Zip-Tips for trapping sulfhydryl-containing peptide segment and preparation method thereof

A peptide and sulfhydryl technology, applied in the field of Zip-Tips (separation tips) and its preparation, can solve the problems of multiple experimental steps, separation speed and separation efficiency that do not meet the needs of rapid development, long time and other problems, and achieve a wide range of applications. The prospect, the production method is simple and novel, and the cost is low

Inactive Publication Date: 2013-08-07
FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the current limited separation speed and separation efficiency do not meet the needs of rapid development in the future, so there is an urgent need to develop a simpler and more efficient separation technology
For example, using related chromatographic techniques, sulfhydryl-containing peptides can be captured by disulfide bond exchange or derivatization methods, but these methods generally require many experimental steps and take a long time

Method used

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  • Zip-Tips for trapping sulfhydryl-containing peptide segment and preparation method thereof
  • Zip-Tips for trapping sulfhydryl-containing peptide segment and preparation method thereof
  • Zip-Tips for trapping sulfhydryl-containing peptide segment and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1. Preparation of novel Zip-Tips for capturing thiol-containing peptides.

[0021] (1) Pretreatment of the tip: heat the tweezers with a candle, and then flatten the end of the common tip.

[0022] (2) Preparation of gold-modified polymers: 10.02 mg of epoxy-based materials and 1.59 mg of glutathione were dissolved in 1 mL of borax buffer solution with pH=9-10, shaken at 37 ℃ for 24 hours, and then centrifuged to obtain The precipitate was reacted with 1 mL of gold colloid that had been synthesized at 37 °C for another 1 hour, and then centrifuged.

[0023] (3) Preparation of new Zip-Tips: Fill the prepared gold-modified polymer into the pretreated tip with a pipette under pressure. At this time, the liquid flows out, while the filler remains in the tip due to its large particle size. .

Embodiment 2

[0024] Example 2. Zip-Tips are applied to capture peptides containing sulfhydryl groups, and combined with MALDI-TOF-MS for efficient and rapid analysis.

[0025] (1) Sample injection: inject the sample from the upper end of the Zip-Tips with a pipette gun, and apply pressure through the pipette gun to make the sample flow through the packing slowly.

[0026] (2) Elution: first use NH 4 HCO 3 (10mM) + 5% acetonitrile (volume ratio) to wash away weakly bound interfering substances, and then use 70% acetonitrile and 0.1% TFA (volume ratio) to elute peptides containing sulfhydryl groups from Zip-Tips.

[0027] (3) Spotting the target: pipette 1uL of the analyte onto the MALDI target plate, and let it dry naturally in the air at room temperature; take 0.5uL of the CHCA solution and drop it on the analyte layer, and let it dry naturally in the air at room temperature.

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Abstract

The invention belongs to the technical field of isolation analysis, and particularly relates to a Zip-Tips (isolation gun) for trapping a sulfhydryl-containing peptide segment and a preparation method thereof. The method provided by the invention comprises the following steps: oscillating a polymer material containing epoxide group and glutathione in an alkaline buffer solution, centrifuging, carrying out oscillating reaction on the precipitate and synthesized gold size, centrifuging to obtain a gold-modified polymer material, and finally, filling the gold-modified polymer material in the gun; and flattening the tail end of the gun with heated tweezers while ensuring that only liquid flows out but the filler can not spill over, thereby obtaining the novel Zip-Tips for sensitively trapping a sulfhydryl peptide segment. The method provided by the invention has the advantages of simple operation steps and very low cost, can be used repeatedly, and can be used for quickly and sensitively analyzing the sulfhydryl-containing peptide segment when being in combination with high-flux MALDI-TOF-MS (matrix-assisted laser desorption / ionization time-of-flight mass spectrometry).

Description

technical field [0001] The invention belongs to the technical field of separation and analysis, and in particular relates to Zip-Tips (separation tip) used in proteomics sample pretreatment steps and a preparation method thereof. Background technique [0002] In the fields of biochemistry and biomedicine, mass spectrometry helps to discover and detect many important peptides. Peptides and materials are captured and separated by a specific force, and then analyzed quickly and efficiently with mass spectrometry. Among them, MALDI-TOF-MS (matrix-assisted laser desorption ionization time-of-flight mass spectrometry) has become a very popular analytical technique due to its advantages of fast, sensitive, and high-throughput. However, before the peptide mixture is detected by MALDI-TOF-MS, the peptide mixture needs to be separated and enriched for better analysis. This means that the means of separation and enrichment plays a very important role in MALDI-TOF-MS analysis. However...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N1/28G01N1/34
Inventor 高明霞张祥民万丹陈奇
Owner FUDAN UNIV
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