Journal: Blood Vessels, Thrombosis & Hemostasis

Article Title: A second generation of C3 humanized rats for preclinical evaluation of human C3 inhibitors

doi: 10.1016/j.bvth.2026.100138

Figure Lengend Snippet: The second-generation hC3 rats express human, but not rat C3 in their plasma and tissues, at levels higher than those in the first-generation hC3 rats. (A) Plasma from both human and rat samples were collected, and the presence of human (hC3) and rat C3 (rC3) were evaluated by western blots. Purified human C3 (hC3), rat C3 (rC3), human C3 depleted serum (hC3-dpl), and C3 knockout rat serum rat plasma (rC3-KO) were used as controls. Arrows pointed to the human or rat C3 band. (B) Plasma hC3 levels in the second-generation hC3 rats (both male and female) were evaluated using a human C3 ELISA kit (Hycult) and compared to the levels in the first-generation hC3 rat plasma. (C) Total RNA was extracted from the liver, retina, spleen, kidney, and brain of WT, second-generation, and first-generation hC3 rats, and then semiquantitative reverse transcription PCR was used to detect and quantitate levels of human C3 (hC3) transcripts using the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as internal controls. 1st gen, first generation; 2nd gen, second generation.

Article Snippet: Blood levels of human C3 protein were measured in the identified human C3 Tg rats using an enzyme-linked immunosorbent assay (ELISA) kit (Hycult, catalog no. HK366) to identify the founder rats with close-to-physiological expression levels of human C3.

Techniques: Clinical Proteomics, Western Blot, Purification, Knock-Out, Enzyme-linked Immunosorbent Assay, Reverse Transcription

Journal: Nucleic Acids Research

Article Title: Deep learning-guided dual-fitness evolution of T7 RNA polymerase for enhanced stability and activity

doi: 10.1093/nar/gkag259

Figure Lengend Snippet: Comparison between WT and mutant T7 RNAP. ( a ) Selection of the improved dual-fitness mutant. Comparison of thermal unfolding curves of the WT and M1 determined by ( b ) far-UV CD spectroscopy and ( c ) DSC. Effects of temperature on activity of M1 under ~100 bp ( d ) and ~1000 bp ( e ) templates. ( f ) The dependence of dsRNA impurities for M1 on temperature. ( g ) The RNA yield (bar) and dsRNA impurities (scatter) under the ~1000 bp template for the WT and mutant. ( h ) The kinetic stability of the proteins is determined by monitoring residual enzymatic activity after incubation for 15 min at various temperatures. The \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{upgreek} \usepackage{mathrsfs} \setlength{\oddsidemargin}{-69pt} \begin{document} $T_{50}^{15}$\end{document} values for the WT, M8, M10, and M11 are 42.1, 51.1, 52.4, and 52.2°C, respectively. ( i ) RNA yield (bar) and dsRNA content (scatter) of the WT, M10, and M11 under different length templates. ( j ) Diagram illustrating the process from transcription to preparation of mRNA–LNPs, followed by cell transfection and characterization of expression efficiency. ( k ) The expression efficiency in HEK293T cells of mRNA encoding luciferase transcribed by the WT at 37°C and M10 at 50°C. The relative activity shown in (a) and (d) is based on the WT at 37°C. The residual activity shown in (h) is based on the maximum activity of each protein.

Article Snippet: The Cap1 capping kit and dsRNA ELISA (enzyme-linked immunosorbent assay) kit were acquired from Hzymes Biotech.

Techniques: Comparison, Mutagenesis, Selection, Circular Dichroism, Activity Assay, Incubation, Transfection, Expressing, Luciferase

Journal: Science Advances

Article Title: Inflammatory arthritis irAE may represent a unique autoimmune disease primarily driven by T cells but likely not autoantibodies

doi: 10.1126/sciadv.aea4262

Figure Lengend Snippet: ( A ) Expression of CD27 and IgD on CD19 + B cells. Right: Percentage of IgD + CD27 − naïve B cells and IgD − CD27 − DN B cells. ( B ) Expression of CD27 and CD38 on CD19 + B cells. Right: Percentages of CD27 hi CD38 hi ASCs and CD27 + CD38 − memory B cells. ( C ) Expression of CD138 on CD27 hi CD38 hi ASCs. Right: Percentage of CD138 + ASCs and CD138 − ASCs. ( D ) Expression of CD11c and CD21 on CD19 + B cells. Right: Percentage of CD11c + CD21 − CD19 + B cells. [(A) to (D)] HC ( n = 63), irAE ( n = 33), RAC ( n = 46), and ICI ( n = 20). ( E to G ) GSEA was performed on the B cells between irAE and ICI. (E) Significantly enriched pathways in B cells from irAE and ICI. GSEA plots of IFN-α and IFN-γ response (F), and oxidative phosphorylation (G). ( H ) The volcano plots of the citrullinated or noncitrullinated relative IgG or IgM isotype autoantigen levels comparing RA versus HC, irAE versus ICI, irAE versus HC, or irAE versus RA. The autoantigens were labeled when P < 0.01. TNF-α reactivities resulted from the administration of anti–TNF-α therapy for the treatment of RA. ( I ) Immunoglobulin isotype levels in the plasma were measured by multiplex assay. HC ( n = 22), irAE ( n = 34), RAC ( n = 46), and ICI ( n = 26). ( J ) CXCL13 levels in the plasma were measured by enzyme-linked immunosorbent assay (ELISA). HC ( n = 20), irAE ( n = 34) RAC ( n = 47), and ICI ( n = 23). ( K ) B-cell activating factor (BAFF) levels in the plasma were measured by multiplex assay. HC ( n = 21), irAE ( n = 34), RAC ( n = 47), and ICI ( n = 18). Data in graphs represent mean ± SEM. Significance was tested by one-way ANOVA [(A) to (D) and (I) to (K)] and logistic regression (H). [(A) to (D) and (F) to (K)] ICI, ICI control.

Article Snippet: For CXCL13, IL-21, and CX3CL1 measurements, the following kits were used: Human CXCL13/BLC/BCA-1 Quantikine Enzyme-Linked Immunosorbent Assay (ELISA) (R&D Systems, catalog no. DCX130), Human IL-21 DuoSet ELISA (R&D Systems, catalog no. DY8879-05), and Human CX3CL1/Fractalkine DuoSet ELISA (R&D Systems, DY365); all steps were performed according to the manufacturer’s instructions.

Techniques: Expressing, Phospho-proteomics, Immunopeptidomics, Labeling, Clinical Proteomics, Multiplex Assay, Enzyme-linked Immunosorbent Assay, Control