Development and characterization of a glyceraldehyde-3-phosphate dehydrogenase(GAPDH)monoclonal antibody for human health assessment

我々は解糖系に密接に関わるGAPDH酵素に対する新規のモノクローナル抗体(mNI-7)を作製した。この抗体は細胞質内に存在する約38kDaの分子を認識する抗体で、プログラム死(アポトーシス)を起こした細胞に反応した。このことは、この抗体によって認識されるGAPDHがアポトーシスを起こした細胞を検出するためのモニタリングマーカーの一つとして利用できることを意味している。そこで、我々は健康人(125人)と慢性関節リウマチ患者(50人)の血清中のGAPDHレベルをmNI-37抗体を用いた簡易ラテックス凝集反応で検査を行なったところ、慢性関節リウマチ患者の血清中に有意にGAPDHが存在していることがわかった(P<0.001)。mNI-37抗体は疾病や健康を診断評価するための非常に有益な抗体であることが明らかになった。A unique monoclonal antibody (mAb), designated mNI-37 (mouse IgM), was generated. This mAb recognized a protein with a molecular weight of approximately 38kDa which exactly corresponded to glyceraldehyde-3-phosphate dehydrogenase (GAPDH), as assessed by enzyme-linked immunoassay (EIA) and immunoblotting analyses. The GAPDH antigen defined by mNI-37 was not expressed on the cell surfaces of various lymphoid cells or cultured cell lines. However, this mAb was found to strongly react with cultured cell lines treated with 70%-ethanol, or cells undergoing apoptosis induced by treatment with mitomycin-C (MMC) or CH-11 (anti CD95,anti-Fas) mAb, suggesting that the GAPDH antigen defined by mNI-37 is abundant in the cytoplasm. In addition, the reactive pattern of mNI-37 was very similar to that of the apoptosis-related (apoptosis cell marker) mAb, APO2.7. On the other hand, it is noteworthy that the level of the GAPDH molecule detected by mNI-37 was markedly elevated in sera from patients with rheumatoid arthritis (RA) as compared with the sera from normal healthy donors (P < 0.001), suggesting that GAPDH is an excellent marker of cell destruction resulting, e.g. apoptotic responses such as autoimmune-mediated cell killing