Fig 1: Plasma triggering receptor expressed on myeloid cells 2 (TREM2) levels can accurately discriminate patients with NAFLD activity score (NAS) ≥4 and subjects with NAS <4 in the derivation cohort. (A) Quantitative enzyme‐linked immunosorbent assay analysis of TREM2 levels in plasma extracted from patients with NAS ≥4 and subjects with NAS <4 categorized according to NAS (n = 48). (B) Diagnostic accuracy of plasma TREM2 levels. (C) Diagnostic accuracy of plasma TREM2, tissue inhibitor metalloproteinase 1 (TIMP1), tissue inhibitor metalloproteinase 2 (TIMP2), and matrix metalloproteinase 2 (MMP2) between patients with NAS ≥4 and subjects with NAS <4 (n = 28). (D) Correlation between hepatic TREM2 messenger RNA levels and plasma TREM2 levels in patients with biopsy‐proven NAFLD (n = 27).
Fig 2: Amine PEGylated (1 kDa) TIMP2 displays an increased serum half-life. (A) Standard curve comparison between TIMP2 and amine PEGylated TIMP2 (TIMP2-a-PEG(n)). (B) Comparison of calculated serum concentrations of TIMP2 and TIMP2-a-PEG(n) between 0 and 28 h. (C,D) Serum half-life of (C) TIMP2 and (D) TIMP2-a-PEG(n), calculated after peak concentrations at 30 min (TIMP2) and 2 h (TIMP2-a-PEG(n)).
Fig 3: Histidine labeling of TIMP2 using bis-sulfone PEG (10 kDa). (A) Reaction series with varying ratios of TIMP2-to-BS-PEG. Unlabeled (Unl.), TIMP2-H-PEG(1), TIMP2-H-PEG(2), and excess (Nx) PEGylated versions are highlighted. (B) Analysis of purified BS-PEG labeled TIMP2 by Coomassie stain, Barium Iodide (BaI2) stain, and reverse zymography. (C) Quantification of the reverse zymogram band intensity. (D) Immunoblot and (E) densitometry illustrating the stability of TIMP2 versus TIMP2-H-PEG(1) in human serum at 37 °C. (F) Circular dichroism of TIMP2 versus TIMP2-H-PEG(1). (G) Tryptic digestion of TIMP2-H-PEG(1). followed by immunoblot using an anti-6×-His antibody and Barium Iodide (BaI2) staining, to demonstrate that the PEG is present on the his-tag.
Fig 4: Bis-sulfone reaction scheme for chemical conjugation to histidines. (A) Bis-sulfones undergo elimination of a sulfinic acid to give monosulfone. (B) Target conjugation sequence for bis-sulfone linkage to histidines. (C) Proposed reaction steps involved in bis-sulfone conjugation to target histidines. (D) Proposed predominant products of PEG-bis-sulfone conjugation to TIMP2.
Fig 5: TIMP2-H-PEG(1) displays an increased serum half-life. (A) Standard curve comparison between TIMP2 and TIMP2-H-PEG(1). (B) Comparison of calculated serum concentrations of TIMP2 and TIMP2-H-PEG(1) between 0 and 28 h, and 0 and 96 h (TIMP2-H-PEG(1)*). (C) Serum half-life of TIMP2, TIMP2-H-PEG(1) (0–28 h), and TIMP2-H-PEG(1)* (0–96 h), calculated after peak concentrations at 30 min.
Supplier Page from Abcam for Human TIMP2 ELISA Kit