Abstract
BackgroundWe aimed to develop a time‐resolved fluorescence immunoassay (TRFIA) for detecting soluble T‐cell immunoglobulin and mucin domain 3 (sTim3) in serum samples and to demonstrate a preliminary application of this method in membranous nephropathy (MN).MethodssTim3 TRFIA was developed, and the sTim3 concentration in the serum of patients with MN and healthy individuals was detected using a sandwich method.ResultsThe sensitivity of the developed sTim3 TRFIA was 0.66 ng/mL, higher than that of an enzyme‐linked immunosorbent assay (ELISA) (1.11 ng/mL). The detection range was 0.66‐40 ng/mL. The intra‐assay coefficient of variation (CV) for sTim3 was 1.64%‐4.68%, and the inter‐assay CV was 5.72%‐9.32%. The cross‐reactivity to interleukin 6 (IL‐6) and kidney injury molecule 1 (KIM‐1) was 0.25% and 0.04%, respectively. The average recovery was 105.26%. The sTim3 concentration in patients with MN was considerably higher than that in healthy individuals (P < .001). The sTim3 concentration in the serum of patients with MN was significantly increased from G1 to G4 based on the Jonckheere‐Terpstra test (P < .001). Thus, we used sTim3 as a diagnostic indicator for distinguishing between healthy individuals and patients with MN as well as between different stages of MN.ConclusionWe successfully established TRFIA to detect sTim3 in serum. We then applied this method to patients with MN, demonstrating for the first time that TRFIA is a valid diagnostic tool to detect sTim3 in serum.
Highlights
T-cell immunoglobulin and mucin domain 3 (Tim3) is a type 1 transmembrane receptor that can inhibit the activation of T helper 1 (Th1)
It was confirmed that another source of sTim[3] is spleen cells. sTim[3] mRNA is only found in spleen cells, and the activation of spleen cells leads to upregulation of soluble T-cell immunoglobulin and mucin domain 3 (sTim3).7 Xiao et al[8] found that sTim[3] and soluble programmed cell death protein 1 can enhance the cell-mediated immune response induced by the simian immunodeficiency virus (SIV) vaccine in mice
Based on the existing literature, this is the first study of time-resolved fluorescence immunoassay (TRFIA) used for the detection of sTim[3] and its application to test sera from patients with membranous nephropathy (MN)
Summary
T-cell immunoglobulin and mucin domain 3 (Tim3) is a type 1 transmembrane receptor that can inhibit the activation of T helper 1 (Th1)cells after ligand binding.[1]. Tim[3] has a soluble form (sTim3) and a membrane-bound form. High expression of membrane-bound Tim[3] was observed in common autoimmune diseases, like IgA nephropathy[9] and systemic lupus erythematosus. These studies demonstrate that Tim[3] is involved in pathogenesis; Tim[3] may be a potential therapeutic target for these diseases.[10]. We aimed to develop a time-resolved fluorescence immunoassay (TRFIA) for detecting soluble T-cell immunoglobulin and mucin domain 3 (sTim3) in serum samples and to demonstrate a preliminary application of this method in membranous nephropathy (MN). Methods: sTim[3] TRFIA was developed, and the sTim[3] concentration in the serum of patients with MN and healthy individuals was detected using a sandwich method. We applied this method to patients with MN, demonstrating for the first time that TRFIA is a valid diagnostic tool to detect sTim[3] in serum
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