Raw HiPCO SWNTs (Unidym) were suspended in an aqueous solution containing 1 wt% sodium deoxycholate by 1 hour of bath sonication. This suspension was then ultracentrifuged at 300,000 g to collect the supernatant, and 0.75 mg·ml−1 of DSPE-mPEG(5 kDa) (1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethyleneglycol, 5000)], Laysan Bio) along with 0.25 mg·ml−1 of DSPE-PEG(5 kDa)-NH2 (1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethyleneglycol, 5000)], Laysan Bio) was added to the supernatant. The resulting suspension was sonicated for 5 min and dialyzed against 1x phosphate buffer saline (PBS) at pH 7.4. These amine-functionalized SWNTs were conjugated with IRDye800 dye molecules. Briefly, the as-made SWNT solution was concentrated down to ~2 µM after removal of excess surfactant through 30-kDa centrifugal filters (Amicon) and was mixed with 0.1 mM IRDye800 NHS ester (LI-COR) dissolved in dimethylsulfoxide (DMSO). The reaction was allowed to proceed for 1 h at pH 7.4 before removing excess IRDye800 by filtration through the 30-kDa filters.
Irdye800 nhs ester
Manufactured by LI COR
Sourced in United States
IRDye800 NHS ester is a near-infrared fluorescent dye that can be used for labeling proteins, peptides, and other biomolecules. It has an excitation maximum at 774 nm and an emission maximum at 789 nm, allowing it to be detected in the near-infrared range.
Automatically generated - may contain errors
Lab products found in correlation
Dspe mpeg 5 kda, by Laysan Bio (2 mentions)
Dspe peg 5 kda nh2, by Laysan Bio (2 mentions)
Milli q purification system, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Tween 20, by Merck Group (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
Phosphate buffered saline (pbs), by Merck Group (1 mentions)
Cholesteryl chloroformate, by Merck Group (1 mentions)
1 2 ethanediamine, by Merck Group (1 mentions)
1 1 carbonylbis 1h imidazole, by Merck Group (1 mentions)
Polyinosinic polycytidylic acid sodium salt poly 1 c, by Merck Group (1 mentions)
Zeba desalting column, by Thermo Fisher Scientific (1 mentions)
Sodium periodate naio4, by Thermo Fisher Scientific (1 mentions)
Il 1b, by Abcam (1 mentions)
Corm 3, by Merck Group (1 mentions)
Tnf α, by Abcam (1 mentions)
Live dead cell double staining kit, by Merck Group (1 mentions)
Irdye 680 nhs ester, by LI COR (1 mentions)
Interleukin 6 (il 6), by Abcam (1 mentions)
Cck 8, by Beyotime (1 mentions)
8 protocols using irdye800 nhs ester
1
Functionalization of Single-Walled Carbon Nanotubes
2
SARS-CoV-2 Variant Serological Assay
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pGOLD chips were purchased from Nirmidas Biotech Inc. (Palo Alto, USA). The glass chips coated by amino silane were purchased from Corning incorporated (New York, USA). Tween-20, PBS and BSA were purchased from Sigma-Aldrich (Shanghai, China). IRDye800-NHS ester and Cy5-NHS ester were purchased from Licor Biosciences (Nebraska, USA). Fetal bovine serum (FBS) was purchased from Invitrogen (Carlsbad, USA). Recombinant human IgG and human IgM were purchased from Acro biosystems. Goat anti-human IgG antibody and goat anti-human IgM were purchased from Sino Biological. The Wild-type S1 (cat. 40,591-V08H3), Alpha variant S1 (cat. 40,591-V08H12), Beta variant S1 (cat. 40,591-V08H15), Delta variant S1 (cat. 40,591-V08H23) and Omicron variant S1 (cat. 40,591-V08H41) were purchased from Sino Biological. The sera of various variants recovered-individuals were provided by Affiliated Hospital of South University of Science and Technology. And 23 control samples (collected before the SARS-CoV-2 outbreak) were acquired from the WWHS Biotech. Inc. (Shenzhen, China). Other chemicals used in this study were of analytic grade. Ultrapure water (≥ 18.2 MΩ/cm), purified using a Milli-Q purification system (Millipore), was used in all experiments.
3
Functionalization of Single-Walled Carbon Nanotubes
4
Synthesis and Functionalization of γ-PGA
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Poly-(γ-glutamic acid) (γ-PGA; molecular weight =50 kDa) was provided by BioLeaders Corporation (Daejeon, Republic of Korea). Cholesteryl chloroformate (97%), 1,1′-carbonylbis-1H-imidazole, 1,2-ethanediamine, and polyinosinic–polycytidylic acid sodium salt, poly-(I:C), were purchased from Sigma-Aldrich (St Louis, MO, USA). IRDye800 NHS ester was purchased from Li-COR Biosciences (Lincoln, NE, USA).
5
Fluorescent Labeling of C2-Crry and Anti-CII Antibodies
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To show the binding of C2-Crry in vitro to apoptotic FLS, we labeled the protein with IRDye800-NHS ester (Li-COR Biosciences) as we have shown previously (60; 61 (link)). C2-Crry fusion protein was incubated with a 10-fold molar excess of dye for 5 h at 4°C in PBS. Unbound dye was removed with a 7 kDa Zeba desalting column (Thermo Fisher). After labeling C2-Crry with IRDye800 and addition to a Zeba column, the protein was eluted from the column with PBS and stored at 4°C before use according to our previously published study (60; 61 (link)). Similarly, Arthrogen (anti-CII) mAbs were labeled with IRDye 800 (60; 61 (link)).
6
Fluorescent Labeling of Ovalbumin Protein
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Ovalbumin protein was fluorescently labeled for live animal imaging studies using amine reactive IRDye 800-NHS ester (LI-COR). Protein and dye were mixed at a 1:1 equimolar ratio in 2mL 1X PBS (pH 8.4) at RT with continuous mixing for an hour. Labeled protein was purified using Amicon filters (10K MWCO) by centrifugation at 3700g for 10 minutes and washing with 1X PBS. The degree of protein labeling was quantified by absorption at 280nm and 774nm using Nanodrop.
7
Multifunctional Wound Dressing Fabrication
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Chitosan (degree of deacetylation≈95%, 100–200 mps), dextran (Mw: ~70 kDa), 3, 4-dihydroxy hydrocinnamic acid, 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC), lead-oxide (ZnO) were purchased from Aladdin. Sodium periodate (NaIO4) was purchased from Acros. 4-(Methylamino) Pyridine was purchased from TCI. Fibrin glue was purchased from Shanghai yuanye Bio-Technology Co., Ltd. Gelatin type A (from porcine skin, 300 Bloom), Live/Dead Cell Double Staining Kit, CORM-3, and GSNO were purchased from Sigma. CCK-8 was purchased from Beyotime. CD31, IL-6, TNF-a, IL-1b, and all other antibodies are purchased from Abcam. IR@dye 680-NHS ester and IR@dye 800-NHS ester were purchased from LI-COR.
8
Fluorescent Labeling of Microbial Cells
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Samples were always kept in ice unless specified. To remove excipients, 200 mg of EDP1867 drug substance (powder formulation) were resuspended and extensively washed by repeated centrifugation/resuspension (4 min, 9000 x g, 4°C) using sterile labeling buffer (2x PBS buffer pH 8.3). Final microbial cell pellets where resuspended in 1 mL final volume using labeling buffer. The fluorescent labeling reaction was started by addition of IRDye800-NHS Ester (LICOR Biosciences) at 50 µM final concentration (from 10 mM dye stock dissolved in DMSO). The reaction was allowed to continue for 1.5 hours at 22°C with gentle agitation, followed by overnight incubation at 4°C. Non-reacted dye was removed by repeated steps of centrifugation/resuspension in sterile PBS buffer until no fluorescence was detected in the supernatant. Fluorescently labeled microbial cells were resuspended in sterile PBS buffer and cell concentration was quantified using coulter counter instrument (Beckman). The molar concentration of IRDye800® covalently attached to microbial cells was quantified by UV-Vis absorbance and using the dye’s molar extinction coefficient 240.000 M-1cm-1 at 778 nm in PBS buffer (LICOR Biosciences). A suspension of unlabeled bacterial cells was used to subtract any light scatter contributions.
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