Journal: Neural Regeneration Research

Article Title: The circ_0002538/miR-138-5p/plasmolipin axis regulates Schwann cell migration and myelination in diabetic peripheral neuropathy

doi: 10.4103/1673-5374.355979

Figure Lengend Snippet: Characterization of circ_0002538 and its function in SCs. (A) Hierarchical clustering analyses of DEcircRNAs ( n = 3). (B, C) RT-PCR verified five circRNAs with upregulated expression and six circRNAs with downregulated expression, and the results were consistent with the RNA-seq data ( n = 12). The red dotted box highlights the circRNA of interest. Y -axis: Fold changes in circRNA expression compared with the non-diabetic group. * P < 0.05, ** P < 0.01, vs . non-diabetic group (independent-sample t-test). (D) Schematic diagram showing that circ_0002538 was formed by the circularization of KLHL8 exon 2. The red arrow represents the “head-to-tail” splicing site of circ_0002538, confirmed by Sanger sequencing. (E) We used divergent primers and convergent primers to amplify circ_0002538 in cDNA and gDNA. We used β-actin as a negative control. (F) circ_0002538 and KLHL8 mRNA in SCs were detected via RT-PCR after incubation with or without RNase R. Y -axis: fold changes in RNA expression compared with the mock group. *** P < 0.001, vs . mock group (independent-sample t -test). (G) circ_0002538 and KLHL8 mRNA levels were evaluated in the sh-circ_0002538-transfected SCs via RT-PCR. Y -axis: fold changes in RNA expression compared with the sh-NC group. *** P < 0.001, vs . sh-NC group. (H, I) The migrating number of SCs in the sh-circ_0002538 group was lower than that in the sh-NC group in the Transwell assays. ** P < 0.01, vs . sh-NC group (independent-sample t -test). Scale bars: 100 μm. All bar graphs represent the average of at least three independent replicates, and the error bars are the SD. cDNA: Complementary DNA; DPN: diabetic peripheral neuropathy; gDNA: genomic DNA; KLHL8: Kelch-like family member 8; sh-circ_0002538: short hairpin RNA for circ_0002538; sh-NC: normal control for short hairpin RNA.

Article Snippet: To detect the combination of circRNAs and miRNAs, we performed RNA pulldown assays with biotinylated probes according to the manufacturer’s protocol (MCE, Shanghai, China, Cat# HY-K0208).

Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, RNA Sequencing, Sequencing, Negative Control, Incubation, RNA Expression, Transfection, shRNA, Control

Journal: Journal of proteome research

Article Title: Assessment of Streptavidin Bead Binding Capacity to Improve Quality of Streptavidin-based Enrichment Studies

doi: 10.1021/acs.jproteome.0c00772

Figure Lengend Snippet: Product Specifications for Various Streptavidin Bead Products Used in Proteomics Workflows a

Article Snippet: Vector Laboratories , MagnaLINK streptavidin magnetic beads , SPM , M-1003–010 , 2.8 , 60% magnetite content , free biotin: ≥10 nmol/mg biotinylated IgG: ≥0.8 nmol/mg biotinylated oligonucleotides: ≥0.75 nmol/mg , fluorescein-biotin assay.

Techniques: Binding Assay, Magnetic Beads, Plasmid Preparation

Journal: Journal of proteome research

Article Title: Assessment of Streptavidin Bead Binding Capacity to Improve Quality of Streptavidin-based Enrichment Studies

doi: 10.1021/acs.jproteome.0c00772

Figure Lengend Snippet: Product Specifications for Various Streptavidin Bead Products Used in Proteomics Workflows a

Article Snippet: Vector Laboratories , MagnaLINK streptavidin magnetic beads , SPM , M-1003–010 , 2.8 , 60% magnetite content , free biotin: ≥10 nmol/mg biotinylated IgG: ≥0.8 nmol/mg biotinylated oligonucleotides: ≥0.75 nmol/mg , fluorescein-biotin assay.

Techniques: Binding Assay, Magnetic Beads, Plasmid Preparation

Journal: Journal of proteome research

Article Title: Assessment of Streptavidin Bead Binding Capacity to Improve Quality of Streptavidin-based Enrichment Studies

doi: 10.1021/acs.jproteome.0c00772

Figure Lengend Snippet: Categorization of Streptavidin Bead Products Used in This Study Based on Their Behavior in Binding Buffer

Article Snippet: Vector Laboratories , MagnaLINK streptavidin magnetic beads , SPM , M-1003–010 , 2.8 , 60% magnetite content , free biotin: ≥10 nmol/mg biotinylated IgG: ≥0.8 nmol/mg biotinylated oligonucleotides: ≥0.75 nmol/mg , fluorescein-biotin assay.

Techniques: Binding Assay, Magnetic Beads

Journal: Journal of proteome research

Article Title: Assessment of Streptavidin Bead Binding Capacity to Improve Quality of Streptavidin-based Enrichment Studies

doi: 10.1021/acs.jproteome.0c00772

Figure Lengend Snippet: General considerations for evaluating the fit-for-purpose of streptavidin bead products and reporting key parameters to promote repeatability among studies. Key features to consider when implementing streptavidin beads into a sample preparation workflow are shown and include physical bead characteristics, bead handling characteristics and performance, and whether manual or automated sample processing is required. Technical details that should be included when reporting data from studies that implement streptavidin-bead-based enrichment are indicated with a black star.

Article Snippet: Vector Laboratories , MagnaLINK streptavidin magnetic beads , SPM , M-1003–010 , 2.8 , 60% magnetite content , free biotin: ≥10 nmol/mg biotinylated IgG: ≥0.8 nmol/mg biotinylated oligonucleotides: ≥0.75 nmol/mg , fluorescein-biotin assay.

Techniques: Sample Prep

Journal: Journal of proteome research

Article Title: Assessment of Streptavidin Bead Binding Capacity to Improve Quality of Streptavidin-based Enrichment Studies

doi: 10.1021/acs.jproteome.0c00772

Figure Lengend Snippet: Intralot variation in binding capacity of streptavidin bead results in differences in number of proteins identified in a CSC experiment. (A) Total number of proteins and cell surface N-glycoproteins identified containing consensus motif (NxS/T/V/c, where x ≠ P) in CSC experiments (1000 μg total peptide) when using 100 μL each of two different lots of GenScript streptavidin MagBeads (C44251904 and C44241809). (B) Binding capacity as reported by GenScript. The orange dashed line highlights the minimum binding capacity necessary to pass quality control (60 nmol/mL). (C) Peak area replicate comparison view for precursors M, M+1, and M+2 for streptavidin peptides YDSAPATDGSGTALGWTVAWK, NAHSATTWSGQYVGGAEAR, INTGWLLTSGTTEANAWK, and STLVGHFTFTK (technical replicates = 4) detected in the CSC results from panel A.

Article Snippet: Vector Laboratories , MagnaLINK streptavidin magnetic beads , SPM , M-1003–010 , 2.8 , 60% magnetite content , free biotin: ≥10 nmol/mg biotinylated IgG: ≥0.8 nmol/mg biotinylated oligonucleotides: ≥0.75 nmol/mg , fluorescein-biotin assay.

Techniques: Binding Assay

Journal: Journal of proteome research

Article Title: Assessment of Streptavidin Bead Binding Capacity to Improve Quality of Streptavidin-based Enrichment Studies

doi: 10.1021/acs.jproteome.0c00772

Figure Lengend Snippet: AVIDITY assay is compatible with a range of streptavidin bead products. (A) HABA titration curves for GenScript streptavidin MagBeads (lot C44261906), Cytiva Sera-Mag Magnetic neutravidin-coated particles (lot 17015347), and New England Biolabs streptavidin magnetic beads (lot 10051761). (B) Absorbances of free HABA of five different bead types at 25 nmol of HABA are shown as means with SD error bars (n = 3). An absorbance range of 0.9 to 1.5 was suitable for all beads tested. (C) Biotin titration curves for two different lots of GenScript streptavidin MagBeads (lots C44261906 and C44242003) and one lot of Cytiva Sera-Mag SpeedBeads neutravidin-coated magnetic beads (lot 17015347). Changes in absorbance (Δ absorbance) as determined by subtracting the control absorbance reading (beads incubated with HABA) from the experimental absorbance reading (beads incubated with HABA and d-biotin) are shown as means with SD error bars (n = 3).

Article Snippet: Vector Laboratories , MagnaLINK streptavidin magnetic beads , SPM , M-1003–010 , 2.8 , 60% magnetite content , free biotin: ≥10 nmol/mg biotinylated IgG: ≥0.8 nmol/mg biotinylated oligonucleotides: ≥0.75 nmol/mg , fluorescein-biotin assay.

Techniques: Titration, Magnetic Beads, Incubation

Journal: Journal of proteome research

Article Title: Assessment of Streptavidin Bead Binding Capacity to Improve Quality of Streptavidin-based Enrichment Studies

doi: 10.1021/acs.jproteome.0c00772

Figure Lengend Snippet: Binding capacity of streptavidin beads as measured by the AVIDITY assay is predictive of proteomics results. (A) Total numbers of proteins and cell surface N-glycoproteins identified containing the consensus motif (NxS/T/V/c, where x ≠ P) in CSC experiments (1000 μg of total peptide) when using 100 μL each of two different lots of GenScript streptavidin MagBeads and one lot each for Cytiva Sera-Mag SpeedBeads neutravidin-coated magnetic beads (lot 17015347), Cytiva Sera-Mag streptavidin-coated magnetic beads (lot 17013370), and Invitrogen Dynabeads MyOne streptavidin C1 (lot 00945048) are shown. (B) Binding capacity as determined by the AVIDITY assay using 25 nmol HABA and 100 nmol d-biotin shown as means with SD error bars (n = 3).

Article Snippet: Vector Laboratories , MagnaLINK streptavidin magnetic beads , SPM , M-1003–010 , 2.8 , 60% magnetite content , free biotin: ≥10 nmol/mg biotinylated IgG: ≥0.8 nmol/mg biotinylated oligonucleotides: ≥0.75 nmol/mg , fluorescein-biotin assay.

Techniques: Binding Assay, Magnetic Beads