How to improve your Alzheimer Protein Analysis with Laser Microdissection

Typically, special membrane-based slides are used for laser microdissection. Sample preparation for LMD is straightforward, and can be derived from classical preparations for histology.

For downstream proteomics we recommend PET membrane slides (which are almost free of softeners) or DIRECTOR™  slides which allow completely membrane-free ablation.

Paraffin embedding brings the tissue sample into a constitution which can be cut into thin slices. Alternatively you can utilize cryo-sectioning to prepare your slides.

We will deliver the suitable LMD slides for your application.

Amyloid plaques can be visualized by DAB staining based on a β-amyloid antibody. This results in a brown staining of amyloid plaques in brightfield microscopy (right). DAB can be also combined with cresyl violet staining.

An alternative is Congo red which stains amyloid on formalin-fixed, paraffin-embedded (FFPE) as well as frozen tissue sections.

β-amyloid antibodies can also be used for immunofluorescence microscopy which is also applicable with Leica LMD systems.

Leica LMD systems can generate an automatic sample overview that can be used to easily navigate to your regions of interest (ROI).

Your ROIs can be identified and marked automatically by the ADM software module, or you can apply shapes manually. 

• More information on the Leica LMD systems
• Laser microdissection articles & tutorials

Next, the defined ROIs are precisely dissected by the laser under visual control and directly collected via gravity.

Isolate disease affected material or any other ROI with a few mouse clicks and pool as much as you want in one or different reaction vessels if desired.

Also, you can cut your samples directly on the fly with the "Move and Cut" tool, without any predefined shapes.

The collection via gravity allows the use of standard, cost-effective consumables such as PCR tubes or 8-strip tubes.

Proteins can be extracted and prepared for downstream analysis from the dissectates with established protocols. Those might differ depending on the upstream sample embedding method, as well as on the aimed protein analysis technique.

Please find some examples here:

• Cryo samples and Western-Blotting (Del Turco et al.)
• Cryo samples and ESI-MS/MS (Cahill et al.)
• FFPE samples and LC-MS/MS (Drummond et al.)
• Cryo samples and LC-MS/MS (Hondius et al.)

The following techniques for protein analysis have been successfully tested with proteins gained from LMD:

• LC-MS/MS
• ESI-MS/MS
• SDS-PAGE
• Western Blotting
• 2D-DIGE
• MALDI-TOF
• SELDI-TOF
• NMR