This method helps determine whether tumor cell invaded beyond elastic lamina in pT3 gastric cancer. elastic lamina erosion in the prognostic picture for pT3 gastric cancer. The main advantage of this technique is that it's convenient, and accurate approach to identify the elastic lamina, which makes it more precise for pathologist to evaluate whether tumor cells invade beyond elastic lamina in pT3 gastric cancer.
This technique can be towards a pathologic test stage diagnosis of lung cancer and cancer. This method could also be applied to the TNM stage system of lung cancer and colo-rectal cancer. First, immerse the slides in xylene in a Coplin jar for 10 minutes.
Remove the slides from this jar and immerse them in fresh xylene in a different Coplin jar for another 10 minutes. Immerse the slides in 100%ethanol for five minutes, and then transfer them to fresh 100%ethanol for another five minutes. Next, immerse the slides in 95%ethanol for two minutes.
Transfer the slides to a jar containing 70%ethanol for two minutes. After this, rinse the slides briefly with distilled water in a new Coplin jar. First, use tissue paper to wipe off any excess solution on the slide or around the tissue.
Place the slides in conditions that match room temperature and humidity. Add a few drops of potassium permanganate to each slide to oxidize them, making sure that the volume added is enough to cover the tissue section on each slide. After five minutes, rinse the oxidized slides with distilled water in Coplin jars, repeating the rinse two to three times and using a new jar of water for each rinse.
The tissue should have an observable brown color. Now, add a few drops of oxalic acid to each slide to bleach them, making sure that the volume added is enough to cover the tissue section on each slide. After five minute, rinse the bleached slides with distilled water, repeating the rinse two to three times and using a new jar of water for each rinse.
Wash the slides briefly in 95%alcohol. Immerse the washed slides in elastin solution for eight to 24 hours. Then immerse the slides directly in 95%ethanol for one to two minutes to differentiate each tissue section well.
Rinse the slides completely with distilled water. Check microscopically for the blue-black elastic fibers staining and the gray background. Counterstain the slides in a van Gieson solution for one minute, making sure that the volume used is enough to cover the tissue section on each slide completely.
Next, rapidly drop 95%ethanol onto the tissue slices for a few seconds to differentiate each tissue section well. To begin, immerse the slides in 95%alcohol for five minutes to dehydrate them quickly. Transfer the slides to 100%alcohol for five minutes to continue the dehydration.
Then, transfer the slides to fresh 100%alcohol for five additional minutes. Immerse the slides in two different changes of xylene with each immersion lasting for three minutes. After this, add one drop of resinous mounting medium to the slide and place a cover slip on top.
Using a microscope, observe the tissue sections as outlined in the text protocol. In this study, elastic staining is utilized to visually identify subserosal elastic lamina. A representative successful staining reveals that after elastin solution staining, but before van Gieson's counterstaining, the elastic lamina can be clearly seen in the form of filaments that are stained blue-black.
They are close to the serosal mesothelial cells, and a certain amount of distance is still seen between the elastic lamina and the tumor cells. Typical elastic stainings with van Gieson's counterstain demonstrate that the elastic lamina is stained blue-black. The collagen fibers are stained red, and the muscle fibers and cancer cells are stained yellow.
Cancer cells penetrating the elastic lamina is considered elastic lamina invasion. Whereas cancer cells being close to but not invading the elastic lamina is considered elastic lamina non-invasion. If the staining reveals a repetitive structure indicative of multiple layers of elastic lamina, and the tumor does not invade beyond the outermost layer, the sample should be diagnosed as elastic lamina non-invasion.
Elastin solution is a and the staining time is relative long, so it's best to immerse all slides with the elastin staining in a sealed container. After elastin solution staining, should be washed away with distilled water.
