Amicon ultra centrifuge filter

Manufactured by Merck Group

Sourced in United States, Germany

The Amicon Ultra centrifuge filter is a lab equipment product designed for sample preparation and concentration. It features a semi-permeable membrane that allows the passage of small molecules while retaining larger molecules or particles during centrifugation.

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Lab products found in correlation

Qubit protein assay kit, by Thermo Fisher Scientific (2 mentions) Histrap ff column, by GE Healthcare (2 mentions) Flexanalysis 3, by Bruker (1 mentions) Maldi tof ms, by Bruker (1 mentions) Ar 2000 radio tlc imaging scanner, by Bioscan (1 mentions) Stem cell factor (scf), by Amgen (1 mentions) Interleukin 3 (il 3), by Thermo Fisher Scientific (1 mentions) Stem cell factor (scf), by Thermo Fisher Scientific (1 mentions) Procrit, by Amgen (1 mentions) Anti il 33 antibody, by R&D Systems (1 mentions) Thrombopoietin tpo, by Thermo Fisher Scientific (1 mentions) Bovine serum albumin (bsa), by Merck Group (1 mentions) Rbc lysis buffer, by Merck Group (1 mentions) β actin bsm 33036m, by Bioss Antibodies (1 mentions) Rhodamine b rho, by Merck Group (1 mentions) Cytopainter lysored indicator reagent ab176828, by Abcam (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Abcam (1 mentions) Fetal bovine serum (fbs), by Wisent (1 mentions) Trypsin, by Wisent (1 mentions) 3 4 5 dimethylthiazol 2 yl 2 5 diphenyl 2h tetrazolium bromide mtt, by Wisent (1 mentions)

Close spelling variants of this product

Amicon Ultra (740 citations) Amicon filters (181 citations) Amicon Ultra filters (132 citations) Amicon centrifuge filters (23 citations) Centrifuge filter (11 citations) Amicon Ultra-0.5 centrifuge filters (2 citations) Amicon Ultra-15 mL Centrifuge Filters (2 citations)

33 protocols using amicon ultra centrifuge filter

Synthesis and Evaluation of DTPA-AHNP-PEG Conjugate

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For preparation of DTPA conjugated-AHNP-PEG (DTPA-AHNP-PEG), AHNP-PEG and p-SCN-Bn-DTPA (w/w 1:50) was soaked in sodium carbonate buffer at 25 °C for 8 h. The conjugated-compounds were then purified by Amicon ultra centrifuge filters (3 kDa) (Merck Millipore). For molecular weight measurement, AHNP-PEG and DTPA-AHNP-PEG were detected with the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS; Bruker Daltonics, Billerica, MA, USA). The spectra were processed using FlexAnalysis™ 3.0 software (Bruker Daltonics). The DTPA-AHNP-PEG conjugation efficiency with indium-111 (¹¹¹In-DTPA-AHNP-PEG) was evaluated by instant thin layer chromatography (ITLC) (AR-2000 radio-TLC Imaging Scanner; Bioscan, Washington, DC, USA).

Guan S.S., Wu C.T., Chiu C.Y., Luo T.Y., Wu J.Y., Liao T.Z, & Liu S.H. (2018). Polyethylene glycol-conjugated HER2-targeted peptides as a nuclear imaging probe for HER2-overexpressed gastric cancer detection in vivo. Journal of Translational Medicine, 16, 168.

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Erythroid Differentiation of CD34+ Cells

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Purified human CD34+ progenitor cells were derived from GCSF–treated peripheral blood cells of healthy donors. These cells were grown at 37°C with 5% CO2 in serum-free medium consisting of Iscove’s modified Dulbecco’s medium (IMDM) with 1-thioglycerol, BIT9500 supplement (BITS) (Stem Cell Technologies), BSA (Sigma-Aldrich), and the indicated cytokines (PeproTech). The cells initially underwent 72 h of expansion with 100 ng/ml SCF (PeproTech), 100 ng/ml FMS-like tyrosine kinase 3 ligand (FLT3 ligand) (PeproTech), 100 ng/ml thrombopoietin (TPO) (PeproTech), and 50 ng/ml IL-3 (PeproTech). After expansion cells were then seeded in erythroid differentiation medium, which contains recombinant human erythropoietin at 4.5 U/ml (Procrit; Amgen), 10 ng/ml SCF and BIT9500 supplement. Human bronchial epithelial cells (Beas2B) were grown at 37°C with 5% CO2 in DMEM medium with 10% FBS. Horseradish peroxidase-conjugated goat anti-rabbit and anti-mouse antibodies, reverse transcriptase, and real-time PCR SosoFast reagents were purchased from Bio-Rad Laboratories (Hercules, CA). Amicon Ultra centrifuge filters were purchased from EMD Millipore (Billerica, MA). Anti-IL-33 antibody was from R&D Systems (Minneapolis, MN). Anti-LarminA/C antibody was from Santa Cruz (Dallas, TX). Anti-GAPDH and β-actin antibodies and RBC lysis buffer were from Sigma (St. Louis, MO).

Wei J., Zhao J., Schrott V., Zhang Y., Gladwin M., Bullock G, & Zhao Y. (2015). Red blood cells store and release interleukin-33. Journal of investigative medicine : the official publication of the American Federation for Clinical Research, 63(6), 806-810.

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Nanoparticle-Mediated Anticancer Therapy

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FUdR was purchased from Alfa Aesar (Shanghai, China). FUdR15 strands were synthesized by Sangon Biotech Co., Ltd. (Shanghai, China). PHBHHx was purchased from Bluepha (Beijing, China). Rhodamine B (Rho) and Cur were purchased from Sigma-Aldrich (Shanghai, China). DAPI and CytoPainter LysoRed Indicator Reagent (ab176828) were obtained from Abcam Co., Ltd. (Cambridge, MA, USA). Trypsin, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT), antibiotic-antimycotic (100×), and fetal bovine serum (FBS) were purchased from Wisent Biotechnology Co., Ltd. (Nanjing, China). All chemicals were used without further purification. Primary antibodies to Bcl-2 (bsm-33047M), Bax (bsm-33283M), and β-actin (bsm-33036M) were purchased from Bioss Antibodies. Caspase 3, 8, and 9 activity detection kits were purchased from BestBio Biotechnology Co., Ltd. (Shanghai, China). Amicon ultracentrifuge filters were purchased from Merck Millipore Ltd. (Darmstadt, Germany).

Zhang F., Zhang C., Fu S., Liu H., Han M., Fan X., Zhang H, & Li W. (2022). Amphiphilic Cationic Peptide-Coated PHA Nanosphere as an Efficient Vector for Multiple-Drug Delivery. Nanomaterials, 12(17), 3024.

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Purification of His-tagged IMP1 Mutants

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IMP1 and its mutants were inserted into pET28a plasmid containing His tag. The fusion proteins were expressed in E. coli strain Rosetta (Shanghai Weidi Biotechnology, #EC1010) under 0.5 mM IPTG, 25 °C for 4-6 h induction after grown to OD₆₀₀ 0.6-0.8. After IPTG induction, E. coli cell pellets were collected, then resuspended with His Lysis buffer (50 mM Tris-HCl pH8.0, 500 mM NaCl, 10% glycerol, 1.0% Triton X-100, 10 mM Imidazol, 0.2 mM PMSF), and further crushed using high pressure homogenizer. Protein supernatant was collected by centrifugation at 12,500 rpm for 15 min at 4 °C. About 150-200

μ L

Ni-NTA Agarose beads (Abclonal, #AS045) were washed twice using His Lysis buffer and then incubated with protein supernatant at 4 °C for 3-4 h. The beads were collected and washed twice for 10 min using 5 mL His Washing buffer (50 mM Tris-HCl pH 8.0, 500 mM NaCl, 10% glycerol, 20 mM Imidazol). Finally, the protein was eluted from the beads using 500

μ L

His Elution buffer (20 mM Tris-HCl pH 8.0, 500 mM NaCl, 10% glycerol, 300 mM Imidazol). The eluted protein was concentrated using Amicon Ultracentrifuge filters (Merck, #UFC201024). Protein purity and concentration were determined with SDS-PAGE followed by Coomassie blue staining. BSA was used as protein standard sample.

Lu P., Yang J., Li M., Wen S., Zhang T., Yan C., Liu R., Xiao Y., Wang X, & Jiang W. (2023). A desert lncRNA HIDEN regulates human endoderm differentiation via interacting with IMP1 and stabilizing FZD5 mRNA. Genome Biology, 24, 92.

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TLC-based Protein Purification Protocol

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Samples were prepared and analysed by TLC (see above) and 6 μL of the reaction were diluted 1:10 in dH₂O. Proteins were removed by filtration using Amicon Ultra centrifuge filters (3000 MWCO; Sigma-Aldrich). Flow-through was stored at −80 °C until analysis.

Rack J.G., Liu Q., Zorzini V., Voorneveld J., Ariza A., Honarmand Ebrahimi K., Reber J.M., Krassnig S.C., Ahel D., van der Marel G.A., Mangerich A., McCullagh J.S., Filippov D.V, & Ahel I. (2021). Mechanistic insights into the three steps of poly(ADP-ribosylation) reversal. Nature Communications, 12, 4581.

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Zika Virus Antigen Preparation and Splenocyte Assay

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Fourteen days after the booster administration, spleens were collected under aseptic conditions, immersed in RPMI 1640 medium (Invitrogen, USA) and mechanically dissociated to obtain a homogeneous cell suspension as described elsewhere (17 (link), 26 (link)). Either zEDIII antigen (10 μg/mL), inactivated ZIKV strain 17 (10 μg/mL) or medium with an unrelated antigen (inactivated bovine herpesvirus, 10 μg/mL) was used for in vitro re-stimulation antigen.
For antigen production, the supernatant medium of ZIKV-infected cells was clarified using low speed centrifugation and filtered through a 0.45 μm filter. Viral particles were purified by ultracentrifugation at 100,000 × g on a 20% sucrose cushion. The viral pellet was inactivated with 0.05% formalin at 22°C for 7 days. The formalin was then removed by extensive dialysis against PBS (pH 7.2). Antigen concentration was then carried out using Amicon® Ultra Centrifuge Filters (molecular weight cut-off of 30 K, Sigma-Aldrich, USA). Antigen inactivation was confirmed by three passages in Vero cells. The amount of viral protein antigen was quantified by fluorimetry in a Qubit apparatus (Thermo Fisher Scientific, USA) using the Qubit protein assay kit.

Cibulski S., Varela A.P., Teixeira T.F., Cancela M.P., Sesterheim P., Souza D.O., Roehe P.M, & Silveira F. (2021). Zika Virus Envelope Domain III Recombinant Protein Delivered With Saponin-Based Nanoadjuvant From Quillaja brasiliensis Enhances Anti-Zika Immune Responses, Including Neutralizing Antibodies and Splenocyte Proliferation. Frontiers in Immunology, 12, 632714.

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Preparation and Characterization of Apo-AcpP

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Apo-AcpP was prepared from a mixture of apo- and holo-AcpP according to a previous published protocol (Kosa, 2012 (link)). Final reaction concentrations: 50 mM potassium phosphate (pH 7.0), 4 mM ATP, 12.5 mM MgCl2, 0.01 µg/µL MBP-CoaA, 0.01 µg/µL MBP-CoaD, 0.01 µg/µL MBP-CoaE, 0.008 µg/µL Sfp (native), 0.6 µg/µL AcpP, 5% DMSO and 500 µM crosslinker probe. As a negative control, the crosslinker probe was replaced with equal volume of DMSO. After the addition of all components, reactions were gently mixed and incubated at 37 °C for 3 hours or overnight. The reaction mixture was centrifuged to remove precipitation, filtered using 0.45 µm syringe filter, and loaded on HiTrap Q anion exchange column to remove the coupling enzymes and unreacted crosslinking probes. The loaded AcpP eluted at ~0.3 M NaCl in 25 mM sodium phosphate (pH 8.0). The eluted protein was collected and concentrated using Amicon Ultra Centrifuge Filters (Millipore) with 3 kDa molecular weight cut off. Loading of DH6, DH8, DH10, and DH10X onto AcpP were monitored using MALDI-TOF (AB SCIEX TOF/TOF 5800), which has a 0.5% margin of error.

Finzel K., Nguyen C., Jackson D.R., Gupta A., Tsai S.C, & Burkart M.D. (2015). Probing the Substrate Specificity and Protein-Protein Interactions of the E. coli Fatty Acid Dehydratase, FabA. Chemistry & biology, 22(11), 1453-1460.

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Production and Purification of Viral Vectors

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Viral vectors were produced in-house for AAV8-CKIIa-stGtACR2-FusionRed (6.1 × 10¹² vg/ml) as follows. HEK 293 cells were transfected with purified helper (E4, E2a and VA), AAV8 Rep/Cap and transfer plasmids containing the pyramidal cell specific CKIIa promoter, stGtACR2 (soma targeting Guillardiatheta anion-conducting channelrhodopsins) and FusionRed DNA using a polyethylenimine method (Boussif et al., 1995 (link)). Subsequently, cells were manually harvested, centrifuged and pelleted. Crude viral particles in the supernatant were then removed. The virus was column purified using an AKTA (GE Life Sciences) chromatography system and concentrated using Amicon ultracentrifuge filters (Millipore). Viral titer was measured by real-time PCR using serial dilutions then aliquoted and frozen at −80°C. Control experiments were also conducted using another viral vector, AAV8-CKIIa-GFP (8 × 10¹² vg/ml), purchased from UNC Vector Core (aliquoted and stored at −80°C). All procedures involving viral vectors were done under Newcastle University safety guidelines and approved by the Microbiological Hazards and Genetic Modification Safety Advisory Sub-Committee.

McLeod F., McDermott E., Mak S., Walsh D., Turnbull M., LeBeau F.E., Jackson A., Trevelyan A.J, & Clowry G.J. (2024). AAV8 vector induced gliosis following neuronal transgene expression. Frontiers in Neuroscience, 18, 1287228.

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Serglycin Detection in Conditioned Media

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A total of 1 × 10⁶ cells were plated in 100 mm dishes and cultured in regular medium. After 24 h in culture, cells were starved for 24 h. Culture supernatants were collected, centrifuged at 3000 r.p.m. for 5 min and concentrated with Amicon Ultracentrifuge filters (10 kDa molecular weight cutoff pore size; Millipore, Boston, MA, USA). Serglycin was detected in concentrated CM by immunoblotting.

Chu Q., Huang H., Huang T., Cao L., Peng L., Shi S., Zheng L., Xu L., Zhang S., Huang J., Li X., Qian C, & Huang B. (2016). Extracellular serglycin upregulates the CD44 receptor in an autocrine manner to maintain self-renewal in nasopharyngeal carcinoma cells by reciprocally activating the MAPK/β-catenin axis. Cell Death & Disease, 7(11), e2456-.

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Proteomic profiling of microglia-fibroblast crosstalk

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FBS-free medium was used in LC-MS/MS analysis. Specifically, mouse microglia were cultured in FBS-free DMEM containing 5μM hemoglobin for 24 h. The resulting microglia-conditioned medium was used to treat human fibroblasts for 24 h to collect fibroblast-conditioned medium. The microglia- and fibroblast-conditioned media were concentrated using Amicon Ultra centrifuge filters (MWCO 10 kDa, Millipore, Z677108), and proteins were separated in gradient SDS-PADE. Hemoglobin bands were cut off from the gel and the remaining gel pieces were pooled and submitted to the Proteomics and Mass Spectrometry Facility for LC-MS/MS analysis. Briefly, proteins were digested using an in-gel trypsin digestion protocol. Mass spectrometry analyses were performed on a Thermo-Fisher LTQ Orbitrap Elite Mass Spectrometer coupled with a Proxeon Easy NanoLC system (Waltham, MA). Data were acquired using Xcalibur software (version 2.2, Thermo Fisher Scientific). Protein identification and modification characterization were performed using Thermo Proteome Discoverer (version 1.4) with Mascot (Matrix Science 2.7) and UniProt database. The spectra of possible modified peptides were inspected further to verify the accuracy of the assignments.

Xu L., Nirwane A., Xu T., Kang M., Devasani K, & Yao Y. (2022). Fibroblasts repair blood-brain barrier damage and hemorrhagic brain injury via TIMP2. Cell reports, 41(8), 111709.

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