Opti tof 384 well insert

Manufactured by Thermo Fisher Scientific

Sourced in United States

The Opti-TOF™ 384-well Insert is a laboratory equipment designed to facilitate sample preparation and analysis in 384-well microplate formats. It provides a consistent and efficient platform for various analytical techniques.

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Lab products found in correlation

4800 maldi tof tof analyzer, by Thermo Fisher Scientific (4 mentions) 4700 calibration mixture, by Thermo Fisher Scientific (3 mentions) Dhb matrix, by Merck Group (1 mentions) Orbitrap fusion tribrid mass spectrometer, by Thermo Fisher Scientific (1 mentions) Cyano 4 hydroxycinamic acid, by Merck Group (1 mentions) Trypsin, by Promega (1 mentions) Cyano 4 hydroxy cinnamic acid chca, by Merck Group (1 mentions)

5 protocols using opti tof 384 well insert

Characterization of MUC5AC O-linked Oligosaccharides

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To characterize MUC5AC O-linked oligosaccharides, permethylated glycans were dissolved in methanol (20 μL). A small aliquot (1 μL) was spotted onto a MALDI plate (Opti-TOF-384 well insert, Applied Biosystems) and crystallized with 1 μL of 2,5-dihydroxybenzoic acid (DHB matrix, 10 mg/mL; Sigma-Aldrich, 149357) in 50% methanol/water. Data were obtained from an Applied Biosystems SCIEX MALDI TOF/TOF 5800 mass spectrometer in reflector positive-ion mode ^{42 (link), 63 (link), 64 (link)}.
To confirm glycan structure by ESI-MSⁿ, a small aliquot (2 μL) of permethylated O-glycans was dissolved in ESI-MS direct infusion buffer (33% H₂O, 33% acetonitrile, and 33% 1 mM NaOH-methanol (1:1 vol/vol)). Further, glycans were infused on an Orbitrap Fusion Tribrid mass spectrometer (Thermo Scientific) through a nanospray ionization (NSI) probe. The MSⁿ spectra (collision-induced dissociation, CID and higher energy collisional dissociation, HCD) of the glycans were acquired at high resolution by total ion mapping (TIM) program and detected on Orbitrap / Ion trap ^{42 (link), 65 (link)}.
Data analysis was performed using Data Explorer V4.5, and the assignment of glycan structures was based on the primary m/z coupled with MS/MS fragmentation patterns using the Expasy tool (https://web.expasy.org/glycomod/) and GlycoWorkbench 1.1 software.

Evert C., Loesekann T., Bhat G., Shajahan A., Sonon R., Azadi P, & Hunter R.C. (2019). Generation of 13C-labeled MUC5AC mucin oligosaccharides for stable isotope probing of host-associated microbial communities. ACS infectious diseases, 5(3), 385-393.

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Protein Identification by MALDI-TOF MS

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Protein spots were excised from gels with a sterile scalpel and placed into Eppendorf tubes. Proteins were digested with trypsin (Promega, Madison, WI) as previously described [11 (link)]. For matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) analysis, peptides were concentrated using a POROS R2, Oligo R3 column (Applied Biosystems, Foster City, CA). After washing the column with 70% acetonitrile, 100% acetonitrile, and 50 mM ammonium bicarbonate, samples were applied to the R2, R3 column and eluted with cyano-4-hydroxycinamic acid (Sigma, St. Louis, MO) in 70% acetonitrile and 2% formic acid onto the MALDI plate (Opti-TOF 384-well Insert, Applied Biosystems) [12 (link)]. MALDI-TOF MS analysis was performed on a 4800 MALDI-TOF/TOF Analyzer (Applied Biosystems) equipped with a 355-nm Nd:YAG laser. The pressure in the TOF analyzer was approximately 0.00010132 Pa. Mass spectra were obtained in reflectron mode with an accelerating voltage of 20 kV and sum of 500 laser pulses, and calibrated using the 4700 calibration mixture (Applied Biosystems). Data Explorer 4.4 (PerSeptive Biosystems, Framingham, MA) was used for monoisotopic mass data acquisition and extraction.

Lee E.J., Yang S.H., Kim K.J., Cha H., Lee S.J., Kim J.H., Song J., Chun K.H, & Seong J. (2017). Inter-alpha Inhibitor H4 as a Potential Biomarker Predicting the Treatment Outcomes in Patients with Hepatocellular Carcinoma. Cancer Research and Treatment : Official Journal of Korean Cancer Association, 50(3), 646-657.

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MALDI-TOF/TOF Analysis of BSA Peptides

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Bovine serum albumin (BSA) peptides (Amresco, USA) were N-terminally labeled as described above as control. The peptides were dissolved in 10 μl 0.1% TFA, and 0.5 μl of each sample was mixed with 0.5 μl of a matrix solution that contained 5 mg/ml CHCA (Sigma, USA), 70% ACN, and 0.1% TFA. The peptides were spotted directly onto a MALDI plate (Opti-TOF™ 384-well Insert, Applied Biosystems, USA) and crystallized with the matrix. Dried peptides were analyzed on a 4800 MALDITOF/TOF™ Analyzer (Applied Biosystems) that was equipped with a 355-nm Nd:YAG laser. The pressure in the TOF analyzer was approximately 7.6 × e⁻⁰⁷ Torr.
The mass spectra were obtained in the reflectron mode over an m/z range of 800–3500 Da with an accelerating voltage of 20. External calibration was performed using des-Arg-Bradykinin (904,468 Da), angiotensin 1 (1,296.685 Da), Glu-Fibrinopeptide B (1,570.677 Da), adrenocorticotropic hormone (ACTH) (1–17) (2,093.087 Da), and ACTH (18–39) (2,465.199) (4700 calibration mixture, Applied Biosystems). Raw data were reported by 4000 SERIES EXPLORER, v4.4 (Applied Biosystems).

Min H., Han D., Kim Y., Cho J.Y., Jin J, & Kim Y. (2014). Label-Free Quantitative Proteomics and N-terminal Analysis of Human Metastatic Lung Cancer Cells. Molecules and Cells, 37(6), 457-466.

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MALDI-TOF MS Analysis of Peptides

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For the MALDI-TOF MS analysis, the peptides were concentrated by a POROS R2, Oligo R3 column (Applied Biosystems, Foster city, CA, USA). After washing the column with 70% acetonitrile, 100% acetonitrile and then 50 mM ammonium bicarbonate, samples were applied to the R2, R3 column and eluted with α-cyano-4-hydroxycinnamic acid (HCCA) (Sigma, St. Louis, MO) dissolved in 70% acetonitrile and 2% formic acid onto the MALDI plate (Opti-TOF™ 384-well Insert, Applied Biosystems) (20 (link)). MALDI-TOF MS was performed on a 4800 MALDI-TOF/TOF™ Analyzer (Applied Biosystems) equipped with a 355-nm Nd:YAG laser. The mass spectra were obtained in the reflectron mode with an accelerating voltage of 20 kV and sum from either 500 laser pulses calibrated using the 4,700 calibration mixture (Applied Biosystems).

Yang J.I., Kong T.W., Kim H.S, & Kim H.Y. (2015). The Proteomic Analysis of Human Placenta with Pre-eclampsia and Normal Pregnancy. Journal of Korean Medical Science, 30(6), 770-778.

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Peptide Mass Fingerprinting Protocol

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For peptide mass fingerprinting (PMF), the spots were excised from the stained gel and in-gel digestion was carried out. The excised spots were destained with 50 mM ammonium bicarbonate in 40% acetonitrile, and dried with a speed vac (Heto Lab Equipment, Allerod, Denmark). The destained spots were rehydrated in 12.5 ng/μL trypsin in 50 mM ammonium bicarbonate. The rehydrated spots were placed on ice for 45 min and treated with 50 mM ammonium bicarbonate (10 μL). The spots were then incubated at 37 °C for 12 h.
For MALDI-TOF-MS analysis, the peptides were desalted and concentrated using a POROS R2 and Oligo R3 column (Applied Biosystems, Fostercity, CA, USA). The column was washed sequentially with 70% acetonitrile, 100% acetonitrile, and 50 mM ammonium bicarbonate. The samples were applied and eluted with cyano-4-hydroxycinnamic acid (CHCA) (Sigma, St. Louis, MO, USA) dissolved in 70% acetonitrile and 2% formic acid onto the MALDI plate (Opti-TOF™ 384-well Insert, Applied Biosystems) [35 (link)]. MALDI-TOF-MS was performed on a 4800 MALDI-TOF/TOF™ Analyzer (Applied Biosystems) equipped with a 355-nm Nd:YAG laser (TOF analyzer pressure: approximately 7.6 × 10⁻⁷ Torr). The mass spectra were obtained in the reflection mode (accelerating voltage of 20 kV and the sum from either 500 laser pulses), and calibrated using the 4700 calibration mixture (Applied Biosystems).

Chun S., Ahn S., Yeom C.H, & Park S. (2016). Exosome Proteome of U-87MG Glioblastoma Cells. Biology, 5(4), 50.

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