The tumor tissue was fixed and embedded in paraffin. After that, the samples were cut into 3-µm-thick slices and stained with immunohistochemistry. The primary antibodies used in the study were as follows: anti-CCL11 (1:500; Abcam; ab203586), CD4 (1:200; Abcam; ab183685), and Foxp3 (1:2000; Abcam; ab75763). The primary antibody was incubated at 4℃ overnight. After washing with PBS, secondary antibody was added and incubated at room temperature for 1 h. With (DAB) color development, hematoxylin inhibits the nucleus. Finally, the staining quantity and intensity were comprehensively analyzed. The IHC results were evaluated using the intensity score. IHC scores were determined by multiplying the score for staining intensity (0, negative; 1, weak; 2, moderate; 3, strong) with the score or positive area (0, less than 5%; 1.5–25%; 2.26–50%; 3.51–75%; 4, greater than 75%). Scores of 0 to 7 were considered as a low expression and scores of 8–12 as a high expression. The scoring criteria referred to the following literature (Shan et al. 2013 (link); Tuo et al. 2022 (link)).
Free full text: Click here
