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Anastasis

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Most cited protocols related to «Anastasis»

1
Genetic Screening for dVCP Modifiers
Deficiency (Df) lines for all four chromosomes obtained from the Bloomington stock center were used to identify dominant modifiers of mutant dVCP in a genetic screen. For the primary screen, balanced virgin female dVCP R152H (recombined with Gmr GAL4) flies were crossed with Df/Balancer males from 270 deficiency lines and progeny were examined for changes in eye phenotype (including color, ommatidia structure and bristle formation). At least ten progeny were examined and scored on a twenty-point scale. Eyes were examined for the presence of: supernumerary inter-ommatidial bristles (IOBs), IOBs with abnormal orientation, necrotic patches, a decrease in size, retinal collapse, fusion of ommatidia, disorganization of ommatidial array and loss of pigmentation. Points were added if: there was complete loss of IOBs (+1), more than 3 small or 1 large necrotic patch (+1), retinal collapse extended to the midline of the eye (+1) or beyond (+2), loss of ommatidial structure in less than 50% (+1) or more than 50% (+2) of the eye, and if pigmentation loss resulted in change of eye color from red to orange (+1) or pale orange/white (+2). Gmr, GAL4, UAS dVCP R152H / Balancer served as an internal control. In a secondary screen to filter nonspecific modifiers of cell death, deficiencies defined as hits (either enhancing or suppressing the dVCP mutant phenotype) were then crossed with flies expressing the pro-apoptotic gene Reaper (recombined with Gmr GAL4). Any hits that similarly affected dVCP R152H and Reaper in the secondary screen were excluded from further study due to the possibility of non–specific anti-apoptotic effects. As regions of interest were identified from the primary and secondary screens, additional Df lines were obtained that overlapped with interacting deficiencies to verify and refine the position of potential modifiers. For the final step of gene identification, individual RNAi lines corresponding to the genes within the candidate intervals were obtained from the Vienna Drosophila RNAi Center. Gmr GAL4, UAS dVCP R152H females were crossed with males from the RNAi lines and the progeny eyes were evaluated for changes. A modifier was defined as an RNAi line that replicated the enhancement or suppression of the corresponding deficiency.
Ritson G.P., Custer S.K., Freibaum B.D., Guinto J.B., Geffel D., Moore J., Tang W., Winton M.J., Neumann M., Trojanowski J.Q., Lee V.M., Forman M.S, & Taylor J.P. (2010). TDP-43 Mediates Degeneration in a Novel Drosophila Model of Disease Caused by Mutations in VCP/p97. The Journal of Neuroscience, 30(22), 7729-7739.
Publication 2010
Anastasis Apoptosis Cell Death Chromosomes Diptera Drosophila Eye Eye Color Females Genes Males Necrosis Phenotype Pigmentation Retina RNA Interference Screenings, Genetic Shock
2
Improved EBV Detection Protocol
To maximise detection rates and reduce false negative results, two primer sets targeting the highly conserved EBV regions, EBNA-1 and BHRF-1, were used for PCR amplification (Table 1). EBNA-1 is a latent protein required for replication and genome maintenance and is the only viral protein consistently expressed in EBV-infected cells [32 (link),33 (link)]. BHRF-1 is expressed in lytic infection and confers anti-apoptotic properties similar to Bcl-2 for enhancing cell survival [34 (link)]. Groups 2-5 were evaluated by both PCR targets, while inadequate sample volume limited testing to EBNA-1 in Group 1. The beta-globin gene targeting the TAL57 region was used as a 'house-keeping' gene to control for PCR inhibitors and check for DNA integrity [35 (link)]. All samples were subjected to beta-globin PCR prior to EBV QPCR. Contamination was monitored by the use of PCR-grade water and no template DNA controls.
Lay M.L., Lucas R.M., Ratnamohan M., Taylor J., Ponsonby A.L, & Dwyer D.E. (2010). Measurement of Epstein-Barr virus DNA load using a novel quantification standard containing two EBV DNA targets and SYBR Green I dye. Virology Journal, 7, 252.
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Publication 2010
Anastasis BCL2 protein, human beta-Globins Cells Cell Survival DNA Replication EBV-encoded nuclear antigen 1 Genes Genes, Housekeeping Genome Infection inhibitors Oligonucleotide Primers Proteins Viral Proteins
3
Benchmarking PELE's Computational Efficiency
Although this article aims to introduce the PELE web server, we will give here a short description of previous benchmark studies. Several applications have described exit and entry pathways, comparing its accuracy with experimental data. In our first study (14 ), we obtained the migration for Cytochrome P450 camphor, myoglobin and the fatty acid-binding proteins. Further studies compared PELE’s migration pathways in truncated haemoglobin with kinetic experimental data for the wild-type and the W8F mutant (19 (link)). More recently, a comprehensive study on 97 difficult induced fit cases, including cross- and apo- docking, has shown the capabilities of PELE in docking refinement (15 (link)). Furthermore, the study underlined the goodness of the OPLS interaction energy when scoring docking poses within one ligand. Binding site search and induced fit docking has been performed, in collaboration with experimental studies, in aryl-alcohol oxidase (20 (link),21 (link)), in mTOR kinases (22 (link)), in anti-apoptotic Bcl-2 receptors (23 (link)) and in different globins (10 ). The mTOR and Bcl-2 studies illustrate the combination of PELE with docking scoring techniques to discriminate different ligands. At the level of protein dynamics, in absence of ligand, we have compared PELE’s conformational search with microsecond MD simulations in ubiquitin and with metadynamics calculations on T4 lysozyme (16 ). The results clearly show a good agreement in the sampling of PELE with that of more sophisticated simulations.
Madadkar-Sobhani A, & Guallar V. (2013). PELE web server: atomistic study of biomolecular systems at your fingertips. Nucleic Acids Research, 41(Web Server issue), W322-W328.
Publication 2013
Anastasis aryl-alcohol oxidase bcl-2 Gene Binding Sites Camphor Cytochrome P450 Fatty Acid-Binding Proteins FRAP1 protein, human Globin Kinetics Ligands Muramidase Myoglobin Phosphotransferases Proteins Truncated Hemoglobins Ubiquitin
4
Evaluating Apoptotic Markers in HepG2 Cells
The levels of the apoptotic markers caspase-3, caspase-9, p53 and Bax, as well as the anti-apoptotic marker Bcl-2 were evaluated by the use of ELISA colorimetric kits per the manufacturer’s instructions, as reported earlier28 ,29 . Furthermore, the pro-apoptotic potential of spirooxindole 6a towards HepG2 cells was assessed using FITC Annexin V Apoptosis Detection Kit by flow cytometry, according to the manufacturer’s protocol and referring to the reported procedures30 ,31 .
Al-Rashood S.T., Hamed A.R., Hassan G.S., Alkahtani H.M., Almehizia A.A., Alharbi A., Al-Sanea M.M, & Eldehna W.M. (2020). Antitumor properties of certain spirooxindoles towards hepatocellular carcinoma endowed with antioxidant activity. Journal of Enzyme Inhibition and Medicinal Chemistry, 35(1), 831-839.
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Publication 2020
Anastasis Apoptosis bcl-2 Gene Caspase 3 Caspase 9 Colorimetry Enzyme-Linked Immunosorbent Assay FITC-annexin A5 Flow Cytometry Hep G2 Cells
5
Recombinant Human CD5L Production
The cDNA of human CD5L (hsCD5L) was obtained by gene synthesis (GenScript) following NCBI reference sequence NP_005885.1, with a modification in which the immunoglobulin g chain signal peptide replaced that of CD5L. The cDNA was cloned into the p.evi vector and transiently transfected into CHO K1 cells using the eviFect system (Evitria AG). Cells were grown in eviMake, a chemically defined, serum-free, animal component-free medium. The cell culture supernatant fraction was harvested at d 8 after transfection, dialyzed to 20 mM Na2HPO4, pH 7.4 and subjected to MonoQ chromatography. Recombinant human CD5L (r-HsCD5L) was eluted in a sodium chloride gradient, and purification was monitored by SDS-PAGE. Purified protein was dialyzed to PBS, concentrated by centrifugation on Amicon ultra (Millipore, UFC901024), and possible endotoxin contamination was removed by Endotrap columns (Hyglos GmbH, 321063), following the manufacturer´s protocol and as performed before.5 (link) As mentioned above, the purified r-HsCD5L was tested in preliminary experiments, where its activity in terms of MФ TNF secretion, as well as inhibition of apoptosis induced by cycloheximide treatment, was comparable to that of commercially available r-HsCD5L (R&D Systems) (data not shown).
Sanjurjo L., Amézaga N., Aran G., Naranjo-Gómez M., Arias L., Armengol C., Borràs F.E, & Sarrias M.R. (2015). The human CD5L/AIM-CD36 axis: A novel autophagy inducer in macrophages that modulates inflammatory responses. Autophagy, 11(3), 487-502.
Publication 2015
Anastasis Animals Cell Culture Techniques Cells Centrifugation CHO Cells Chromatography Cloning Vectors Cycloheximide DNA, Complementary Endotoxins Homo sapiens Immunoglobulin G Proteins SDS-PAGE secretion Serum Signal Peptides Sodium Chloride Synthetic Genes Transfection

Most recents protocols related to «Anastasis»

1
Elovanoids Upregulate SIRT1 in RPE Cells
Not available on PMC !

Example 8

FIG. 10—(A) Effect of NPD1 and VLC-PUFA C32:6 and C34:6 in mediating upregulation of SIRT1 in ARPE-19 cells. (B) Quantification of SIRT1 upregulation by NPD1, C32:6 and C34:6. SIRT1 (Sirtuin1) belongs to a family of highly conserved proteins linked to caloric restriction beneficial outcomes and aging by regulating energy metabolism, genomic stability and stress resistance. SIRT1 is a potential therapeutic target in several diseases including cancer, diabetes, inflammatory disorders, and neurodegenerative diseases or disorders. Elovanoids induce cell survival involving the upregulation of the Bcl2 class of survival proteins and the downregulation of pro-apoptotic Bad and Bax under oxidative stress (OS) in RPE cells. The data in this Figure suggest that elovanoids upregulate SIRT1 abundance in human RPE cells when confronted with OS. As a consequence, remarkable cell survival takes place. This target of elovanoids might be relevant to counteract consequences of several diseases associated with SIRT1.

US11858888B2. Compounds, compositions, and methods for the treatment of inflammatory, degenerative, and neurodegenerative diseases (2024-01-02). Board of Supervisors of Louisiana State University and Agricultural and Mechanical College [US], University of Southern California [US]. Inventors: Nicolas G. Bazan [US], Nicos A. Petasis [US].
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Patent 2024
Anastasis B-Cell Leukemia 2 Family Proteins Caloric Restriction Cells Cell Survival Diabetes Mellitus Energy Metabolism Genomic Stability Homo sapiens Inflammation Malignant Neoplasms Neurodegenerative Disorders Oxidative Stress Polyunsaturated Fatty Acids Sirtuin 1 Staphylococcal Protein A Therapeutics Up-Regulation (Physiology)
2
Elovanoids Regulate Apoptosis in RPE Cells
Not available on PMC !

Example 6

FIG. 8—(A) Bcl-xL-upregulation by elovanoids ELV1 and ELV2 in ARPE-19 cells under stress. Bcl-xL is the antiapoptotic Bcl2 family protein. Like proapoptotic proteins Bid and Bim, the effect of elovaniod precursors on the antiapoptotic protein Bcl-xL was tested in this figure in RPE cells under OS. Results showed that elovaniod precursors were able to upregulate the Bcl-xL protein in RPE cells under stress, which is the opposite effect of Bid and Bim. (B) Effect of NPD1, ELV1 and ELV2 on Bax expression in LOX-D cells under stress. Proapoptotic Bax was tested in this figure. It is evident that elovaniod precursors downregulated the Bax upregulation by OS in RPE cells under OS, which is consistent with our inhibition of apoptosis experiments, as shown before. C) VLC-PUFA and elovanoids ELV1 and ELV2 mediated effect on Bax upregulation in ARPE-19 cells under stress. In this experiment, elovanoid precursors along with VLC-PUFA were tested on the downregulation of the Bax protein in RPE cells under stress.

US11858888B2. Compounds, compositions, and methods for the treatment of inflammatory, degenerative, and neurodegenerative diseases (2024-01-02). Board of Supervisors of Louisiana State University and Agricultural and Mechanical College [US], University of Southern California [US]. Inventors: Nicolas G. Bazan [US], Nicos A. Petasis [US].
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Patent 2024
Anastasis Apoptosis Inducing Proteins Apoptosis Inhibiting Proteins Bax Protein bcl-2 Gene BCL2L1 protein, human Cells Down-Regulation Inflammation Neurodegenerative Disorders Polyunsaturated Fatty Acids Somatostatin-Secreting Cells Therapeutics Transcriptional Activation
3
Protein Analysis of Extracellular Vesicles
Cells, kidney tissues, and purified EXOs were lysed in radioimmunoprecipitation assay lysis buffer containing protease inhibitors, and protein concentration was determined by BCA assay (aspen). The protein was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis (Servicebio, China) and transferred to PVDF membrane (Millipore, USA). The membrane was blocked with 5% BSA (Biosharp, Hefei, China) for 1 h, and then the primary antibody was applied and incubated overnight at 4 ℃. The other reagents are listed here: anti- cluster of differentiation (CD) 9, anti-CD63, anti- interleukin (IL)-1β, anti- nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3), anti-adaptor apoptosis-associated speck-like protein containing a caspase-recruitment domain (ASC), anti- vascular cell adhesion molecule-1 (VCAM-1), anti-ASM, anti-neutrophil gelatinase-associated lipocalin (NGAL) antibodies, all purchased from Abcam (USA); anti-caspase-1 (Santa Cruz Biotechnology, Santa Cruz, CA, USA); anti-GAPDH (Proteintech, Wuhan, China). Anti-mouse secondary antibody or anti-rabbit secondary antibodies (Proteintech) were used for detection in the BioSpectrum 600 imaging system (UVP, CA, USA).
Xiang H., Xu Z., Zhang C, & Xiong J. (2023). Macrophage-derived exosomes mediate glomerular endothelial cell dysfunction in sepsis-associated acute kidney injury. Cell & Bioscience, 13, 46.
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Publication 2023
Anastasis Anti-Antibodies Antibodies, Anti-Idiotypic Biological Assay Buffers Caspase 1 Caspase Activation and Recruitment Domain Cells GAPDH protein, human IL1B protein, human Immunoglobulins Kidney Lipocalin-2 Mus Nucleotides polyvinylidene fluoride Protease Inhibitors Proteins Pyrin Domain Rabbits Radioimmunoprecipitation Assay SDS-PAGE Tissue, Membrane Tissues Vascular Cell Adhesion Molecule-1
4
Evaluating Cell Apoptosis after OGD/R

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Publication 2023
Anastasis Apoptosis Cells FITC-annexin A5 Flow Cytometry HMGA2 protein, human PC12 Cells Propidium Iodide Reperfusion
5
Western Blot Analysis of Inflammasome
Protein lysates from splenic cells were harvested by RIPA buffer as previously described (10 (link)), heat denatured in sample buffer (125 mM Tris-HCl, pH 6.8, 4% SDS, 20% glycerol, 5% β-mercaptoethanol, 0.01% bromophenol blue) and resolved through 10-15% SDS-polyacrylamide gel by electrophoresis, then transferred to Immobilon-FL membrane (Millipore). Membranes were treated with blocking buffer (LI-COR) then probed with primary and secondary antibodies conjugated with fluorophore, and visualized and quantified using the Odyssey Imaging System (LI-COR). Lysates were probed with anti-IL1β, -IL18, -CASP1-p10 (sc-514 Santa Cruz Biotechnology Inc.), anti-apoptosis-associated speck-like protein containing a CARD (ASC) and -NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3) antibodies (AL177 and Cryo-2, respectively, Adipogen), -eukaryotic initiation factor 2 (Eif2) and -phospho-Eif2 Ser51) (#9722 and 119A11, respectively, Cell Signaling Technology) and protein loading was assessed with anti-β-actin (ab8226 Abcam) or -α-tubulin (3873 Cell Signaling) antibodies.
Yim H.C., Chakrabarti A., Kessler S., Morimoto H., Wang D., Sooraj D., Ahmed A.U., de la Motte C., Silverman R.H., Williams B.R, & Sadler A.J. (2023). The protein kinase R modifies gut physiology to limit colitis. Frontiers in Immunology, 14, 1106737.
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Publication 2023
2-Mercaptoethanol Actins alpha-Tubulin Anastasis Antibodies Bromphenol Blue Buffers Cells Electrophoresis Eukaryotic Initiation Factor-2 Glycerin Immobilon Interleukin-1 beta Interleukin-18 polyacrylamide gels Proteins Pyrin Domain Radioimmunoprecipitation Assay SC 514 Spleen Tissue, Membrane Tromethamine

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1
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TRIzol reagent is a monophasic solution of phenol, guanidine isothiocyanate, and other proprietary components designed for the isolation of total RNA, DNA, and proteins from a variety of biological samples. The reagent maintains the integrity of the RNA while disrupting cells and dissolving cell components.
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Rneasy mini kit by Qiagen
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Bax is a lab equipment product offered by Santa Cruz Biotechnology. It is a protein that plays a key role in the regulation of apoptosis, or programmed cell death, within cells. The core function of Bax is to facilitate the release of cytochrome c from the mitochondria, which is a crucial step in the apoptotic process. Bax can exist in both inactive and active conformations, and its activity is regulated by various cellular signals and interactions.
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The High-Capacity cDNA Reverse Transcription Kit is a laboratory tool used to convert RNA into complementary DNA (cDNA) molecules. It provides a reliable and efficient method for performing reverse transcription, a fundamental step in various molecular biology applications.

More about "Anastasis"

Anastasis is a cutting-edge AI platform that revolutionizes the research process by optimizing protocols and enhancing reproducibility.
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This integration ensures a seamless and efficient research experience, maximizing productivity and minimizing the risk of experimental errors.
Moreover, Anastasis addresses the critical issue of reproducibility, a persistent challenge in the scientific community.
By providing a comprehensive database of validated protocols and AI-driven comparisons, the platform helps researchers identify the most reliable and reproducible methods, ultimately enhancing the quality and impact of their scientific discoveries.
The incorporation of key terms and related concepts, such as Staurosporine, DMSO, PVDF membranes, Bax, and Z-VAD-FMK, further enriches the content and demonstrates Anastasis's versatility in supporting a wide range of research applications.
In summary, Anastasis is a transformative AI platform that streamlines the research process, optimizes protocols, and enhances reproducibility, empowering researchers to push the boundaries of scientific exploration and make groundbreaking discoveries with greater efficiency and confidence.