Monday, November 18, 2013

Record Keeping……..Never Too Much: A Lesson Learned in Laser Micro Dissection (LMD)

A major research activity that we perform for our scientists is Laser Micro Dissection (LMD). Over the years we have performed LMD on many of our donated tissue samples.  We have microdissected breast tissue and lymph node tissue including both normal and tumor cell types. A good part of this is performed on OCT embedded tissue.  The first step usually is to prepare H&E section(s) and review the section/area of interest with the pathologist.  Often times we draw lines on the coverslip to mark the areas of interest and use as a reference while performing LMD.  This slide will be saved in our file for future reference (see Figure 1).

The next step is cutting tissue sections with the laser microdissecting microscope. In our establishment, we use Leica AS LMD (Leica microsystems). Cut tissue sections are then mounted on PEN foil slides (W. Nuhsbaum Inc.), stained and LMD performed to cut out the areas of interest. 
 Many times, both cell types of interest are represented in the same OCT block (as shown in Figure 1 and 2 above) and often one of the cell types is much smaller compared to the other.  When genomic data for example, is generated from these cells, there could be potential questions raised about cell contamination. In order to understand if this was a possibility or not, an image of the LMD slide and written record of how LMD was performed (that is, both cell types were obtained from the same slide, or each cell type was obtained from independent slides during LMD) will provide clear answers about any potential contamination of cells during the downstream processing into nucleic acids (DNA/RNA). 
Record keeping is therefore very essential here and you can never have too much detail maintained during LMD processing.  We have always recorded how many sections we cut from each block and used for LMD, how the cells were captured within each slide and images of the LMD slide.  For example, if we are studying tumor and adjacent normal tissue from the same donor, we document if we captured both normal and tumor from separate slides or from the same slide.  However, even detailed notes cannot beat the usefulness of an image of the slide. It is beneficial to capture the H&E image of the slide and an image of the LMD slide before and after performing LMD.  If an imaging system is unavailable, another option would be to save the PEN foil slide and the original H&E slide.  Both of these options will tell you exactly what areas of the slide were laser microdissected. This exercise may seem tedious and you may need a lot of storage space in the months or years ahead after performing multiple LMD, but when you are faced with a research scientist who has questions regarding the tissue morphology, and who is trying to eliminate potential cell contamination in the interpretation of research results, you will be glad you kept all of these different pieces of information!

No comments:

Post a Comment