Enriched fructans oligosaccharides were analysed by MALDI-ToF-MS using a 4700 Proteomics Analyser (Applied Biosystems), as described by Maslen et al. [41 ]. The matrix was 2,5-dihydroxybenzoic acid (10 mg ml−1 dissolved in 50% MeOH). Three different T1 plants and three different T4 plants were used for this analysis. The MALDI-ToF-MS was done with three technical replicates. The results were statistically analyzed by Student’s t test.
4700 proteomics analyser
Manufactured by Thermo Fisher Scientific
Sourced in United States
The 4700 Proteomics Analyzer is a mass spectrometry instrument designed for the analysis of proteins. It features a time-of-flight (TOF) analyzer and a matrix-assisted laser desorption/ionization (MALDI) ion source. The instrument is capable of performing high-throughput analysis of protein samples.
Automatically generated - may contain errors
6 protocols using 4700 proteomics analyser
1
MALDI-ToF-MS Analysis of Enriched Fructans
2
MALDI-ToF/ToF-MS/MS Analysis of Glycans
Check if the same lab product or an alternative is used in the 5 most similar protocols
Per-methylated samples were analysed by MALDI-ToF/ToF-MS/MS (4700 Proteomics Analyser, Applied Biosystems, Foster City, CA, USA) as previously described [50] (link), using 2,5-dihydroxybenzoic acid (2,5-DHB) matrix (10 mg ml−1 dissolved in 50% MeOH). The above tandem mass spectrometer uses a 200 Hz frequency triple Nd-YAG laser operating at 355 nm wavelength. High energy MALDI-CID spectra were acquired with an average 10,000 laser shots/spectrum, using a high collision energy (1 kV). The oligosaccharide ions were allowed to collide in the CID cell with argon at a pressure of 2×10−6 Torr.
+ Expand
Per-methylated samples were analysed by MALDI-ToF/ToF-MS/MS (4700 Proteomics Analyser, Applied Biosystems, Foster City, CA, USA) as previously described [50] (link), using 2,5-dihydroxybenzoic acid (2,5-DHB) matrix (10 mg ml−1 dissolved in 50% MeOH). The above tandem mass spectrometer uses a 200 Hz frequency triple Nd-YAG laser operating at 355 nm wavelength. High energy MALDI-CID spectra were acquired with an average 10,000 laser shots/spectrum, using a high collision energy (1 kV). The oligosaccharide ions were allowed to collide in the CID cell with argon at a pressure of 2×10−6 Torr.
3
In-Gel Tryptic Digestion and MALDI-TOF-TOF MS
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The protein spots of interest were excised and de-stained with 25 mmol/l ammonium bicarbonate/50% acetonitrile (CAN), and in-gel digestion was performed with 0.01 µg/µl trypsin (Promega, Madison, Wisconsin, USA) in 25 mmol/l ammonium bicarbonate for 15 h at 37°C. The hydrolysates were collected, and the tryptic peptides were extracted from the gel pieces sequentially with 5% TFA at 40°C for 1 h and then 2.5% TFA/50% ACN at 30°C for 1 h. The extracts were pooled, lyophilized and stored at -20°C until use. Gel pieces from a 'blank' region and from the BSA molecular mass marker were used as negative and positive controls, respectively.
Subsequently, the peptide mixtures were re-dissolved in 0.5% TFA, and 1 µl of peptide solution was mixed with an equal volume of matrix (4-hydroxy-a-cyanocinnamic acid in 30% ACN/ 0.1% TFA). Then, the peptides were spotted on the target plate. Individual protein peptides were identified by MALDI-TOF-TOF mass spectrometry on a 4700 Proteomics Analyser (Applied Biosystems, Foster City, California, USA). The mass spectra were used to examine human protein sequences in the Swiss-Prot database using the Mascot database search algorithm (version 1.9).
Subsequently, the peptide mixtures were re-dissolved in 0.5% TFA, and 1 µl of peptide solution was mixed with an equal volume of matrix (4-hydroxy-a-cyanocinnamic acid in 30% ACN/ 0.1% TFA). Then, the peptides were spotted on the target plate. Individual protein peptides were identified by MALDI-TOF-TOF mass spectrometry on a 4700 Proteomics Analyser (Applied Biosystems, Foster City, California, USA). The mass spectra were used to examine human protein sequences in the Swiss-Prot database using the Mascot database search algorithm (version 1.9).
4
MALDI-ToF-MS/MS Analysis of Oligosaccharides
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Native or reductively aminated samples were analysed by MALDI-ToF-MS/MS (4700 Proteomics Analyser; Applied Biosystems, http://www.appliedbiosystems.com ), as previously described (Maslen et al., 2007 (link)), using 2,5-dihydroxybenzoic acid (2,5-DHB) matrix (10 mg ml−1 dissolved in 50% MeOH). The above tandem mass spectrometer uses a 200-Hz frequency triple Nd-YAG laser operating with a wavelength of 355 nm. High-energy MALDI-CID spectra were acquired with an average of 10 000 laser shots/spectrum, using a high collision energy (1 kV). The oligosaccharide ions were allowed to collide in the CID cell with argon at a pressure of 2 × 10−6 Torr (0.27 mPa).
5
Protein Identification via MALDI-TOF MS
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For the identification of the target protein, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used. Proteins were visualized with CBB-G250 after SDS-PAGE, and then the target bands were excised from the gel and washed with 25 mM NH HCO , 40 % (v/v) ethanol five times. The gel was cut into pieces and dehydrated with 1 ml of acetonitrile and dried in vacuo. Following this, 0.1 µg/ml trypsin in 25 mM NH HCO was added to the gel pieces and incubated at 37 °C for 16 h. Peptide fragments were extracted from the gel pieces with 50 % (v/v) acetonitrile, 5 % (v/v) trifluoroacetic acid for 30 min. The extracts were dried in vacuo, dissolved in 5 µl of 50 % (v/v) acetonitrile, 0.1 % (v/v) trifluoroacetic acid, and subjected to an ABI 4700 proteomics analyzer (Applied Biosystems). Mass fingerprints of tryptic peptides dissolved in 5 µl of 50 % (v/v) acetonitrile, 0.1 % (v/v) trifluoroacetic acid were generated by MALDI-TOF-MS using an Applied Biosystems 4700 Proteomics Analyser with TOF/TOF optics in the MS mode. A Nd:YAG laser (355 nm) was used to irradiate the sample. The spectra were acquired in reflection mode in the mass range 700-3200 Da. Amino-acid sequences of the fragments were determined in MS/MS mode with DeNovo Explorer software (Applied Biosystems).
6
MALDI-TOF-MS Analysis of Glycans
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The isolated oligosaccharides were analysed by MALDI-TOF-MS. The matrix was a dihydroxybenzoic acid solution in 50% ACN (10 mg.ml -1 ). The analyses were carried on a 4700 Proteomics Analyser with TOF/TOF optics (Applied Biosystems, Framingham, MA). The mass spectrometer had a 200 Hz frequency-tripled Nd-YAG laser operating at a wavelength of 355 nm. A total of 1500 shots were acquired in the MS mode. Spectra from m/z 900 to 3000 were recorded. Glycans were detected as [M+Na] + ions.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!