Journal: Nutrients

Article Title: β-Cryptoxanthin Improves p62 Accumulation and Muscle Atrophy in the Soleus Muscle of Senescence-Accelerated Mouse-Prone 1 Mice

doi: 10.3390/nu12082180

Figure Lengend Snippet: Repression of muscle atrophy by β-cryptoxanthin in the soleus muscle of SAMP1 mice. ( A ) Immunofluorescent staining of laminin in the soleus muscle of the SAMR1 (R1), SAMP1-control (P1con), and SAMP1-CX (P1cx) groups. Scale bar: 100 μm. ( B ) Distribution of soleus muscle fiber cross-sectional area (CSA). ( C ) Average size of the soleus muscle fiber CSA. ( D ) Western blot analyses of MyHC type I and GAPDH. MyHC type I expression was normalized to anti-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) expression. Values are represented as mean ± standard deviation. n = 5–8 per group. * p < 0.05, statistical significance compared with the SAMP1-control group.

Article Snippet: Tissue homogenates and cell lysates were centrifuged at 20,000× g for 15 min, and the supernatants were subjected to sodium dodecyl sulfate–polyacrylamide gel electrophoresis, followed by Western blot analysis using the following antibodies: rabbit polyclonal beclin-1 (cell signaling; Danvers, MA, USA), anti-p62, anti-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) [ ], and anti-ubiquitin (cell signaling) antibodies; rabbit monoclonal anti-LC3 (clone D3U4C; cell signaling), anti-atrogin-1 (clone EPR9148(2); Abcam; Cambridge, UK), anti-p70S6K (clone 49D7; cell signaling), anti-phospho-p70S6K (clone 108D2; cell signaling), anti-AMPKα (clone 40H9; cell signaling), anti-mTOR (clone 7C10; cell signaling), and anti-phospho-mTOR (clone D9C2; cell signaling) antibodies; mouse monoclonal anti-β-actin (clone 2D4H5; cell signaling) and anti-myosin heavy chain (MyHC) (clone BA-D5 that recognizes MyHC type I; Developmental Studies Hybridoma Bank, University of Iowa, Iowa City, IA, USA) antibodies; and goat polyclonal anti-MuRF-1 antibody (R&D Systems, Minneapolis, MN, USA).

Techniques: Staining, Control, Western Blot, Expressing, Standard Deviation

Journal: Skeletal Muscle

Article Title: Elusive sources of variability of dystrophin rescue by exon skipping

doi: 10.1186/s13395-015-0070-6

Figure Lengend Snippet: Variability of dystrophin protein expression, as shown by IF after PMO injection. a Representative images of C57BL/10 (WT) and b PMO-treated mdx tibialis anterior sections stained for dystrophin. The WT control shows uniform IF staining for dystrophin. Insert at ×40 shows expected staining pattern for dystrophin-positive fibers. b PMO-treated mdx tibialis anterior shows a mosaic staining pattern and clustering of positive fibers. The yellow line represents the border between the tibialis anterior and EDL. Quantification was performed on the entire area of the muscle section. c Representative images of mouse mdx-6 showing variability between the muscles of the same animal. Images were selected to show positive fiber clustering and do not represent total area quantification. d IF quantification of diaphragm, gastrocnemius, heart, quadriceps, tibialis anterior, and triceps for all mice ( n = 6). e Geographic variability observed within the highly rescued triceps from mouse mdx-1. All tissues were sectioned (10-μm thick), stained, and probed with goat anti-rabbit IgG Alexa 594 antibody. Dystrophin-positive fibers were normalized to the area of the muscle section and the WT percentage of positive fibers. Original magnification for a, b, e = ×20; scale bar , 500 μm; for c = ×40; scale bar , 100 μm

Article Snippet: Muscle fiber types were identified using the following antibodies: mouse IgG2b monoclonal anti-type 1 MHC (clone BA-D5, 1:50), mouse IgG1 monoclonal anti-type 2a MHC (clone SC-71, 1:50), mouse IgM monoclonal anti-type 2b MHC (clone BF-F3, 1:5), and mouse IgG1 monoclonal anti-embryonic MHC (clone F1.652, 1:25), all obtained from the Developmental Studies Hybridoma Bank at the University of Iowa (Ames, IA, USA) [ ].

Techniques: Expressing, Injection, Staining, Control, Muscles

Journal: Cell

Article Title: Protection from SARS-CoV-2 Delta one year after mRNA-1273 vaccination in rhesus macaques coincides with anamnestic antibody response in the lung

doi: 10.1016/j.cell.2021.12.002

Figure Lengend Snippet: mRNA-1273 provides durable protection in the lower airway from B.1.617.2 (A and B) Representative images of lung samples 7 days after B.1.617.2 challenge from 4 NHPs that received mRNA-1273 (A) or mRNA control (B). Top row, detection of SARS-CoV-2 antigen by immunohistochemistry with a polyclonal anti-N antibody. Antigen-positive foci are marked by red arrows. Bottom row, hematoxylin and eosin (H&E) staining illustrating the extent of inflammation and cellular infiltrates. Images at 10× magnification with black bars for scale (100 μm). (C) SARS-CoV-2 antigen and inflammation scores in the left cranial (Lc) lobe, right middle (Rmid) lobe, and right caudal (Rc) lobe of the lungs 7 days after B.1.617.2 challenge. Antigen scoring legend: –, no antigen detected; +/−, rare to occasional foci; +, occasional to multiple foci; ++, multiple to numerous foci; +++, numerous foci. Inflammation scoring legend: –, minimal to absent inflammation; +/−, minimal to mild inflammation; +, mild to moderate inflammation; ++, moderate to severe inflammation; +++, severe inflammation. Horizontal rows correspond to individual NHPs depicted above (A and B).

Article Snippet: Goat anti-human IgD-FITC (polyclonal) , Southern Biotech , Cat#2030-02; RRID: AB_2795624.

Techniques: Control, Immunohistochemistry, Staining

Journal: Cell

Article Title: Protection from SARS-CoV-2 Delta one year after mRNA-1273 vaccination in rhesus macaques coincides with anamnestic antibody response in the lung

doi: 10.1016/j.cell.2021.12.002

Figure Lengend Snippet:

Article Snippet: Goat anti-human IgD-FITC (polyclonal) , Southern Biotech , Cat#2030-02; RRID: AB_2795624.

Techniques: Labeling, Virus, Neutralization, Recombinant, Transfection, Staining, Multiplex Assay, Diagnostic Assay, Software

Journal: Frontiers in Plant Science

Article Title: Transient Expression of Tetrameric Recombinant Human Butyrylcholinesterase in Nicotiana benthamiana

doi: 10.3389/fpls.2016.00743

Figure Lengend Snippet: Map of gene constructs. (A) pTRBO-prBChE-KDEL, (B) pTRBO-prBChE, and (C) p35S-P19, 35S: Cauliflower Mosaic Virus (CaMV) promotor, RAmy3DSP : rice alpha-amylase 3D gene signal peptide, Replicase : replicase gene of tobacco mosaic virus (TMV), MP : movement protein, BChE : codon optimized sequences for the human butyrylcholinesterase gene. 3XFLAG : specific amino acid codon sequences used for immunodetection and purification of the protein. KDEL : Codons encoding lysine, aspartic acid, glutamic acid, leucine, specifying the endoplamsmic retention sequence. P19: P19 gene from Tomato Bushy Stunt Virus (TBSV).

Article Snippet: The blots were developed with either 1:2,500 dilution of monoclonal anti-FLAG M2-Peroxidase (HRP) antibody (Sigma–Aldrich, St. Louis, MO, USA) in 5% NFDM solution or with 1:200 mouse monoclonal anti-BChE antibody (D-5; Santa Cruz biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution followed by 1:2000 goat anti-mouse HRP conjugated secondary antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution.

Techniques: Construct, Virus, Immunodetection, Purification, Sequencing

Journal: Frontiers in Plant Science

Article Title: Transient Expression of Tetrameric Recombinant Human Butyrylcholinesterase in Nicotiana benthamiana

doi: 10.3389/fpls.2016.00743

Figure Lengend Snippet: (A) SDS-PAGE and (B) Western blot of ER retained and apoplast targeted prBChE protein extracted using different extraction buffers. (A) Coomassie stained gel with 15 mU: lane1: (prBChE-ER pH 4 extract), lane 2: (prBChE-ER pH 8 extract), lane 3 (prBChE pH 4 extract), lane 4 (prBChE pH 8 extract), lane 5: (3 μg of equine BChE control) loaded under non-reduced conditions. Lane M shows the pre-stained protein molecular weight standards along with the molecular weight in kDa. (B) Western blot analysis using 1:200 mouse anti-BChE antibody and 1:2,000 goat anti-mouse HRP conjugated antibody.

Article Snippet: The blots were developed with either 1:2,500 dilution of monoclonal anti-FLAG M2-Peroxidase (HRP) antibody (Sigma–Aldrich, St. Louis, MO, USA) in 5% NFDM solution or with 1:200 mouse monoclonal anti-BChE antibody (D-5; Santa Cruz biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution followed by 1:2000 goat anti-mouse HRP conjugated secondary antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution.

Techniques: SDS Page, Western Blot, Extraction, Staining, Control, Molecular Weight

Journal: Frontiers in Plant Science

Article Title: Transient Expression of Tetrameric Recombinant Human Butyrylcholinesterase in Nicotiana benthamiana

doi: 10.3389/fpls.2016.00743

Figure Lengend Snippet: (A) SDS-PAGE and (B) Western blot of purified prBChE protein compared to serial dilutions of PEG-rBChE protein control. (A) Coomassie stained gel: lanes 1 and 2 contain duplicate loadings of 7 μg of prBChE protein. Lanes 3 to 6 contain serial dilutions of PEG-rBChE control (lane 3: 1, lane 4: 3, lane 5: 7, and lane 6: 10 μg), all loaded under reduced conditions. Lane M shows the pre-stained protein molecular weight standards along with the molecular weight in kDa. Because the goat recombinant BChE is PEGylated, its monomeric mobility on SDS-PAGE is approximately 200 kDa. However, the unPEGylated goat BChE co-migrates with prBChE as revealed by SDS-PAGE. (B) Western blot analysis using 1:200 mouse anti-BChE antibody and 1:2,000 goat anti-mouse HRP conjugated antibody.

Article Snippet: The blots were developed with either 1:2,500 dilution of monoclonal anti-FLAG M2-Peroxidase (HRP) antibody (Sigma–Aldrich, St. Louis, MO, USA) in 5% NFDM solution or with 1:200 mouse monoclonal anti-BChE antibody (D-5; Santa Cruz biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution followed by 1:2000 goat anti-mouse HRP conjugated secondary antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution.

Techniques: SDS Page, Western Blot, Purification, Control, Staining, Molecular Weight, Recombinant

Journal: Frontiers in Plant Science

Article Title: Transient Expression of Tetrameric Recombinant Human Butyrylcholinesterase in Nicotiana benthamiana

doi: 10.3389/fpls.2016.00743

Figure Lengend Snippet: Initial reaction velocity kinetics of BTCh hydrolysis by BChE enzyme. The hydrolysis rate of BTCh substrate was monitored by measuring BChE activity (prBChE vs. human BChE control) over a large substrate concentration (10 μM–7.5 mM) in 0.1 M phosphate buffer pH 7.4, 0.267 mM DNTB.

Article Snippet: The blots were developed with either 1:2,500 dilution of monoclonal anti-FLAG M2-Peroxidase (HRP) antibody (Sigma–Aldrich, St. Louis, MO, USA) in 5% NFDM solution or with 1:200 mouse monoclonal anti-BChE antibody (D-5; Santa Cruz biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution followed by 1:2000 goat anti-mouse HRP conjugated secondary antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution.

Techniques: Activity Assay, Control, Concentration Assay

Journal: Frontiers in Plant Science

Article Title: Transient Expression of Tetrameric Recombinant Human Butyrylcholinesterase in Nicotiana benthamiana

doi: 10.3389/fpls.2016.00743

Figure Lengend Snippet: Oligomer distribution analysis by Western blot using (A) non-reducing and (B) reducing conditions SDS-PAGE analysis. Western blot analysis was developed using 1:200 mouse anti-BChE antibody and 1:2,000 goat anti-mouse HRP conjugated antibody of 0.25 μg of equine BuChE control (lane 1) and 0.25 μg pure prBChE protein (lane 2). Lane M shows the pre-stained protein molecular weight standards along with the molecular weight in kDa.

Article Snippet: The blots were developed with either 1:2,500 dilution of monoclonal anti-FLAG M2-Peroxidase (HRP) antibody (Sigma–Aldrich, St. Louis, MO, USA) in 5% NFDM solution or with 1:200 mouse monoclonal anti-BChE antibody (D-5; Santa Cruz biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution followed by 1:2000 goat anti-mouse HRP conjugated secondary antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution.

Techniques: Western Blot, SDS Page, Control, Staining, Molecular Weight

Journal: Frontiers in Plant Science

Article Title: Transient Expression of Tetrameric Recombinant Human Butyrylcholinesterase in Nicotiana benthamiana

doi: 10.3389/fpls.2016.00743

Figure Lengend Snippet: (A) Coomassie stained, (B) Western blot, and (C) activity stained Native gel analysis of prBChE, compared to BChE control. (A,B) Coomassie stained and Western blot 7.5% gel with 3 μg of purified prBChE (lane 3) compared to BChE mammalian serum enzymes (1.5 μg of human protein: lane 1, 3 μg equine protein: lane 2). (C) Activity stained of 0.25 mU pure prBChE (lane 2) and 0.25 mU equine control (lane 1).

Article Snippet: The blots were developed with either 1:2,500 dilution of monoclonal anti-FLAG M2-Peroxidase (HRP) antibody (Sigma–Aldrich, St. Louis, MO, USA) in 5% NFDM solution or with 1:200 mouse monoclonal anti-BChE antibody (D-5; Santa Cruz biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution followed by 1:2000 goat anti-mouse HRP conjugated secondary antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution.

Techniques: Staining, Western Blot, Activity Assay, Control, Purification

Journal: Frontiers in Plant Science

Article Title: Transient Expression of Tetrameric Recombinant Human Butyrylcholinesterase in Nicotiana benthamiana

doi: 10.3389/fpls.2016.00743

Figure Lengend Snippet: (A) Coomassie stained and (B) Western blot of Native gel analysis of denatured prBChE, compared to denatured equine BChE control. (A) Coomassie stained gel, lane 1: 1 μg equine BChE control under native conditions, lane 2: 1 μg of denatured equine BChE control under reducing conditions, lane 3: 1 μg prBChE under native conditions, lane 4: 1 μg of denatured prBChE under reducing conditions. (B) Western blot analysis developed with 0.5 μg equine BChE control and prBChE using 1:200 mouse anti-BChE antibody and 1:2,000 goat anti-mouse HRP conjugated antibody.

Article Snippet: The blots were developed with either 1:2,500 dilution of monoclonal anti-FLAG M2-Peroxidase (HRP) antibody (Sigma–Aldrich, St. Louis, MO, USA) in 5% NFDM solution or with 1:200 mouse monoclonal anti-BChE antibody (D-5; Santa Cruz biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution followed by 1:2000 goat anti-mouse HRP conjugated secondary antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution.

Techniques: Staining, Western Blot, Control

Journal: Frontiers in Plant Science

Article Title: Transient Expression of Tetrameric Recombinant Human Butyrylcholinesterase in Nicotiana benthamiana

doi: 10.3389/fpls.2016.00743

Figure Lengend Snippet: (A) Western blot analysis of a native gel of crude preparation of prBChE. Western blot analysis was performed using 1:200 mouse anti-BChE antibody and 1:2,000 goat anti-mouse HRP conjugated antibody of crude extract preparation of 0.25 μg prBChE (lane 3) compared to 0.25 μg of equine control (lane 1) and 0.25 μg pure prBChE protein (lane2). (B) Native gel analysis of different prBChE variants. 45 mU of prBChE-ER (lane 1), prBChE (lane 2), prBChE-AWF (lane 3) and equine control (lane 4) were loaded to a 7.5% gel and stained for BChE activity according to the method of .

Article Snippet: The blots were developed with either 1:2,500 dilution of monoclonal anti-FLAG M2-Peroxidase (HRP) antibody (Sigma–Aldrich, St. Louis, MO, USA) in 5% NFDM solution or with 1:200 mouse monoclonal anti-BChE antibody (D-5; Santa Cruz biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution followed by 1:2000 goat anti-mouse HRP conjugated secondary antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA) in 5% NFDM solution.

Techniques: Western Blot, Control, Staining, Activity Assay