Journal: RSC Advances

Article Title: Pre-clinically evaluated visual lateral flow platform using influenza A and B nucleoprotein as a model and its potential applications †

doi: 10.1039/d1ra01361k

Figure Lengend Snippet: Specificity of the system against different related respiratory viruses including influenza (Flu) A and B, RSV, adenovirus (ADV), parainfluenza (PI) virus, and other non-related proteins including avidin, CRP, haemoglobin (Hb) protein, and insulin. The presence of the signal at either TL1 or TL2 position, together with the C-line signal, indicates the positive result for influenza A and B virus, respectively.

Article Snippet: Primary mouse monoclonal antibodies against influenza A nucleoprotein and influenza B nucleoprotein, secondary anti-mouse monoclonal with and without alkaline phosphatase enzyme (ALP) conjugation antibodies, and all respiratory virus protein antigens including recombinant influenza A nucleoprotein, influenza B virus-infected Madin–Darby Canine Kidney (MDCK) Cell lysate, RSV, parainfluenza virus, and adenovirus proteins were supplied from Fitzgerald, MA, USA, East Coast Bio, TN, USA, and Meridian Life Sciences, TN, USA, and Seracare (MA, USA).

Techniques: Avidin-Biotin Assay

Journal: Anesthesiology

Article Title: Intrathecal Injection of Metabotropic Glutamate Receptor Subtype 3 and 5 Agonist/Antagonist Attenuates Bone Cancer Pain by Inhibition of Spinal Astrocyte Activation in a Mouse Model

doi: 10.1097/aln.0b013e31823de68d

Figure Lengend Snippet: Fig. 2. Quantitative real-time reverse transcription-polymer- ase chain reaction analysis of changes of spinal glial fibrillary acidic protein (GFAP), mGluR3, and mGluR5 messenger RNA (mRNA) expression after the surgery on mice. Spinal GFAP (A), mGluR3 (B), and mGluR5 (C) mRNA expression progres- sively increased as time went on after day 7 in tumor-bearing mice. Each group used five mice. Data were presented as fold change of control (day 0) mean SD. # P 0.05 versus day 0. * P 0.05 versus sham mice.

Article Snippet: Membranes were blocked in PBS/5% skim milk/0.1% Tween 20 for 2 h at room temperature, followed by overnight incubation at 4°C with anti mGluR3 (goat affinity purified polyclonal antibody; 1:500; Santa Cruz Biotechnology, Santa Cruz, CA), anti mGluR5 (rabbit affinity purified polyclonal antibody; 1:500; Abcam, Hong Kong, China), anti-GFAP (goat affinity purified polyclonal antibody; 1:1000; Santa Cruz Biotechnology) or anti- -actin (rabbit affinity purified polyclonal antibody; 1:1000;Cell Signaling, Danvers, MA) diluted in blocking buffer.

Techniques: Reverse Transcription, Polymer, Expressing, Control

Journal: Anesthesiology

Article Title: Intrathecal Injection of Metabotropic Glutamate Receptor Subtype 3 and 5 Agonist/Antagonist Attenuates Bone Cancer Pain by Inhibition of Spinal Astrocyte Activation in a Mouse Model

doi: 10.1097/aln.0b013e31823de68d

Figure Lengend Snippet: Fig. 3. Changes of spinal glial fibrillary acidic protein (GFAP), mGluR3, and mGluR5 protein expression over time in tumor- bearing mice and sham group mice. Western blot for -actin, GFAP, mGluR3, and mGluR5 resulted in products of 45, 50, 110 and 150 kDa, as expected (markers show predicted band sizes) (A and B). Densitometric quantification of -actin, GFAP, mGluR3, and mGluR5 immunoreactivity on Western blots (C, D, and E). Each group used five mice. Data were presented as fold change of control (day 0) mean SD. # P 0.05 versus day 0. * P 0.05 versus sham mice.

Article Snippet: Membranes were blocked in PBS/5% skim milk/0.1% Tween 20 for 2 h at room temperature, followed by overnight incubation at 4°C with anti mGluR3 (goat affinity purified polyclonal antibody; 1:500; Santa Cruz Biotechnology, Santa Cruz, CA), anti mGluR5 (rabbit affinity purified polyclonal antibody; 1:500; Abcam, Hong Kong, China), anti-GFAP (goat affinity purified polyclonal antibody; 1:1000; Santa Cruz Biotechnology) or anti- -actin (rabbit affinity purified polyclonal antibody; 1:1000;Cell Signaling, Danvers, MA) diluted in blocking buffer.

Techniques: Expressing, Western Blot, Control

Journal: Anesthesiology

Article Title: Intrathecal Injection of Metabotropic Glutamate Receptor Subtype 3 and 5 Agonist/Antagonist Attenuates Bone Cancer Pain by Inhibition of Spinal Astrocyte Activation in a Mouse Model

doi: 10.1097/aln.0b013e31823de68d

Figure Lengend Snippet: Fig. 5. Effects of chronic intrathecal administration of APDC (mGluR3 agonist,150 nmol/5 l), LY 341495 (mGluR3 antag- onist,15 nmol/5 l), CHPG (mGluR5 agonist, 300 mol/5 l), MTEP (mGluR5 antagonist, 150 nmol/5 l), morphine (5 mol/5 l), fluorocitrate (0.75 nmol/5 l), Vehicle 1 (dilute NaOH in 0.9% saline; 5 l) and Vehicle 2 (dilute DMSO in 0.9% saline; 5 l) on hind limb pain behaviors. (A) The num- ber of flinches during 2 min. (B) Paw withdrawal mechanical threshold to von Frey filaments. (C) Paw withdrawal thermal latencyto radiant heat. Animals were treated once daily on days 14–20 after surgery with the drugs or vehicles. Pain behaviors were assessed on day 7 after the first intrathecal injection. Ipsi ipsilateral hind limb to the surgery; Contra Contralateral hind limb to the surgery. Each column repre- sented mean SD for eight animals. If the two groups were marked with different letters (a, b, c, or d), it showed the data were of statistical difference between these two groups (P 0.05).

Article Snippet: Membranes were blocked in PBS/5% skim milk/0.1% Tween 20 for 2 h at room temperature, followed by overnight incubation at 4°C with anti mGluR3 (goat affinity purified polyclonal antibody; 1:500; Santa Cruz Biotechnology, Santa Cruz, CA), anti mGluR5 (rabbit affinity purified polyclonal antibody; 1:500; Abcam, Hong Kong, China), anti-GFAP (goat affinity purified polyclonal antibody; 1:1000; Santa Cruz Biotechnology) or anti- -actin (rabbit affinity purified polyclonal antibody; 1:1000;Cell Signaling, Danvers, MA) diluted in blocking buffer.

Techniques: Saline, Injection

Journal: Anesthesiology

Article Title: Intrathecal Injection of Metabotropic Glutamate Receptor Subtype 3 and 5 Agonist/Antagonist Attenuates Bone Cancer Pain by Inhibition of Spinal Astrocyte Activation in a Mouse Model

doi: 10.1097/aln.0b013e31823de68d

Figure Lengend Snippet: Fig. 6. Effects of intrathecal administration of APDC (mGluR3 agonist; 150 nmol/5 l), LY 341495 (mGluR3 antagonist; 15 nmol/5 l), CHPG (mGluR5 agonist; 300 mol/5 l), MTEP (mGluR5 antagonist; 150 nmol/5 l), morphine (5 mol/5 l), fluorocitrate (0.75 nmol/5 l), Vehicle 1 (dilute NaOH in 0.9% saline; 5 l) and Vehicle 2 (dilute DMSO in 0.9% saline; 5 l) on spinal glial fibrillary acidic protein (GFAP) messenger RNA (mRNA) (A) and protein (B and C) expression. Animals were treated with once-daily injections on days 14–20 after surgery with the drugs or vehicles. The spinal cord was removed on day 7 after the first intrathecal injection. Each group used five mice. The mRNA and protein data were presented as fold change of control (day 0) mean SD. If the two groups were marked with different letters (a, b, c, or d), it showed the data were of statistical difference between these two groups (P 0.05).

Article Snippet: Membranes were blocked in PBS/5% skim milk/0.1% Tween 20 for 2 h at room temperature, followed by overnight incubation at 4°C with anti mGluR3 (goat affinity purified polyclonal antibody; 1:500; Santa Cruz Biotechnology, Santa Cruz, CA), anti mGluR5 (rabbit affinity purified polyclonal antibody; 1:500; Abcam, Hong Kong, China), anti-GFAP (goat affinity purified polyclonal antibody; 1:1000; Santa Cruz Biotechnology) or anti- -actin (rabbit affinity purified polyclonal antibody; 1:1000;Cell Signaling, Danvers, MA) diluted in blocking buffer.

Techniques: Saline, Expressing, Injection, Control

Journal: Anesthesiology

Article Title: Intrathecal Injection of Metabotropic Glutamate Receptor Subtype 3 and 5 Agonist/Antagonist Attenuates Bone Cancer Pain by Inhibition of Spinal Astrocyte Activation in a Mouse Model

doi: 10.1097/aln.0b013e31823de68d

Figure Lengend Snippet: Fig. 7. Effects of intrathecal administration of APDC (mGluR3 agonist; 150 nmol/5 l), LY 341495 (mGluR3 antagonist; 15 nmol/5 l), CHPG (mGluR5 agonist; 300 mol/5 l), MTEP (mGluR5 antagonist; 150 nmol/5 l), morphine (5 mol/5 l), fluorocitrate (0.75 nmol/5 l), Vehicle 1 (dilute NaOH in 0.9% saline; 5 l) and Vehicle 2 (dilute DMSO in 0.9% saline; 5 l) on tumor growth. The ratio of maximum thigh circumference (ipsi/contra) of mice indicated the size of the tumor. Animals were treated once daily injections on days 14–20 after sur- gery with the drugs or vehicles. Data were presented as mean SD. If the two groups were marked with different letters (a, b), it showed the data were of statistical difference between these two groups (P 0.05).

Article Snippet: Membranes were blocked in PBS/5% skim milk/0.1% Tween 20 for 2 h at room temperature, followed by overnight incubation at 4°C with anti mGluR3 (goat affinity purified polyclonal antibody; 1:500; Santa Cruz Biotechnology, Santa Cruz, CA), anti mGluR5 (rabbit affinity purified polyclonal antibody; 1:500; Abcam, Hong Kong, China), anti-GFAP (goat affinity purified polyclonal antibody; 1:1000; Santa Cruz Biotechnology) or anti- -actin (rabbit affinity purified polyclonal antibody; 1:1000;Cell Signaling, Danvers, MA) diluted in blocking buffer.

Techniques: Saline

Journal:

Article Title: Involvement of Actin Microfilaments in the Replication of Human Parainfluenza Virus Type 3

doi:

Figure Lengend Snippet: Protein distribution between the SOL fraction and CSK framework. (A) CV-1 cells were infected with HPIV3 and, at different times postinfection (p.i.) (as indicated), pulse-labeled with [35S]methionine for 10 min and chased for 60 min. The labeled cells were fractionated into CSK and SOL fractions with an extraction buffer as described by Hamaguchi et al. (24). The fractions were analyzed by SDS-polyacrylamide gel electrophoresis followed by fluorography and autoradiography. The migration positions of molecular size markers in kilodaltons are shown on the right. (B) Western blot analysis of the proteins in soluble and cytoskeletal fractions. The distribution of HPIV3 NP and the cytoskeletal component actin between the SOL and CSK fractions was determined by the same procedure as described above, except the cells were not labeled and the NP and actin were analyzed by Western blotting with anti-RNP that detects primarily NP and anti-actin antibodies, respectively.

Article Snippet: The fixed cells were stained with goat anti-HPIV3 antibody (Biodesign) and anti-goat IgG conjugated with biotin followed by FITC-conjugated avidin.

Techniques: Infection, Labeling, Polyacrylamide Gel Electrophoresis, Autoradiography, Migration, Western Blot

Journal:

Article Title: Involvement of Actin Microfilaments in the Replication of Human Parainfluenza Virus Type 3

doi:

Figure Lengend Snippet: Detection of HPIV3 RNAs in the CSK framework. CV-1 cells, grown on coverslips, were infected with HPIV3 at 1 PFU/cell. The cells were treated with CSK buffer and hybridized with genome sense or antisense NP RNA labeled with digoxigenin. The coverslips were treated with anti-digoxigenin antibody coupled to alkaline phosphatase (Boehringer Mannheim), and the signal was detected with nitroblue tetrazolium and X-phosphate as the substrate. The coverslips were then mounted on slides and visualized under a phase-contrast microscope. Similarly treated mock-infected CV-1 cells served as the control. Mock-infected CV-1 cells (A) and HPIV3-infected cells (B) hybridized with antisense NP RNA (T7 transcript of NP cDNA clone in pGEM4Z) and mock infected CV-1 cells (C) and HPIV3-infected cells (D) hybridized with NP mRNA (SP6 transcript of NP clone in pGEM4Z) are shown.

Article Snippet: The fixed cells were stained with goat anti-HPIV3 antibody (Biodesign) and anti-goat IgG conjugated with biotin followed by FITC-conjugated avidin.

Techniques: Infection, Labeling, Microscopy

Journal:

Article Title: Involvement of Actin Microfilaments in the Replication of Human Parainfluenza Virus Type 3

doi:

Figure Lengend Snippet: Distribution of HPIV3-specific RNAs between the CSK and SOL fractions from infected CV-1 cells

Article Snippet: The fixed cells were stained with goat anti-HPIV3 antibody (Biodesign) and anti-goat IgG conjugated with biotin followed by FITC-conjugated avidin.

Techniques: Infection

Journal:

Article Title: Involvement of Actin Microfilaments in the Replication of Human Parainfluenza Virus Type 3

doi:

Figure Lengend Snippet: Inhibition of HPIV3 replication by CD. CV-1 cells were infected with HPIV3 at 5 PFU/cell in the presence of different concentrations of CD, as indicated. At 24 h post infection, the progeny virus released into the medium was collected and measured by plaque assay. CV-1 cells were infected with vesicular stomatitis virus (VSV) at 5 PFU/cell in the presence of CD. At 12 h postinfection, the progeny virus released into the medium was measured.

Article Snippet: The fixed cells were stained with goat anti-HPIV3 antibody (Biodesign) and anti-goat IgG conjugated with biotin followed by FITC-conjugated avidin.

Techniques: Inhibition, Infection, Plaque Assay

Journal:

Article Title: Involvement of Actin Microfilaments in the Replication of Human Parainfluenza Virus Type 3

doi:

Figure Lengend Snippet: Inhibition of intracellular RNP synthesis by CD. (A) CV-1 cells were infected with HPIV3 in the presence or absence of 8 mg of CD per ml. Intracellular RNP was isolated by the method described by Toneguzzo and Ghosh (43). The level of NP was determined by Western blot analysis with anti-RNP antibody. RNP; purified viral RNP was used as control. (B) Inhibition of mRNA synthesis by CD. CV-1 cells were infected with HPIV3 in the presence or absence of 8 mg of CD per ml. Total RNA was isolated from the cells, and the levels of NP mRNA were determined by Northern blot analysis with 32P-labeled NP cDNA. The same blot was hybridized with 32P-labeled glyceraldehyde-3-phosphate dehydrogenase (GAPDH) cDNA and used as an internal control.

Article Snippet: The fixed cells were stained with goat anti-HPIV3 antibody (Biodesign) and anti-goat IgG conjugated with biotin followed by FITC-conjugated avidin.

Techniques: Inhibition, Infection, Isolation, Western Blot, Purification, Northern Blot, Labeling

Journal:

Article Title: Involvement of Actin Microfilaments in the Replication of Human Parainfluenza Virus Type 3

doi:

Figure Lengend Snippet: Association of HPIV3 RNP with the CSK framework in vivo. CV-1 cells grown on coverslips were infected with HPIV3 at 1 PFU/cell, and at 12 h postinfection the cells were treated with CSK buffer containing rhodamine-conjugated phalloidin. The cells were washed with the same buffer followed by PBS and fixed. The fixed CSK structures were treated with anti-HPIV3 antibody that detects the RNP-associated proteins NP and P. The coverslips were then treated with biotin-conjugated anti-goat antibody followed by fluorescein-conjugated avidin. Similar staining of mock-infected cells served as control. Mock-infected (A) or HPIV3-infected (C) CV-1 cells treated with CSK buffer containing rhodamine-phalloidin and mock-infected (B) or HPIV3-infected (D) CV-1 cells treated with anti-HPIV3 antibody followed by biotin-conjugated anti-goat immunoglobulin plus fluorescein-conjugated avidin are shown.

Article Snippet: The fixed cells were stained with goat anti-HPIV3 antibody (Biodesign) and anti-goat IgG conjugated with biotin followed by FITC-conjugated avidin.

Techniques: In Vivo, Infection, Avidin-Biotin Assay, Staining

Journal:

Article Title: Involvement of Actin Microfilaments in the Replication of Human Parainfluenza Virus Type 3

doi:

Figure Lengend Snippet: Specificity of the interaction between the RNP and actin microfilaments. To determine the specificity of the RNP-actin interaction, the association of RNP with another cytoskeletal protein, tubulin, was examined. CV-1 cells, grown on coverslips, were infected with HPIV3 at 1 PFU/cell and at 12 h postinfection either incubated in PBS (A and B) or treated with CSK buffer (C and D) for 3 min on ice. The cells were washed with PBS and fixed. The coverslips were treated with anti-tubulin or anti-HPIV3 antibody. They were then treated with fluorescein-conjugated anti-mouse IgG for anti-tubulin (A and C) or biotin-conjugated anti-goat antibody followed by Texas red-conjugated avidin for HPIV3 antigens (B and D).

Article Snippet: The fixed cells were stained with goat anti-HPIV3 antibody (Biodesign) and anti-goat IgG conjugated with biotin followed by FITC-conjugated avidin.

Techniques: Infection, Incubation, Avidin-Biotin Assay

Journal:

Article Title: Involvement of Actin Microfilaments in the Replication of Human Parainfluenza Virus Type 3

doi:

Figure Lengend Snippet: Effect of CD on the distribution of RNP in vivo. CV-1 cells, grown on coverslips, were infected with HPIV3 at 1 PFU/cell in the presence of 8 μg of CD per ml. At 12 h postinfection, the cells were extracted with CSK buffer containing rhodamine-conjugated phalloidin. The CSK structures remaining on the coverslips were washed with the CSK buffer followed by PBS. The CSK structures were fixed and treated with anti-HPIV3 antibody followed by biotin-conjugated anti-goat Ig and fluorescein-conjugated avidin. Similarly treated HPIV3-infected CV-1 cells served as controls. Mock-infected (A) or HPIV3-infected (C) CV-1 cells treated with 8 μg of CD per ml were treated with CSK buffer containing rhodamine-conjugated phalloidin. Mock-infected (B) or HPIV3-infected (D) CV-1 cells were treated with anti-HPIV3 antibody followed by biotin-conjugated anti-goat Ig plus fluorescein conjugated avidin.

Article Snippet: The fixed cells were stained with goat anti-HPIV3 antibody (Biodesign) and anti-goat IgG conjugated with biotin followed by FITC-conjugated avidin.

Techniques: In Vivo, Infection, Avidin-Biotin Assay