CellCut

The sample captured on the silicone membrane is ready for your downstream analysis. There are several ways to remove or process the sample from the silicone membrane depending on the throughput. For detailed guidance, please refer to the "Sample Lysis" protocols:

Sample Lysis Using Isolation Caps and CapStrips_Ice-Assisted Transfer 

Sample Lysis in EVO 96 Plate_Ice-Assisted Transfer 

Sample Lysis for MMI Isolation Caps_Sonication

To inspect and ensure you have captured your ROI, you can use the CellTools inspection function to move the sticky cap over a blank area or clean slide and bring it into focus. Using an objective of choice, you can visualise the cap when brought down into focus and locate all of your cut samples, giving yourself peace of mind. 

All common formats such as XML, JSON, CSV, and TIFF are supported for importing cell shapes. Additional formats can also be accommodated if needed. Our software team is happy to develop a dedicated interface to ensure the correct and seamless import of other file types. 

For precise alignment within the MMI CellCut system, three reference points are initially identified on the whole slide image. It is important to ensure to select distinct and recognisable features of the sample—such as specific cells, anatomical structures, or geometrically unique areas of the tissue—as reference points. After importing the shapes into the LMD system, the same three reference points must then be identified on the live image. This alignment process overlays the coordinates from the whole slide image onto those of the LMD system, enabling accurate positioning and seamless integration of the shapes. 

The MMI CellCut laser microdissection system is highly suitable for seamless integration into a Deep Visual Proteomics (DVP) workflow. Designed for maximum accuracy, this platform enables the contamination-free isolation of even the smallest single cells and subcellular compartments (such as nuclei) from tissue sections.

For high-throughput DVP workflows, the MMI CellCut HT laser microdissection system is particularly recommended. It features automated isolation into 96-well plates, allowing researchers to automatically and precisely deposit single cells or distinct cellular populations into individual wells.

The integration process includes the following key steps:

1. Cell Segmentation & Import: Following high-resolution imaging and AI-driven cell segmentation, the generated cell shapes and coordinates are directly imported and aligned within the MMI CellCut software.

    

2. Automated Laser Microdissection: The system automatically performs laser microdissection to cut and collect the targeted cells into the 96-well plate, ensuring high sample recovery and morphological integrity.

    

3. Downstream Processing: Once isolated via laser microdissection, the samples in the 96-well plate are ready for the subsequent DVP workflow steps, including protein extraction, enzymatic digestion, peptide purification, and ultimately, ultra-high-sensitivity mass spectrometry (LC-MS/MS) analysis. 

No. Overlapping of shapes is prevented by the predefined target positioning (PTP). PTP enables controlled collection of single cells or larger regions of interest on the sticky silicone membrane. This allows hundreds of samples to be placed next to each other without overlapping during laser microdissection. 

Isolated Leukocytes on a Single Cap

The laser used in the MMI CellCut laser microdissection system operates with low pulse energy and an extremely short pulse width of 500 picoseconds (ps). In contrast to conventional long-pulse lasers, significantly less laser energy is required for the cutting process, which minimises potential damage to the sample. Despite the lower energy per pulse, the high repetition rate delivers sufficient overall energy to efficiently cut even challenging tissues such as bone, making the process both precise and gentle during laser microdissection.

Laser Profiles of a Short-Pulse Laser and a Long-Pulse Laser

Detailed specifications of the MMI laser microdissection systems can be found here

Membrane slides are essential for laser microdissection because they act as a supportive carrier that is cut along with the sample, ensuring both structural integrity and successful isolation.

During the cutting process, target tissue sections or single cells are precisely excised using the gentle UV laser. Instead of cutting the sample directly on bare glass, the UV laser cuts through both the sample and the underlying inert membrane (such as PET, PEN, or PPS) of the membrane slide.

Despite featuring this specialised polymer layer, membrane slides are highly versatile; they can be handled like standard glass slides and are fully compatible with various staining methods, including classical histological stains and complex immunohistochemical (IHC) protocols. This makes them the ideal foundation for any high-precision laser microdissection workflow. 

Yes, the CellCut laser microdissection system is compatible with several inverted microscopes. In particular, it can be integrated with Evident IX73, IX83, and IX85, Nikon Ti2, and Zeiss Axio Observer microscopes. Other inverted microscope models may also be compatible. Please feel free to contact us to discuss your specific microscope and integration options. 

With the MMI CellCut, the maximum sample thickness you can successfully cut and isolate depends on the material, but the system is highly adaptable. While standard tissue sections are typically much thinner, the MMI CellCut can easily process and isolate thicker specimens, including FFPE tissue sections and other specialised materials with thicknesses over 100 µm.

To achieve clean cuts in thicker or denser samples, the platform offers two advanced features:

1. Adjustable Laser Power: Users can easily increase laser intensity within the software to cut through denser materials without damaging the surrounding tissue. Alternatively, the system can be equipped with a high-power UV Laser, providing the extra cutting strength needed for particularly thick or dense specimens.

    

2. Z-Drill Functionality: This feature allows CellCut to perform laser microdissection by cutting the same shape multiple times at consecutively adjusted Z-levels (focus planes). By stepping through the sample vertically, the laser can cleanly slice through thick specimens layer by layer. 

Laser Microdissection is defined as the precise cutting of microscopic tissue area using a laser and the subsequent separation of the dissected area from surrounding tissue. Importantly, laser dissection microscopes differ in the particular method to isolate target cells after cutting. MMI, for example, isolates cut tissue by employing adhesive isolation caps, which gently and reliably take up the sample of any size and shape.  

Typically, laser microdissection systems are based on research microscopes. Lasers are required to precisely cut tissue on this microscopic scale, and they are coupled into the optical paths of the dissection microscopes. For cutting, the sample is mounted on membrane slides to enable both for precise cutting and for subsequent cell isolation procedures.

The laser microdissection microscopes on the market today mainly differ in the laser types they use and how the laser is applied to cut the sample. The MMI CellCut uses a low-damage laser with low power but high pulse frequency to allow for efficient and precise cutting and to not compromise tissue integrity. Moreover, the laser is fixed during the cutting process to ensure that the laser is always in focus and the sample is efficiently cut.

Importantly, the laser dissection microscopes differ in the particular method to isolate target cells after cutting. MMI for example isolates cut tissue by employing adhesive isolation caps, multi-well plates and other formats, which gently and reliably take up the sample of any size and shape 

The laser microdissection technology has improved over the last year to make the cutting process more reliable and more precise. Thus, also single and rare cells can be excised from tissue sections, living cells, or smears and swabs.

Interestingly, the MMI CellCut microdissection microscope is highly optimised to specifically cut single cells. The low-damage laser allows to cut single target cells without impairing their DNA or RNA quality, or the integrity of living cells. To be able to cut single living cells, MMI also offers a range of dedicated consumables. Thus, physiological conditions and contamination-free handling is supported throughout the experiment.

Intriguingly, the patented adhesive cap technology used by the MMI CellCut allows to keep the tissue in position during cutting. Thus, excised tissue cannot drop off because of airflow or static forces, a phenomenon that often occurs with dry sample of very small sizes such as single cells. Moreover, the different objectives and fluorescence filters implemented in the laser dissection microscope can be combined and optimally employed to identify and select single target cells. Image analysis tools integrated into the MMI CellTools software package allows finding even very rare cells in the sample.