Peptides were synthesized using the solid-phase-9-fluorenylmethocycarbonyl1 (F-moc) method on a Rink amide-4-methylbenzhydrylamine resin and a Liberty microwave peptide synthesizer (CEM Co., Matthews, NC, USA). Hydroxybenzotriazole (0.1 M) dissolved in piperidine and dimethylformamide and 0.45 M 2-(1H-benzotriazole-1-yil)-1,1,3,3-tetramethyluroniunm hexafluorophosphate dissolved in dimethylformamide were used as linkage reagents. After washing with dichloromethane, cleavage was performed in a solution of trifluoroacetic acid, phenol, water, and triisopropylsilane for 2 h at 25 °C. The crude peptides were diluted with ice-cold ether for precipitation. The peptides were added to tubes and then completely dried. The dried peptides were diluted in distilled water and then purified by RP-HPLC on a Jupiter C18 column (4.6 × 250 mm, 300 Å, 5 μm; Phenomenex, Torrance, CA, USA). The molecular weights of the peptides were confirmed by MALDI-TOF MS (Kratos Analytical, Inc., Chestnut Ridge, NY, USA). Projections of the predicted three-dimensional structures were confirmed using PEP-FOLD3 (https://bioserv.rpbs.univ-paris-diderot.fr/services/PEP-FOLD3/ ). PEP-FOLD3 can be used for structural characterization of the peptides, followed by visualization using PyMOL [48 (link),49 (link)]. The HeliQuest site (http://heliquest.ipmc.cnrs.fr ) was used to generate helical wheel diagrams.
Maldi tof ms
Manufactured by Shimadzu
Sourced in Japan, United States
The MALDI-TOF MS (Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry) is a type of mass spectrometry instrument. It is used for the analysis and identification of large biomolecules, such as proteins, peptides, and oligonucleotides, by measuring their mass-to-charge ratios.
Automatically generated - may contain errors
Lab products found in correlation
Jupiter c18 column, by Phenomenex (2 mentions)
Micro bio spin p 6 column, by Bio-Rad (2 mentions)
Hplc labsolution, by Shimadzu (1 mentions)
3 3 dipropyl thiadicarbocyanine iodide disc3 5, by Thermo Fisher Scientific (1 mentions)
Phospholipid, by Avanti Polar Lipids (1 mentions)
F moc solid phase peptide synthesis, by GL Biochem (1 mentions)
Inertsil ods sp, by Shimadzu (1 mentions)
Cary 50 uv vis spectrophotometer, by Agilent Technologies (1 mentions)
C18 column, by Shimadzu (1 mentions)
Alexa 680 c2 maleimide, by Thermo Fisher Scientific (1 mentions)
High performance liquid chromatography (hplc), by Shimadzu (1 mentions)
Avance 3 400, by Bruker (1 mentions)
Modified trypsin, by Promega (1 mentions)
Glu c, by Promega (1 mentions)
Lcms it tof system, by Shimadzu (1 mentions)
Microbank, by Pro-Lab Diagnostics (1 mentions)
Fp 8200 spectrofluorometer, by Jasco (1 mentions)
Disc3 5, by Thermo Fisher Scientific (1 mentions)
Jes fa200 esr spectrometer, by Bruker (1 mentions)
F 7000 fluorescence spectrophotometer, by Hitachi (1 mentions)
39 protocols using maldi tof ms
1
Peptide Synthesis and Structural Characterization
2
Solid-Phase Fmoc Peptide Synthesis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Peptides were synthesized via the solid-phase F-moc method on a Rink amide-4-methylbenzhydrylamine resin using a Liberty microwave peptide synthesizer (CEM Co.) (Park et al., 2020 (link)). Linkage reagents included 0.1 M hydroxybenzotriazole in piperidine/dimethylformamide and 0.45 M 2-(1H-benzotriazole-1-yil)-1,1,3,3-tetramethyluroniunm hexafluorophosphate in dimethylformamide. Following dichloromethane washes, cleavage occurred in a trifluoroacetic acid, phenol, water, and triisopropylsilane mixture for 2 h at 25 °C. Crude peptides were precipitated using ice-cold ether, dried, reconstituted in distilled water, and purified by RP-HPLC on a Jupiter C18 column (Phenomenex). Peptide molecular weights were determined by Matrix-Assisted Laser Desorption/Ionization—Time of Flight Mass Spectrometry (MALDI-TOF MS) (Kratos Analytical, Inc.).
3
Synthesis of Photosensitizer-Peptide Conjugates
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The purchased peptide NH2-FFVLGGGC (60 mg, 1 eq.) and Ad-ss-Py (31.41 mg, 1.2 eq.) were mixed in 3 mL DMF and stirred in the dark for 12 h at 30 °C under a nitrogen atmosphere to obtain Ad-ss-pep. Then Ce6 (53.84 mg, 1 eq.), EDC (28.10 mg, 3 eq.), and NHS (31.15 mg, 3 eq.) were dissolved in 2 mL DMF and stirred for 2 h at room temperature to activate the carboxyl of Ce6. The latter solution and TEA (37.52 μL, 3 eq.) were added into the former one and allowed to react for 24 h in the dark at 30 °C under a nitrogen atmosphere. The final collected solution was dialyzed with DMSO(MWCO1000) for 12 h and with deionized water for 24 h at room temperature.
The preparation of Ad-pep-Ce6 is similar to the Ad-ss-pep-Ce6, with only the first step being slightly different. The peptide NH2-FFVLGGGC (60 mg, 1 eq.), Ad-Mal (45.45 mg, 2 eq.), and TEA (the pH of the solution was adjusted to 7.5) were mixed in DMF and stirred at 30 °C under a nitrogen atmosphere for 24 h. The subsequent reaction with Ce6 was the same as described above. The final product and intermediate product of Ad-ss-pep-Ce6 and Ad-pep-Ce6 were verified by 1H NMR spectrum (Agilent), LC‒MS (PerkinElmer), and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS, SHIMADZU, AXIMA-TOF, Japan) (the matrix was sinapic acid).
The preparation of Ad-pep-Ce6 is similar to the Ad-ss-pep-Ce6, with only the first step being slightly different. The peptide NH2-FFVLGGGC (60 mg, 1 eq.), Ad-Mal (45.45 mg, 2 eq.), and TEA (the pH of the solution was adjusted to 7.5) were mixed in DMF and stirred at 30 °C under a nitrogen atmosphere for 24 h. The subsequent reaction with Ce6 was the same as described above. The final product and intermediate product of Ad-ss-pep-Ce6 and Ad-pep-Ce6 were verified by 1H NMR spectrum (Agilent), LC‒MS (PerkinElmer), and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS, SHIMADZU, AXIMA-TOF, Japan) (the matrix was sinapic acid).
4
MALDI-TOF–MS Protein Analysis Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Analyzes were performed using MALDI-TOF–MS (AXIMA Performance, Shimadzu Co., Japan) with peak purified by RP-HPLC added to 20 mg. ml−1 sinapic acid saturated in 50% acetonitrile and 0.1% trifluoroacetic acid as a matrix, and analyzes were performed in positive linear mode.
5
Exenatide-SRHWKFL Peptide Hydrazide Synthesis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Example 44
Production of Peptide Hydrazide Compound Using Exenatide-SRHWKFL Peptide
500 μL of a reaction liquid (a 100 g/mL Exe-SRHWKFL solution, a 100 mM HEPES buffer (pH of 8.2), 1 mM NiSO4, 50% methanol) was prepared, and a reaction was performed at 37° C. overnight to obtain a methyl ester compound. Then, 21 μL of hydrazine monohydrate was added to the reaction liquid (the final concentration of hydrazine monohydrate in the reaction liquid was 5%), and the mixture was reacted at 25° C. for 3 hours to obtain a hydrazide compound. The reaction liquid was ultrafiltrated with Amicon Ultra 0.5 mL 3 kDa (Merck Millipore), replaced with a 20 mM sodium phosphate buffer (pH of 7), and concentrated to 25 μL. Then, 250 μL of a reaction liquid (200 mM NaH2PO4, 6 M guanidine hydrochloride, 20 mM NaNO2, pH of 3) containing 25 μL of the concentrate was prepared, and a reaction was performed on ice for 1 hour. To 125 μL of this reaction liquid, 125 μL of a reaction liquid (200 mM Na2HPO4, 6 M guanidine hydrochloride, 200 mM MPAA, 10 mM Cys-Lys (Biotin)) was added and reacted at a pH of 7 at 25° C. overnight to obtain a compound in which Cys-Lys (Biotin) was added to Exenatide. The reaction product was confirmed by molecular weight analysis using MALDI-TOF MS (an AXIMA (registered trademark)-TOF2 laser ionization time-of-flight-mass-spectrometry device, produced by Shimadzu Corporation).
6
Synthesis of TfR-T12-PEG2000-DSPE Conjugate
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To synthesize TfR-T12-PEG2000-DSPE, TfR-T12 (THRPPMWSPVWP, 8 μmol) and NHS-PEG2000-DSPE (8 μmol) were dissolved in DMF (2 mL), and N-methylmorpholine (200 μL) was added into the mixture. The mixture was stirred using a magnetic stirrer at room temperature for 48 h under nitrogen gas protection. Then, the crude product was transferred into a regenerated cellulose dialysis tubing (MWCO, 3000 Da) and dialyzed against deionized water for 24 h to remove unreacted raw materials and dimethyl formamide (DMF). The mixture was freeze dried, and the product was obtained. The product was confirmed by MALDI-TOF-MS (Shimadzu, Japan).
7
Synthesis and Purification of Periplanetasin-4 Peptides
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
All peptides were chemically synthesized by Anygen (Korea), using the solid-phase peptide synthesis method with Fmoc (9-fluorenyl-methoxycarbonyl) chemistry. Periplanetasin-4 and its analogs were manually synthesized. Assembly of the peptides was achieved with a 60-min cycle for each residue at ambient temperature using a reactor with a specially-designed shape. The crude peptide was repeatedly washed with diethyl ether and dissolved in 0.1 mM ammonium bicarbonate, water and acetonitrile. The mixture was then freeze-dried in a lyophilizer after the salts were excluded. The peptides were purified using preparative reverse-phase high-performance liquid chromatography (RP-HPLC) on C18 columns (20 × 250 mm; Shim-pack; Shimadzu, Japan). The purity of the peptides was verified with analytical RP-HPLC, and the peptide masses were confirmed using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS; Shimadzu). The course of the reaction was monitored using HPLC. The purity of the periplanetasin-4 and the three analogs was >90%.
8
Synthesis and Purification of SHLP Peptides
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Small humanin-like peptide 1 to 6 (SHLP1-6) were purchased from AnyGen [http://www.anygen.com/eng/ ]. For each peptide, the Fmoc-Ser (tBu)-Wang resin was allowed to swell in dimethylformamide (DMF). The amino acid was then deprotected using a solution of 20% piperidine in DMF. Coupling was carried out using Hexafluorophosphate Benzotriazole Tetramethyl Uronium (HBTU), N-methylmorpholine (NMM) in DMF. Stepwise deprotection and coupling of amino acids was repeated until the desired peptides was synthesized. The peptides were cleaved from dried resin using a trifluoroacetic acid (TFA) solution containing 2.5% 1, 2 ethanedithiol (EDT), 5% thioanisole and 5% distilled water. Crude peptides were precipitated in ether, and dried under vacumm. Peptides were purified using commercial columns (YMC-Triart C18/S-5 μm/12 nm.5μm (20 x 250 mm) and each fraction was obtained at a flow rate of 1 mL/min. The collected fractions were analyzed via HPLC (Shimadzu HPLC LabSolution) and MALDI-TOF MS (AXIMA Assurance, Shimadzu). The fractions containing the pure peptide were mixed and then lyophilized. SHLP2 were freshly prepared at 1 µg/µL by diluting in saline, and were freshly used for mice. The information of SHLP peptide sequence used in study were listed in Supplementary Table 3 .
9
Folate-Conjugated Oligonucleotide Synthesis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Folate was conjugated to the amino-modified overhang complement strand, as described by Erben et al. [15 (link)] Sulfo-SMCC and SPDP were used in the synthesis process as heterobifunctional cross-linkers. Amino-modified oligos overhang complement was re-suspended in phosphate buffered saline (137 mM NaCl 2.7 mM KCl, 8 mM Na2HPO4 and 2 mM K2HPO4, PBS pH 7.4), at a final concentration of 1mM. The DNA solution was combined with a saturated sulfo-SMCC solution (2.9 mg/mL) at 1:2 (v/v) in PBS (pH 7.4), and incubated for 1 h. Bio-Rad Micro Bio-Spin P-6 columns were used to remove excess cross-linkers. Folate was dissolved in PBS (pH 7.4) to 1 mM, combined with SPDP solution (5 mg/mL in DMSO) at 25:2 (v/v) and incubated for 1 h. Tris(2-carboxyethyl) phosphine hydrochloride was dissolved in PBS (pH 7.4) and added to folate solution at 1:10 (v/v). After incubation for 30 min, DNA and folate solutions were combined and incubated at 4 °C overnight. N-ethylmaleimide was dissolved in PBS (pH 7.4) and added to DNA-folate conjugate solution at 1:25 (v/v), to bind any unreacted thiol groups. After incubation for 30 min, a Bio-Rad Micro Bio-Spin P-6 column was used to transfer the conjugates into TM buffer (10 mM Tris-HCl and 5 mM MgCl2, pH 8). The product was confirmed by using MALDI-TOF-MS (Shimadzu, Japan).
10
Conjugation of CPP-PVGLI-PEG5000 to DSPE-PEG2000-MAL
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
At pH 6.8, the maleimide group of DSPE-PEG2000-MAL efficiently reacted with the sulfhydryl group of CPP-PVGLI-PEG5000 peptide (1:1, a molar ratio), and then adjusted pH of the reaction solution to 9.0 with N-methylmorpholine. The reaction solution was stirred overnight at room temperature (RT), and dialyzed overnight (MWCO. 8000 Da) to remove unbound material. Subsequently, the solution was lyophilized and stored at −20°C. Then, it was characterized by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS, Shimadzu, Japan).
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!