Journal: Genes & Diseases

Article Title: Dual-mode aptamer-driven biosensing platform for ultrasensitive and mutation-resilient detection of the SARS-CoV-2 nucleocapsid protein

doi: 10.1016/j.gendis.2025.101943

Figure Lengend Snippet: Workflow of NP14 aptamer screening and development of the MD ELAAA detection platform. (A) Schematic illustration of the X-aptamer protein SELEX process for isolating aptamers. (B) Schematic illustration of the ultrasensitive detection of the SARS-CoV-2 N protein via the MD ELAAA platform.

Article Snippet: X-Aptamer libraries were acquired from AM Biotechnologies (Houston, Texas, USA); His-Tag magnetic beads (Invitrogen, DynabeadsTM His-Tag Isolation & Pulldown, 10103D), SARS-CoV-2 N protein, and anti-SARS-CoV-2 N protein monoclonal antibodies (anti-SARS-CoV-2 N protein mAb, Cat: 40143-MM05, 40588-R001) were purchased from Sino Biological.

Techniques:

Journal: Genes & Diseases

Article Title: Dual-mode aptamer-driven biosensing platform for ultrasensitive and mutation-resilient detection of the SARS-CoV-2 nucleocapsid protein

doi: 10.1016/j.gendis.2025.101943

Figure Lengend Snippet: Binding affinity and stability characterization of the NP14 aptamer. (A) Magnetic bead (12.5 mg/mL, 3 μL) flow assay for the binding of the aptamer to the His-tag SARS-CoV-2 N protein (1 μg). (B) Flow cytometry analysis of the binding of 300 nM FAM-labeled aptamer NP14 to magnetic beads coated with the SARS-CoV-2 N protein. (C) Flow cytometry analysis of the binding of 300 nM FAM-labeled NP14 to magnetic beads coated with the SARS-CoV-2 N protein at different temperatures (4 °C, 25 °C, and 37 °C). (D) The binding affinity of NP14 for the SARS-CoV-2 N protein was validated via the use of 2 μg/mL SARS-CoV-2 N protein and biotin-labeled NP14 at different concentrations (0, 2.5, 5, 10, 20, 50, 100, 150, and 200 nM). (E) Determination of the Kd value of aptamer NP14 (15.625, 31.25, 62.5, 125, 250, and 500 nM) via surface plasmon resonance. (F) Confocal analysis of 300 nM FAM-labeled aptamer NP14 with SARS-CoV-2 N protein-coated magnetic beads (scale bar = 30 μm).

Article Snippet: X-Aptamer libraries were acquired from AM Biotechnologies (Houston, Texas, USA); His-Tag magnetic beads (Invitrogen, DynabeadsTM His-Tag Isolation & Pulldown, 10103D), SARS-CoV-2 N protein, and anti-SARS-CoV-2 N protein monoclonal antibodies (anti-SARS-CoV-2 N protein mAb, Cat: 40143-MM05, 40588-R001) were purchased from Sino Biological.

Techniques: Binding Assay, Flow Cytometry, Labeling, Magnetic Beads, SPR Assay

Journal: Genes & Diseases

Article Title: Dual-mode aptamer-driven biosensing platform for ultrasensitive and mutation-resilient detection of the SARS-CoV-2 nucleocapsid protein

doi: 10.1016/j.gendis.2025.101943

Figure Lengend Snippet: Structural basis and binding mechanism of NP14 interaction with the SARS-CoV-2 N protein. (A) Molecular simulation of the binding mode between aptamer NP14 and the SARS-CoV-2 N protein ( http://www.rcsb.org , ID:6VYO) via AutoDock. (B) Enlarged view of the presumed binding area. (C) Nucleic acid sequences and corresponding amino acids involved in the docking model. (D) Secondary structure simulation of aptamer NP14 via the Nupack web server at 37 °C. (E) Secondary structure simulation of the truncated chains NP14a via the Nupack web server at 37 °C. (F) Secondary structure simulation of the truncated chains NP14b via the Nupack web server at 37 °C. (G) Binding analysis of NP14 with truncated NP14a, NP14b, and base-mutated 400 nM NP14a1, NP14a2, NP14a3, NP14a4, NP14b1, NP14b2, NP14b3, NP14b4, and NP14b5 to the SARS-CoV-2 N protein by ELONA. Data were presented as mean ± standard deviation of triplicate results ( n = 3). The NP14 control: ns, not significant; ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001. (H) Circular dichroism spectroscopy of AS1411 (20 μM) and NP14 (10 μM) was performed in PBS buffer (0.01 M, pH = 7.4) at wavelengths ranging from 220 to 320 nm. (I) Domain organization of the SARS-CoV-2 N protein, with numbers indicating domain boundaries. (J) Immunomagnetic beads (40 μL, 10 mg/mL) labeled with Flag antibodies against the truncated overexpressed protein were reacted with 300 nM biotin-labeled NP14 to assess binding. Data were presented as mean ± standard deviation of triplicate results ( n = 3). Compared with the blank control: ∗∗∗∗ p < 0.0001. (K) 250 nM biotin-labeled NP14 was mixed with 250 nM unlabeled N1, A58, A61 and competitive binding was analyzed by ELONA. Data were presented as mean ± standard deviation of four replicate results ( n = 4). Compared with the NP14: ns, not significant; ∗∗∗ p < 0.001. (L) Evaluation of the binding affinity for truncated proteins containing the NTD region at different concentrations of NP14 (0, 2, 5, 10, 20, 50, and 100 nM). Data were presented as mean ± standard deviation of triplicate results ( n = 3).

Article Snippet: X-Aptamer libraries were acquired from AM Biotechnologies (Houston, Texas, USA); His-Tag magnetic beads (Invitrogen, DynabeadsTM His-Tag Isolation & Pulldown, 10103D), SARS-CoV-2 N protein, and anti-SARS-CoV-2 N protein monoclonal antibodies (anti-SARS-CoV-2 N protein mAb, Cat: 40143-MM05, 40588-R001) were purchased from Sino Biological.

Techniques: Binding Assay, Standard Deviation, Control, Circular Dichroism, Spectroscopy, Labeling

Journal: Genes & Diseases

Article Title: Dual-mode aptamer-driven biosensing platform for ultrasensitive and mutation-resilient detection of the SARS-CoV-2 nucleocapsid protein

doi: 10.1016/j.gendis.2025.101943

Figure Lengend Snippet: Specificity and cross-variant recognition of NP14 for the SARS-CoV-2 N protein. (A) ELONA method detection mode diagram. (B) NP14 labeled with 400 nM biotin was used with various proteins (1 μg/mL): SARS-CoV N protein, human coronavirus (HCoV) 229E, OC43, HKU1, SARS-CoV-2 receptor-binding domain (RBD), alpha-fetoprotein (AFP), interleukin-4 (IL-4), bovine serum albumin (BSA), and influenza (InFlu) A and B proteins, to validate the specificity of NP14 via ELISA. Data were presented as mean ± standard deviation of triplicate results ( n = 3). Compared with the SARS-CoV-2 N protein: ns, not significant; ∗∗∗∗ p < 0.0001. (C) Direct detection of SARS-CoV-2 N protein binding activity at various concentrations (0, 0.5, 1, 5, 10, 20, 50, 100, 200, 500, 800, and 1000 ng/mL) via the ELONA platform. Data were presented as mean ± standard deviation of triplicate results ( n = 3). (D – L) Detection of NP14 (biotin-labeled, 400 nM) binding to N recombinant proteins from SARS-CoV-2 variants at different concentrations (0, 5, 10, 20, 50, 100, 200, 500, and 1000 ng/mL) on the direct ELONA platform. Variants included (D) alpha, (E) beta, (F) gamma, (G) delta, (H) omicron B.1.640, (I) omicron BA.2, (J) lambda, (K) omicron BA.1, and (L) omicron BA.4.

Article Snippet: X-Aptamer libraries were acquired from AM Biotechnologies (Houston, Texas, USA); His-Tag magnetic beads (Invitrogen, DynabeadsTM His-Tag Isolation & Pulldown, 10103D), SARS-CoV-2 N protein, and anti-SARS-CoV-2 N protein monoclonal antibodies (anti-SARS-CoV-2 N protein mAb, Cat: 40143-MM05, 40588-R001) were purchased from Sino Biological.

Techniques: Variant Assay, Labeling, Binding Assay, Enzyme-linked Immunosorbent Assay, Standard Deviation, Protein Binding, Activity Assay, Recombinant

Journal: Genes & Diseases

Article Title: Dual-mode aptamer-driven biosensing platform for ultrasensitive and mutation-resilient detection of the SARS-CoV-2 nucleocapsid protein

doi: 10.1016/j.gendis.2025.101943

Figure Lengend Snippet: Comparative sensitivity and specificity of antibody–antibody versus antibody–aptamer sandwich assays. (A) Standard curve for the sandwich assay (1 μg/mL antibody) using the SARS-CoV-2 N protein at various concentrations (0, 0.1, 0.5, 1, 5, 10, 20, 50, 100, 200, 500, and 1000 ng/mL). Data were presented as mean ± standard deviation of triplicate results ( n = 3). (B) Standard curve of the SARS-CoV-2 N protein in the antibody‒aptamer sandwich mode using SARS-CoV-2 N protein at various concentrations (0, 0.2, 0.5, 1, 5, 10, 20, 50, 100, 200, 500, and 1000 ng/mL). Data were presented as mean ± standard deviation of triplicate results ( n = 3). (C) Specificity validation with multiple proteins (1 μg/mL), including: SARS-CoV-2 receptor-binding domain (RBD), alpha-fetoprotein (AFP), interleukin-4 (IL-4), bovine serum albumin (BSA), influenza (InFlu) A and B proteins, to validate the specificity of the antibody–antibody (1 μg/mL) sandwich assay. Data were presented as mean ± standard deviation of triplicate results ( n = 3). Compared with the blank control: ns, not significant; ∗∗ p < 0.01 and ∗∗∗∗ p < 0.0001. (D) Validation was performed using multiple proteins at a concentration of 1 μg/mL, including: SARS-CoV-2 RBD, AFP, IL-4, BSA, InFlu A and B proteins, to validate the specificity of the antibody (1 μg/mL)-aptamer (200 nM) sandwich assay. Data were presented as mean ± standard deviation of triplicate results ( n = 3). Compared with the blank control: ns, not significant; ∗∗∗∗ p < 0.0001.

Article Snippet: X-Aptamer libraries were acquired from AM Biotechnologies (Houston, Texas, USA); His-Tag magnetic beads (Invitrogen, DynabeadsTM His-Tag Isolation & Pulldown, 10103D), SARS-CoV-2 N protein, and anti-SARS-CoV-2 N protein monoclonal antibodies (anti-SARS-CoV-2 N protein mAb, Cat: 40143-MM05, 40588-R001) were purchased from Sino Biological.

Techniques: Standard Deviation, Biomarker Discovery, Binding Assay, Control, Concentration Assay

Journal: Genes & Diseases

Article Title: Dual-mode aptamer-driven biosensing platform for ultrasensitive and mutation-resilient detection of the SARS-CoV-2 nucleocapsid protein

doi: 10.1016/j.gendis.2025.101943

Figure Lengend Snippet: Analytical performance of the MD ELAAA platform in detecting the SARS-CoV-2 N protein and viral cultures. (A) Schematic illustration of the modulation of the Ag shell layer thickness in core–shell AuNFs@Ag nanostructures leading to changes in the localized surface plasmon resonance (LSPR) and light scattering intensity. (B) Standard curve of the MD ELAAA method for different SARS-CoV-2 N proteins (0, 0.005, 0.01, 0.02, 0.05, 0.1, 0.5, 1, 2, and 5 ng/mL). Data were presented as mean ± standard deviation of triplicate results ( n = 3). (C) Validation was performed using multiple proteins at a concentration of 1 ng/mL, including: SARS-CoV-2 receptor-binding domain (RBD), alpha-fetoprotein (AFP), interleukin-4 (IL-4), bovine serum albumin (BSA), influenza (InFlu) A and B proteins, to validate the specificity of the MD ELAAA platform. Data were presented as mean ± standard deviation of triplicate results ( n = 3). The blank control: ns, not significant; ∗∗∗∗ p < 0.0001. (D) Standard curve of the MD ELAAA method for SARS-CoV-2 virus cultures at different concentrations (0, 1, 2, 5, 10, 20, 50, 100, and 200 TCID 50 /mL). Data were presented as mean ± standard deviation of triplicate results ( n = 3). (E) Standard curve of the ELAAA method for SARS-CoV-2 virus cultures at different concentrations (0, 10, 20, 50, 100, 200, 300, 500, and 1000 TCID 50 /mL). Data were presented as mean ± standard deviation of triplicate results ( n = 3).

Article Snippet: X-Aptamer libraries were acquired from AM Biotechnologies (Houston, Texas, USA); His-Tag magnetic beads (Invitrogen, DynabeadsTM His-Tag Isolation & Pulldown, 10103D), SARS-CoV-2 N protein, and anti-SARS-CoV-2 N protein monoclonal antibodies (anti-SARS-CoV-2 N protein mAb, Cat: 40143-MM05, 40588-R001) were purchased from Sino Biological.

Techniques: SPR Assay, Standard Deviation, Biomarker Discovery, Concentration Assay, Binding Assay, Control, Virus

Journal: Genes & Diseases

Article Title: Dual-mode aptamer-driven biosensing platform for ultrasensitive and mutation-resilient detection of the SARS-CoV-2 nucleocapsid protein

doi: 10.1016/j.gendis.2025.101943

Figure Lengend Snippet: Specificity and cross-variant recognition of NP14 for the SARS-CoV-2 N protein. (A) ELONA method detection mode diagram. (B) NP14 labeled with 400 nM biotin was used with various proteins (1 μg/mL): SARS-CoV N protein, human coronavirus (HCoV) 229E, OC43, HKU1, SARS-CoV-2 receptor-binding domain (RBD), alpha-fetoprotein (AFP), interleukin-4 (IL-4), bovine serum albumin (BSA), and influenza (InFlu) A and B proteins, to validate the specificity of NP14 via ELISA. Data were presented as mean ± standard deviation of triplicate results ( n = 3). Compared with the SARS-CoV-2 N protein: ns, not significant; ∗∗∗∗ p < 0.0001. (C) Direct detection of SARS-CoV-2 N protein binding activity at various concentrations (0, 0.5, 1, 5, 10, 20, 50, 100, 200, 500, 800, and 1000 ng/mL) via the ELONA platform. Data were presented as mean ± standard deviation of triplicate results ( n = 3). (D – L) Detection of NP14 (biotin-labeled, 400 nM) binding to N recombinant proteins from SARS-CoV-2 variants at different concentrations (0, 5, 10, 20, 50, 100, 200, 500, and 1000 ng/mL) on the direct ELONA platform. Variants included (D) alpha, (E) beta, (F) gamma, (G) delta, (H) omicron B.1.640, (I) omicron BA.2, (J) lambda, (K) omicron BA.1, and (L) omicron BA.4.

Article Snippet: The SARS-CoV N protein, HCoVs229E, HCoVs-OC43, HCoVs-HKU1, and ELISA kits (Cat: CSB-EL33251, https://www.cusabio.com/ ) were purchased from CUSABIO (Wuhan, China).

Techniques: Variant Assay, Labeling, Binding Assay, Enzyme-linked Immunosorbent Assay, Standard Deviation, Protein Binding, Activity Assay, Recombinant

Journal: Journal of Traditional and Complementary Medicine

Article Title: Chemical characterization of Jing Guan Fang and its application in alleviating coronavirus envelope protein-induced proinflammatory responses in vitro and in vivo

doi: 10.1016/j.jtcme.2025.12.003

Figure Lengend Snippet: JGF inhibits NO, IL-6, and TNF-α production in RAW264.7 and MH-S cells. The cells were treated with JGF (50, 100, 150, 300, 600 μg/mL), 2-E (0.1 μM), DXT (10 μM), or LPS (0.1 μg/mL) for 24 h. ( A ) Cell viability was evaluated using crystal violet. ( B ) NO production was measured using the Griess assay. ( C-D ) IL-6 ( C ) and TNF-α ( D ) levels were determined by ELISA. EC 50 was calculated by CompuSyn software. Data was presented as mean ± standard deviation (SD) for groups (n = 3). Significant differences are denoted as ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001.

Article Snippet: SARS-CoV-2-envelope (2-E) protein (Cat# NBP2–90986) was purchased from Novus Biologicals (Littleton, CO, USA).

Techniques: Griess Assay, Enzyme-linked Immunosorbent Assay, Software, Standard Deviation

Journal: Journal of Traditional and Complementary Medicine

Article Title: Chemical characterization of Jing Guan Fang and its application in alleviating coronavirus envelope protein-induced proinflammatory responses in vitro and in vivo

doi: 10.1016/j.jtcme.2025.12.003

Figure Lengend Snippet: Components of JGF inhibit 2-E-induced inflammation. The RAW264.7 and MH-S cells were co-treated with JGF compounds and 2-E for 24 h. ( A ) The 3D-HPLC fingerprint of JGF. Compound structures were sourced from the PubChem database. The detection wavelength ranged from 200 to 400 nm, and the injection volume was 20 μL. ( B ) Cell viability was evaluated using crystal violet. ( C ) NO production was measured using the Griess assay. ( D-E ) IL-6 ( D ) and TNF-α ( E ) levels were determined by ELISA. Data are presented as mean ± SD (n = 3). Statistical significance was determined relative to the 2-E group. Significant differences are denoted as ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001.

Article Snippet: SARS-CoV-2-envelope (2-E) protein (Cat# NBP2–90986) was purchased from Novus Biologicals (Littleton, CO, USA).

Techniques: Injection, Griess Assay, Enzyme-linked Immunosorbent Assay

Journal: Journal of Traditional and Complementary Medicine

Article Title: Chemical characterization of Jing Guan Fang and its application in alleviating coronavirus envelope protein-induced proinflammatory responses in vitro and in vivo

doi: 10.1016/j.jtcme.2025.12.003

Figure Lengend Snippet: JGF downregulates 2-E-induced iNOS and COX-2 in RAW264.7 and MH-S cells. Cells were treated with JGF (0, 50, 200 μg/mL) or 2-E (0.1 μM) for 24 h. ( A ) Protein levels of iNOS and COX-2 in macrophages were measured by Western blot. ( B-C ) Quantification of iNOS and COX-2 in cells without ( B ) and with ( C ) 2-E stimulation, calculated using ImageJ. Actin was used as the internal control. The non-detected data showed as – or ND. Data are presented as mean ± SD (n = 3). Significant differences are denoted as ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.

Article Snippet: SARS-CoV-2-envelope (2-E) protein (Cat# NBP2–90986) was purchased from Novus Biologicals (Littleton, CO, USA).

Techniques: Western Blot, Control

Journal: Journal of Traditional and Complementary Medicine

Article Title: Chemical characterization of Jing Guan Fang and its application in alleviating coronavirus envelope protein-induced proinflammatory responses in vitro and in vivo

doi: 10.1016/j.jtcme.2025.12.003

Figure Lengend Snippet: JGF inhibits 2-E-induced phosphorylation of STAT3 in RAW264.7 and MH-S cells. Cells were treated with JGF (0, 50, 200 μg/mL) or 2-E (0.1 μM) for 3 h. ( A ) Protein levels of phosphorylated JAK2 and STAT3 were measured by Western blot. ( B-C ) Quantification of phosphorylated JAK2 and STAT3 in cells without ( B ) and with ( C ) 2-E stimulation, calculated using ImageJ. Actin was used as the internal control. Data are presented as mean ± SD (n = 3). Significant differences are denoted as ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.

Article Snippet: SARS-CoV-2-envelope (2-E) protein (Cat# NBP2–90986) was purchased from Novus Biologicals (Littleton, CO, USA).

Techniques: Phospho-proteomics, Western Blot, Control

Journal: Journal of Traditional and Complementary Medicine

Article Title: Chemical characterization of Jing Guan Fang and its application in alleviating coronavirus envelope protein-induced proinflammatory responses in vitro and in vivo

doi: 10.1016/j.jtcme.2025.12.003

Figure Lengend Snippet: JGF inhibits 2-E-induced phosphorylation of ERK1/2 in RAW264.7 and MH-S cells. Cells were treated with JGF (0, 50, 200 μg/mL) or 2-E (0.1 μM) for 3 h. ( A ) Protein levels of phosphorylated JNK1/2, ERK1/2, p38, and p65 were measured by Western blot. ( B-C ) Quantification of phosphorylated JNK1/2, ERK1/2, p38, and p65 in cells without ( B ) and with ( C ) 2-E stimulation, calculated using ImageJ. Actin was used as the internal control. Data are presented as mean ± SD (n = 3). Significant differences are denoted as ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.

Article Snippet: SARS-CoV-2-envelope (2-E) protein (Cat# NBP2–90986) was purchased from Novus Biologicals (Littleton, CO, USA).

Techniques: Phospho-proteomics, Western Blot, Control

Journal: Journal of Traditional and Complementary Medicine

Article Title: Chemical characterization of Jing Guan Fang and its application in alleviating coronavirus envelope protein-induced proinflammatory responses in vitro and in vivo

doi: 10.1016/j.jtcme.2025.12.003

Figure Lengend Snippet: JGF reduces the 2-E-induced proinflammatory cytokines in vivo . ( A ) The experimental scheme for mouse exposure. ( B-F ) Levels of IL-6 ( B ), TNF-α ( C ), IFN-γ ( D ), IL-1β ( E ), and IL-12 ( F ) in lung tissue and serum were measured by ELISA. Data are presented as mean ± SD (n = 9 for serum, except DXT group n = 6; n = 6 for lung tissue, except DXT group n = 3) ( G ) Representative histological images of lung tissue stained with H&E and IHC images for IL-6, TNF-α, and IL-1β expression. ( H-J ) Quantification of IL-6 ( H ), TNF-α ( I ), and IL-1β ( J ) positive areas using ImageJ (n = 3). Significant differences between the control (CTL) group and other groups are denoted by ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001. Significant differences between the 2-E group and 2-E + JGF group are indicated by #p < 0.05, ##p < 0.01, ###p < 0.001.

Article Snippet: SARS-CoV-2-envelope (2-E) protein (Cat# NBP2–90986) was purchased from Novus Biologicals (Littleton, CO, USA).

Techniques: In Vivo, Enzyme-linked Immunosorbent Assay, Staining, Expressing, Control

Journal: Journal of Traditional and Complementary Medicine

Article Title: Chemical characterization of Jing Guan Fang and its application in alleviating coronavirus envelope protein-induced proinflammatory responses in vitro and in vivo

doi: 10.1016/j.jtcme.2025.12.003

Figure Lengend Snippet: Schematics showing the anti-inflammatory mechanism of JGF in 2-E-induced mice macrophages.

Article Snippet: SARS-CoV-2-envelope (2-E) protein (Cat# NBP2–90986) was purchased from Novus Biologicals (Littleton, CO, USA).

Techniques:

Journal: medRxiv

Article Title: Diagnostic Classification for Long Covid Patients identifying Persistent Virus and Hyperimmune Pathophysiologies

doi: 10.64898/2026.04.21.26351402

Figure Lengend Snippet:

Article Snippet: The integrity of the spike protein samples on the surface was tested using an anti-S2 antibody (SinoBiological, 40590-D001), chosen for its specificity to a pan-variant conserved epitope.

Techniques: