Journal: ACS Nano

Article Title: Biomimetic Virus-Like Particles as Severe Acute Respiratory Syndrome Coronavirus 2 Diagnostic Tools

doi: 10.1021/acsnano.0c08430

Figure Lengend Snippet: Overall schematic and workflow of the VLP-based biomimetic SARS-CoV-2 positive controls. (A) Design of SDM from 5′ to 3′: T7 promoter (gray), Qβ hairpin (yellow), N1 (red), N2 (green), and RP (black), T7 terminator (gray). (B) Production of Qβ 1P–C19 and Qβ 2P–C19 VLPs via in vivo assembly. Qβ 1P–C19 VLPs were produced from a one-plasmid system, where the gene of the Qβ CPs and SDM RNAs were cloned in one vector (pCDFDuet-1). Qβ 2P–C19 VLPs were produced from a two-plasmid system, where Qβ CP was derived from pCDFDuet-Qβ and SDM RNA was produced from pET-28a (+). (C) CCMV–C19 VLPs were produced by in vitro reconstitution of the transcribed SDM RNAs with purified CCMV CPs.

Article Snippet: SARS-CoV-2 detection module (SDM) gene from plasmid Qβ 1P–C19 was subcloned into plasmid pET-28a(+) by amplifying with Qβ 2P–C19 forward primer (5′-GAA GAT CTT AAT ACG ACT CAC TAT AGG G-3′) and Qβ 2P–C19 reverse primer (5′-TTT TCC TTT TGC GGC CGC CAA AAA ACC CCT CAA GAC CCG TTT AGA G-3′) using NEB Q5 high fidelity 2X master mix.

Techniques: In Vivo, Produced, Plasmid Preparation, Clone Assay, Derivative Assay, In Vitro, Purification

Journal: ACS Nano

Article Title: Biomimetic Virus-Like Particles as Severe Acute Respiratory Syndrome Coronavirus 2 Diagnostic Tools

doi: 10.1021/acsnano.0c08430

Figure Lengend Snippet: Characterization of VLP-based SARS-CoV-2 positive controls. Agarose gel: Native agarose gel electrophoresis of VLPs packaging the SDM with gels were stained with GelRed nucleic acid stain (RNA stain) and Coomassie blue (protein stain) to show the presence of RNAs and VLPs. DLS: Dynamic light scattering (DLS) of VLPs packaging the SDM; triplicate samples were analyzed, and representative data sets are shown. TEM: Imaging of negatively stained VLPs packaging the SDM using transmission electron microscope (TEM). Average size of 20 particles tabulated by ImageJ software is stated in the inset box. SEC: Analysis of positive controls by size exclusion chromatography (SEC) using a Superose 6 column and GE Healthcare Äkta Purifier chromatography system; protein was detected at 280 nm, and RNA was detected at 260 nm.

Article Snippet: SARS-CoV-2 detection module (SDM) gene from plasmid Qβ 1P–C19 was subcloned into plasmid pET-28a(+) by amplifying with Qβ 2P–C19 forward primer (5′-GAA GAT CTT AAT ACG ACT CAC TAT AGG G-3′) and Qβ 2P–C19 reverse primer (5′-TTT TCC TTT TGC GGC CGC CAA AAA ACC CCT CAA GAC CCG TTT AGA G-3′) using NEB Q5 high fidelity 2X master mix.

Techniques: Agarose Gel Electrophoresis, Staining, Imaging, Transmission Assay, Microscopy, Software, Size-exclusion Chromatography, Chromatography

Journal: ACS Nano

Article Title: Biomimetic Virus-Like Particles as Severe Acute Respiratory Syndrome Coronavirus 2 Diagnostic Tools

doi: 10.1021/acsnano.0c08430

Figure Lengend Snippet: Validation of VLP-based SARS-CoV-2 positive controls in the clinical setting using ddPCR detection of SARS-CoV-2. (A) ddPCR 1-D amplitude plots of SARS-CoV-2 positive controls according to N1, N2, and RP regions. Lane 1: Qβ 1P–C19. Lane 2: Qβ 2P–C19. Lane 3: CCMV–C19. Lane 4: (+) Clinical sample from COVID-19 confirmed patient. Lane 5: (−) Clinical sample from healthy patient for N1 and N2 (negative control); no template control for RP. Amplifications were performed in triplicate. (B) Scatter plot comparing copy numbers of SARS-CoV-2 detection regions (N1, N2, RP) for all positive controls. (C) Tabulated SDM RNA copy number for each SARS-CoV-2 positive controls.

Article Snippet: SARS-CoV-2 detection module (SDM) gene from plasmid Qβ 1P–C19 was subcloned into plasmid pET-28a(+) by amplifying with Qβ 2P–C19 forward primer (5′-GAA GAT CTT AAT ACG ACT CAC TAT AGG G-3′) and Qβ 2P–C19 reverse primer (5′-TTT TCC TTT TGC GGC CGC CAA AAA ACC CCT CAA GAC CCG TTT AGA G-3′) using NEB Q5 high fidelity 2X master mix.

Techniques: Negative Control

Journal: ACS Nano

Article Title: Biomimetic Virus-Like Particles as Severe Acute Respiratory Syndrome Coronavirus 2 Diagnostic Tools

doi: 10.1021/acsnano.0c08430

Figure Lengend Snippet: Characterization of VLP-based SARS-CoV-2 positive controls after 1-month storage under ambient conditions. Agarose gel: Native agarose gel electrophoresis of VLP-based SARS-CoV-2 positive controls reveals intact VLPs; gels were stained with GelRed nucleic acid stain (RNA stain) and Coomassie blue (protein stain) to show the presence of RNAs and VLPs. DLS: Dynamic light scattering of VLPs packaging the SDM; triplicate samples were analyzed, and representative data sets are shown. TEM: Imaging of negatively stained VLPs packaging the SDM using transmission electron microscope. Average size of 20 particles tabulated by ImageJ software was stated in inset box. SEC: Analysis of positive controls by size exclusion chromatography using a Superose 6 column and GE Healthcare Äkta Purifier chromatography system; protein was detected at 280 nm and RNA was detected at 260 nm. See also Figure , showing the characterization of freshly prepared samples.

Article Snippet: SARS-CoV-2 detection module (SDM) gene from plasmid Qβ 1P–C19 was subcloned into plasmid pET-28a(+) by amplifying with Qβ 2P–C19 forward primer (5′-GAA GAT CTT AAT ACG ACT CAC TAT AGG G-3′) and Qβ 2P–C19 reverse primer (5′-TTT TCC TTT TGC GGC CGC CAA AAA ACC CCT CAA GAC CCG TTT AGA G-3′) using NEB Q5 high fidelity 2X master mix.

Techniques: Agarose Gel Electrophoresis, Staining, Imaging, Transmission Assay, Microscopy, Software, Size-exclusion Chromatography, Chromatography

Journal: ACS Nano

Article Title: Biomimetic Virus-Like Particles as Severe Acute Respiratory Syndrome Coronavirus 2 Diagnostic Tools

doi: 10.1021/acsnano.0c08430

Figure Lengend Snippet: Stability of VLP-based SARS-CoV-2 positive controls in respect to time and temperature. RT-qPCR was performed to obtain the C q values. Triplicates were performed on each sample with the error bar showing the standard deviation.

Article Snippet: SARS-CoV-2 detection module (SDM) gene from plasmid Qβ 1P–C19 was subcloned into plasmid pET-28a(+) by amplifying with Qβ 2P–C19 forward primer (5′-GAA GAT CTT AAT ACG ACT CAC TAT AGG G-3′) and Qβ 2P–C19 reverse primer (5′-TTT TCC TTT TGC GGC CGC CAA AAA ACC CCT CAA GAC CCG TTT AGA G-3′) using NEB Q5 high fidelity 2X master mix.

Techniques: Quantitative RT-PCR, Standard Deviation

Journal: Cells

Article Title: A Defective Viral Particle Approach to COVID-19

doi: 10.3390/cells11020302

Figure Lengend Snippet: Harnessing EV properties to combat SARS-CoV-2 infection or treat COVID-19. ( A ) EVs can be used to deliver nucleic acid sequences that either modulate the expression of specific targets, express genes of interests, or encode for viral products. ( B ) EVs can be used to deliver compounds of interest. ( C ) EVs derived from a specific cell type or tissue could be used to mitigate disease and trigger tissue regeneration. ( D ) EVs carrying the spike protein can be used to antagonize viral entry into a host cell. ( E ) EVs carrying the virus entry receptor ACE2 could serve as decoys for the virus. ( F ) EVs carrying viral antigens could be used for vaccine development. The image was generated with BioRender.com (accessed on 13 September 2021).

Article Snippet: Asuragen and SeraCare have announced developments of SARS-CoV-2 positive controls for diagnostics, in which a SARS-CoV-2 detection module for RT-PCR was encapsidated into VLPs from the bacteriophage Qβ and the CCMV virus [ ].

Techniques: Infection, Expressing, Derivative Assay, Generated

Journal: Cells

Article Title: A Defective Viral Particle Approach to COVID-19

doi: 10.3390/cells11020302

Figure Lengend Snippet: The SARS-CoV-2 genome. On entry into the host cell, a coronavirus particle is uncoated, and its single-stranded positive-sense RNA genome enters the cytoplasm. Two-thirds of the coronavirus genome is occupied by two large overlapping open reading frames (ORF1a and ORF1b) that are translated into polyproteins and that are processed to generate 16 non-structural proteins (nsp1 to nsp16). The rest of the genome includes ORFs for the structural proteins and several accessory proteins. The 5′-UTR is 265 nucleotides long, while the 3′-UTR is 358 nucleotides. The major distinction between other coronaviruses related to SARS-CoV and SARS-CoV-2 is in orf3b, Spike and orf8 but especially in the highly variable Spike S1 and orf8, which are recombination hot spots.

Article Snippet: Asuragen and SeraCare have announced developments of SARS-CoV-2 positive controls for diagnostics, in which a SARS-CoV-2 detection module for RT-PCR was encapsidated into VLPs from the bacteriophage Qβ and the CCMV virus [ ].

Techniques:

Journal: Medicine International

Article Title: Anti‑epidermal growth factor receptor monoclonal antibody therapy in locally advanced head and neck cancer: A systematic review of phase III clinical trials

doi: 10.3892/mi.2024.165

Figure Lengend Snippet: RCTs testing the effectiveness of anti-EGFR monoclonal antibody in treating locally advanced head and neck squamous cell carcinoma.

Article Snippet: Gebre-Medhin et al , 2021 (ARTSCAN III trial) , Cetuximab , RCT , Patients ≥18 years with previously untreated squamous cell carcinoma of oropharynx, hypopharynx or larynx, stage III-IV according to UICC TNM classification, 7th edition, without distant metastasis. Aimed for curative treatment with definitive RT (n=298) , RT +cetuximab RT (68 Gy to the primary tumour and lymph node metastases, 54.4 Gy to elective neck volumes given as daily fractions of 2.0 Gy and 1.6 Gy respectively, 5 days per week for seven weeks + Cetuximab (400 mg/m 2 as loading dose 1 week before start of RT, followed by 250 mg/m 2 per week for 7 weeks Patients with T3-T4 tumour underwent another random assignment of 63 Gy or 73.1 Gy given to the primary tumour as daily fractions of 2.15 Gy. Used IMRT, helical tomotherapy or volumetric arc therapy for radiation planning. (n=149) , Chemo-RT RT (68 Gy to the primary tumour and lymph node metastases, 54.4 Gy to elective neck volumes given as daily fractions of 2.0 Gy and 1.6 Gy respectively,5 days per week for seven weeks + Cisplatin 40 mg/m 2 per week for seven weeks Patients with T3-T4 tumour underwent another random assignment of 63 Gy or 73.1 Gy given to the primary tumour as daily fractions of 2.15 Gy Used IMRT, helical tomotherapy or volumetric arc therapy for radiation planning. (n=149) , 39 months , OS: 3-year OS, 78% (95% CI, 71-85) vs. 88% (95% CI, 83-94); adjusted HR, 1.63; 95% CI, 0.93-2.86; P=0.086. Post hoc subgroup analyses: HR for OS, 5.70 (95% CI, 1.67-19.5) for patients with p16 + -OPC. Cetuximab + RT vs. cisplatin + RT (P=0.03) , LRC: Locoregional failure at 3 years, 23% (95% CI, 16-31) vs. 9% (95% CI, 4-14). Adjusted cause-specific HR, 2.49; 95% CI, 1.33-4.66; P=0.0045. Distant failure: NS. Adverse events: Acute mucositis (P=0.035), skin reactions (P=0.001) and acneiform rashes (P<0.001) were more common in the cetuximab group compared with the cisplatin group. Nausea (P=0.001), vomiting (P=0.015), acute kidney injury (P<0.001), tinnitus (P=0.002), dysphagia (P=0.033) and neutropenia (P<0.001) were significantly more common in the cisplatin arm. Late toxicities such as taste alteration and hearing impairment were more common in the cisplatin arm, while late pain and mucosal toxicities were more common in the cetuximab arm.. , ( ) .

Techniques: Control, Computed Tomography, Fractionation, Modification