benefit from the ability to glean molecular profiles of specific, visually identifiable cells or regions of tissue. This article will cover some features to consider when choosing a laser microdissection system, and some expert advice on handling samples.

Dual lasers

Thermo Fisher Scientific plans a Q4 2021 release of the Arcturus Cellect Laser Capture Microdissection System, an update to the current Arcturus XT Laser Capture Microdissection Instrument, which contains both IR and UV lasers. “The gentle IR laser preserves tissue morphology and genetic content of captured and neighboring cells, allowing researchers to easily verify captured material while simultaneously preserving biomolecule integrity to enable research in diverse fields such as oncology and neurology,” says Troy Stearns, Product Manager for Arcturus Laser Capture Microdissection. The more powerful UV cutting laser is suitable for microdissection of dense tissue or large groups of cells. “The [IR and UV lasers] combine for the precision and power needed to drive improved downstream research analysis,” says Stearns.

Thermo Fisher Scientific provides detailed sample-preparation protocols for use with the Arcturus Cellect LCM System. Stearns notes that the dissection and embedding steps are critical for maintaining RNA quality when preparing LCM samples. “Researchers should use the entire adjacent section or dissect a large area of the section (>1000 cells) for RNA isolation,” he says, advising that it’s important to assess the quality of RNA isolated from the adjacent section.

“With a good amount of RNA, one can assess the RNA quantity and quality to make sure that high-quality RNA is obtained, even from a low number of LCM cells.”

Assessing RNA quality is also important for preserved FFPE samples, which may contain chemical modifications or RNA fragmentation that pose distinct challenges to gene-expression analysis, notes Stearns. “The initial paraffin blot preparation and storage can significantly affect the RNA quality,” he says. “Archival FFPE samples present additional challenges due to increased RNA degradation over time.”

A movable cutting laser

Some systems use a fixed laser for cutting tissue (along with a movable microscope stage). In contrast, Leica’s platforms use a movable cutting laser and a fixed stage. Leica Microsystems offers two systems for laser microdissection: the more basic LMD6, and the advanced LMD7, which has a more powerful laser, greater speed, and flexibility to accommodate more sample types. “The combination of laser movement via optics, and gravity collection directly into multi-well-plates, make our systems unique,” says Falk Schlaudraff, Manager Application Management Research Microscopy and Sample Preparation, Life Science division at Leica Microsystems. “Moving the laser for dissection offers high accuracy and speed, and is convenient for the human eye to watch the cutting process as the sample stays fixed.”

The Leica platforms collect samples using a contact-free gravity-based method, which can dispense samples directly into multi-well plates to facilitate subsequent high-throughput work. “Separation of individual cells or nuclei from heterogeneous tissue directly into 96-well plates, ready for downstream analysis, enables automated workflows for meaningful and reliable molecular biology such as spatial ‘omics approaches,” says Schlaudraff. “Dissection and collection of living cells is also very gentle with gravity collection, as the collection vessel can be pre-filled with culture media.”

Catapulting your sample

While gravity is efficient for moving samples, ZEISS’s laser microdissection pressure catapulting method provides another method for contact-free sample collection after laser microdissection. Nathan Lane, Life Sciences PASS Manager West for ZEISS Research Microscopy Solutions, notes that a distinguishing characteristic of the ZEISS PALM MicroBeam IV is that it enables non-contact pure and undamaged specimen collection against gravity, facilitating “more reliable sample extraction, increased purity, downstream RT-PCR analysis, and isolation of single cells.” According to Lane, another advantage to this technological approach of collection against gravity via the pressure catapult is that it improves collection efficiency and prevents smaller samples from being held back by surface tension when depending upon the force of gravity to make them drop. “The dissectate doesn’t always simply fall down, introducing limitations with respect to particles of microscopic diameters, single cells, and cell clusters,” he says.

Further advice

Depending on your particular experimental situation, you may prefer an inverted (such as ZEISS) or upright (such as Leica) microscopy set-up. But no matter the system chosen, it behooves the user to invest time in optimizing protocols for quality and reliable results. Chris Fillmore, Director of the Biorepository and Molecular Pathology Core and Assistant Professor of Pathology at the University of Utah, uses the ZEISS Palm MicroBeam to isolate tumor cells from a variety of cancer types, mainly for the purposes of NGS and single-cell applications. “We had to spend quite a lot of time optimizing conditions for the best sample capture, but the time is well-spent to produce optimal yields,” says Fillmore.

Attention to detail is especially important when it comes to sample preparation. With the traditional adage “Garbage in, garbage out,” Lane advises that careful sample preparation is crucial for quality results, whether RNA, DNA, or protein. “Preservation of the molecular content throughout the entire workflow is key—to put this simply, you risk losing content at every step,” he says. Similarly, Schlaudraff advises researchers to “protect your biomolecules of interest—investing time to establish a working protocol that allows you to identify microscopic targets and prevents degradation and alteration of biomolecules is key.” Conscientious sample handling, along with consideration of your experimental needs, can help you down the path toward successful laser microdissection.