Journal: iScience

Article Title: tANCHOR-cell-based assay for monitoring of SARS-CoV-2 neutralizing antibodies rapidly adaptive to various receptor-binding domains

doi: 10.1016/j.isci.2024.109123

Figure Lengend Snippet:

Article Snippet: Rabbit anti-V5 , Novus Biologicals , Cat# NB600-381.

Techniques: Virus, Recombinant, Saline, Staining, Isolation, Protease Inhibitor, Software, Cell Culture, Enzyme-linked Immunosorbent Assay

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Synapse-specific catecholaminergic modulation of neuronal glutamate release.

doi: 10.1073/pnas.2420496121

Figure Lengend Snippet: Fig. 3. Type I MNs OA receptors include OAMB, which is enriched in the Ib MN and needed to boost locomotion. (A) RNAseq shows expression of five OA receptors in type I MNs. The dashed line shows the maximum normalized counts of any gene observed in attp2 controls. Error bars are ± S.E.M (B) Example image of an NMJ showing a maximum projection confocal image of OAMB receptor staining (green) in a Ib and Is MN pair. Animals had the OAMB receptor CRISPR tagged with 10 copies of the V5 epitope. Staining was against the V5 epitope. (Scale bar, 5 μm.) (C) Bar graph shows the mean ± 95% CI of the mean OAMB staining levels (A.U. arbitrary units) per Ib and Is MNs. Each Ib and Is were paired on the same muscle (10 NMJs, 1 per animal). (**** P <0.0001 by the paired t-test). (D) Mean velocity (mm/s) of control larvae (attp2) (black, n = 15) and animals with OAMB knocked down in type I MNs (oambRNAi; blue, n = 14) crawling in the dark (black bar) followed by under 505 nm light (blue bar). Recording starts at 5 min., following an adjustment period. (E) Mean total displacement for each animal in (D). Attp2 (black, n = 15 animals), oambRNAi (blue, n =14 animals). Red points are mean ± S.E.M. (*** P <0.001; ** P <0.01; n.s., not significant by Student’s paired t-test).

Article Snippet: For OAMB imaging, rat anti- V5 epitope (Novus NBP2- 81037) was used at 1:500.

Techniques: Expressing, Staining, CRISPR, Control

Journal: Retrovirology

Article Title: The HIV-1 late domain-2 S 40 A polymorphism in antiretroviral (or ART)-exposed individuals influences protease inhibitor susceptibility

doi: 10.1186/s12977-016-0298-1

Figure Lengend Snippet: Trans cleavage at the proximal site in the presence of IDV. a Schematic drawing of a model precursor substrate and expected products of trans processing by enzymatically active miniprecursors. b , c Quantification of trans processing product and p6*-PR b enzyme. Band intensity was used to reflect trans processing efficiency and enzyme detection levels. d – g Western blot analyses of the model substrate and four fusion precursors with increasing IDV. Post-nuclear total lysates were resolved on SDS-PAGE and transferred to a PVDF membrane. Each blot was simultaneously probed with mouse HA ( top ) or rabbit V5 ( bottom ) antibodies. The blots were then probed with IR800 goat anti-mouse secondary and IR700 goat anti-rabbit antibody and scanned using a LI-COR Odyssey scanner. The images shown are representative of four experiments

Article Snippet: Fusion precursors and the autoprocessing products were detected using antibodies raised against the HA tag (H9658, Sigma), the V5 tag (600-401-378, Rockland Laboratory) or the FLAG (F1804, Sigma).

Techniques: Western Blot, SDS Page, Membrane

Journal: Science signaling

Article Title: ATM directs DNA damage responses and proteostasis via genetically separable pathways

doi: 10.1126/scisignal.aan5598

Figure Lengend Snippet: (A) In vitro kinase assay with immunoprecipitated CK2. HEK-293T cells stably expressing V5-tagged CK2β were treated with 10 μM ATM inhibitor (KU-55933) or an equivalent amount of DMSO for 16 hours and CK2β was immunoprecipitated with magnetic anti-V5 beads. CK2 was incubated with 1.67 mM ATP, 5 fCi ATP-γ-32P, and 1.15 μg of GST-CK2 substrate for 1 hour, and 32P-labeled substrate was analyzed by phosphorimager. (B) Normalized amount of immunoprecipitated CK2β-V5 from (A) with the levels of CK2β-V5 normalized to levels in mock-treated cells. Quantitation was performed on the Licor system using Image Studio Ver 4.0. P-values: * < 0.05 (C-E) Distribution profile of CK2 subunits CK2α (C), CK2α′ (D), and CK2β (E), in HEK-293T cells after sucrose gradient sedimentation. HEK-293T cells expressing CK2α-V5, CK2α′-V5, or CK2β-V5 were treated with 10 μM KU-55933 or an equivalent amount of DMSO for 16 hours, harvested, and lysed in the absence of detergent. 1 mg of lysate in 500 μl lysis buffer total was added to the top of a sucrose gradient made with 1 ml layers of 50% to 5% sucrose in 5% increments. After ultracentrifugation, 500 μl fractions were collected and analyzed by western blot. (F and G) the distribution profile of CK2α (F) and CK2β (G) from the AT1ABR cells expressing wild-type or CL ATM alleles, analyzed by sucrose gradient sedimentation. Lysates were analyzed as in (C-E) except with induced AT1ABR cells expressing wild-type or CL alleles of ATM as indicated. (H) Sucrose gradient sedimentation pattern of V5-tagged CK2β stably expressed in U2OS cells treated with 10 μM KU-55933 or an equivalent amount of DMSO for 16 hours, or treated with a combination of KU-55933 and 1 mM NAC, as indicated. (I) Analysis of detergent-resistant aggregates in U2OS cells treated with 10 μM KU-55933 or DMSO, or a combination of KU-55933 and 1 mM NAC, as indicated. Aggregate fractions were isolated and compared with total lysate using western blot analysis for stably expressed V5-tagged CK2β. (J) Analysis of detergent-resistant aggregates in U2OS Flp-In cells expressing wt, CL, or 2RA ATM treated with 25 μM arsenite as shown in (I). (K) Aggregate fractions were isolated from U2OS cells expressing shCTRL or ATM shRNA and probed for CK2β, ATM, and β-actin as indicated from two independent cultures. (L) Means ± S.D. of CK2β abundance in aggregates were quantified from 3 independent experiments [including the replicates shown in (K)]. ** p< 0.005.

Article Snippet: V5-tagged CK2 levels were probed using rabbit anti-V5 (Novus Biologicals, NB600-381) and Alexa Fluor 680 goat anti-rabbit (Invitrogen, {"type":"entrez-nucleotide","attrs":{"text":"A21076","term_id":"579037","term_text":"A21076"}} A21076 ) and scanned and quantitated on the Li-Cor Odyssey system.

Techniques: In Vitro, Kinase Assay, Immunoprecipitation, Stable Transfection, Expressing, Incubation, Labeling, Quantitation Assay, Sedimentation, Lysis, Western Blot, Isolation, shRNA