Basic StudiesIdentification of Different Form Tim-3 Proteins by a Unique Set of Tim-3 Monoclonal AntibodiesWang, Zhuocai*; Yan, Guangning*; Cui, Wenzhi*; Gao, Feng†; Chen, Jing†; Luo, Luqiao‡; Zhang, Minghui‡; Li, Zhi‡Author Information *Department of Pathology, Liuhuaqiao Hospital, Guangzhou ‡Department of Pathology, Guangdong Provincial People’s Hospital, Guangdong Academy of Medical Sciences, Guangzhou, Guangdong Province †Department of Pathology, Jingjiang People’s Hospital, Jingjiang, Jiangsu Province, China Reprints: Zhi Li, Department of Pathology, Guangdong Provincial People’s Hospital, Guangdong Academy of Medical Sciences, 106, Zhongshan Road II, Guangzhou 510080, Guangdong Province, China (e-mail: email@example.com). Received June 28, 2019 Accepted September 23, 2019 Online date: October 22, 2019 Journal of Immunotherapy: February/March 2020 - Volume 43 - Issue 2 - p 43-47 doi: 10.1097/CJI.0000000000000303 Buy Metrics Abstract T-cell immunoglobulin and mucin domain-3 (Tim-3) has been suggested to be a critical immune checkpoint target for cancer immunotherapy. However, limited progress with Tim-3 immunotherapy has been achieved over the last decade due to the lack of specific Tim-3 monoclonal antibodies. In this study, we have successfully developed a unique set of Tim-3 antibodies that are able to detect different molecular weights (by Western blot mobility) of Tim-3 proteins ectopically expressed in the same CHO cells. Some of the antibody clones detect only 33 or 55 kDa bands, the rest can recognize both 33 and 55 kDa bands on polyacrylamide gel electrophoresis gel. Antibody clones with 55 kDa specificity uniquely bind to the membrane form of Tim-3 on macrophage, which colocalizes with the CD68, and could be used as a specific marker for tumor-associated macrophage, whereas other clones showed cytoplasmic staining in tumor cells. The membrane form of Tim-3 on tumor-associated macrophages may bear significant roles for clinical application of Tim-3, but less likely for cytoplasmic one. The availability of this unique set of antibodies will be critical for an ultimate understanding of Tim-3 function in tumor microenvironment and potential clinical applications. Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.