Esophageal Adenocarcinoma Methods and Protocols

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●● Press the continue button in the program and repeat the pro-

cedures above.

●● The procedure will be repeated until all the spots designed by

the template are made.

●● Close the program.

●● At the end, the program will generate an Excel report.

Automatic TMA is similar to the semi-automatic method. The

process of punching the holes in “recipient” block and “donor”

blocks as well as transferring the tissue to the recipient blocks is

totally automatic.

1. After the completion of insertion of samples into the “recipi-
   ent” blocks, put the block on a glass slide (facing down) and
   heat the “recipient” block in an oven at 42 °C for 40 min.
   After baking, place the block with the glass slide on a cool-
   ing plate for 10 min. Repeat the process of heating and cooling
   between 3 and 5 times. As a result, the tissue is strongly
   adhered to the surrounding paraffin.


2. Place the block on a cold plate to solidify. Figure 3 shows
   the TMA of a population of patients with esophageal
   adenocarcinoma.


3. Cut the TMA block into thin tissue sections by microtome.
   The best thickness for use is normally 4 μm. The microtome
   system consists of a blade carrier with combined transfer
   bridge, a control unit, and heated water bath. From the blade,
   the sections were cut. These sections are transferred to the
   water bath via a transfer bond.


4. The sections should be removed from the water bath quickly
   after they have expanded out to avoid melting of sensitive tis-
   sue spots (e.g., lipid-rich tissues like breast and brain). It needs
   to transport the section from the blade to a water bath manu-
   ally by using, e.g., a brush.
   Place the section on a superfast glass slide. The time the
   sections can be left in the water depends on the type of paraffin
   waxed used and the water temperature, and we suggest a tem-
   perature of 37–39 °C for the water bath.


5. For the best preparation (avoid detachment of the sections),
   put the slides in a slide holder for drying at room temperature
   overnight. The slides are then baked at 37 °C for 12–24 h.

After the cutting of the TMA by microtome, stain with hematoxy-

lin and eosin to make sure that the samples have representative

tissues (Fig. 4 ). Then, the TMA blocks could be cut for testing the

markers in the research works. Lastly, scan the hematoxylin and

eosin stained as well as other stained sections of the TMA into digi-

tal images for proper assessment.

3.3 Fully Automated
Method for Making
TMA

3.4 Section TMAs
by Microtome

3.5 Staining
and Processing

Nassim Saremi and Alfred K. Lam