Modified clearing technique to identify lymph node metastases in post-irradiated surgical specimens from rectal...

The aim of this study is to describe a modification of the original method of clearing lymph nodes in specimens from rectal adenocarcinoma and its yield in comparison to a control group whose specimens were studied by a manual technique. There were 32 consecutive patients with locally advanced rectal adenocarcinoma. All patients received preoperative radiation therapy (PRT) at doses of 45 Gy, concomitant with 5-fluorouracil at the doses of 450 mg/m2 IV bolus, days 1-5 and 28-33 of PRT; 4-8 weeks later radical surgery with a curative intent was performed. Twenty two specimens were studied by a manual technique consisting of fixation in 10% formalin. Ten specimens were studied with the modified clearing technique consisting of administration through the inferior mesenteric artery of methylene blue; fixation with 10% formalin plus 0.01% of methylene blue; clearing the fat with a sequence of 95% alcohol-100% acetone-100% xylene. In the 32 specimens, 413 lymph nodes were found. Using the manual technique 104 lymph nodes were found (average 4.7 lymph nodes per specimen); 309 lymph nodes by the modified clearing technique (average 30.9 lymph nodes per specimen), (p < 0.0001). Using the manual technique 23 out of 104 lymph nodes (22%) contained metastases. All metastatic lymph nodes measured > 5 mm; 18 out of 23 metastatic lymph nodes were located on the perirectal area (level 2); and five were located in the root of the inferior mesenteric artery (level 5). Using the modified clearing technique 278 out of 309 (89%) lymph nodes measured < 5 mm; 15 out of 309 (4.8%) lymph nodes contained metastases; 9 out of 15 (60%) measured > 5 mm and were located on levels 2 and 5; and six, located on level 5, measured < 5 mm. The modified clearing technique is a quick, easy and reproducible method for identifying lymph nodes in post-irradiated surgical specimens. In comparison to the traditional manual method of searching for lymph nodes, it improves in the detection of small lymph nodes (< 5 mm), allowing better staging in patients with rectal adenocarcinoma.