Analysis method of target board derivatization and MALDI-TOF-MS of reducible carbohydrate chain

A MALDI-TOF-MS and derivatization technology, applied in the field of glycobiology, can solve the problems of not being able to detect neutral sugar chains and acidic sugar chains at the same time, decrease in detection sensitivity, signal loss, etc., so as to improve ionization efficiency, Improved detection sensitivity and strong versatility

Active Publication Date: 2017-10-20
西安泰乐星生物科技有限公司
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Problems solved by technology

[0011] In order to solve the problem that the target derivatization method before MALDI-TOF-MS analysis of reducing sugar chains in the prior art cannot be applied to the detection of neutral sugar chains and acidic sugar chains at the same time, and for those containing unstable functional groups such as saliva Acidic sugar chains with acid groups are likely to cause sample degradation, signal loss, and detection sensitivity. On the one hand, the present invention provides a target derivatization method for reducing sugar chains; on the other hand, it provides the target derivatization method. The MALDI-TOF-MS analysis method of the target plate sample sample obtained by the method

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  • Analysis method of target board derivatization and MALDI-TOF-MS of reducible carbohydrate chain
  • Analysis method of target board derivatization and MALDI-TOF-MS of reducible carbohydrate chain

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specific Embodiment

[0100] The embodiments in the present invention are about the target plate derivatization method of reducing sugar chains and the MALDI-TOF-MS analysis method of the target plate spot sample after target plate derivatization. The research strategy of the sugar chain target plate derivatization method and MALDI-TOF-MS analysis method of the present invention is as follows:figure 1 as shown,

[0101] In the derivatization of the target plate of the present invention, Girard reagent T is selected, and the reaction conditions are mild. For groups with unstable functional groups, the derivatization conditions of the present invention can effectively inhibit or prevent sample degradation; the derivatization efficiency is close to 100%. Significantly improve the ionization efficiency of sugar chains in mass spectrometry detection, and thus have very high detection sensitivity. Girard T reagent is used as the derivatization reagent, because of its structure with hydrazine group and po...

Embodiment 1

[0103] Example 1: Target plate derivatization of maltodextrin (Maltodextrin) and MALDI-TOF-MS analysis after derivatization analysis

[0104] Maltodextrin (Maltodextrin) is a macromolecular reducing oligosaccharide. Weigh maltodextrin and use ultrapure water as a solvent to prepare a sugar chain solution with a mass concentration of 0.10 mg / mL; weigh Girard reagent T , be dissolved in the mixed solvent of methanol, acetic acid and water to obtain the acidic Girard reagent T solution, in the mixed solvent, methanol: acetic acid: water (v: v: v) = 1: 3: 6, and the mass concentration is prepared as 13.36 mg / mL of Girard's reagent T solution. The matrix DHB was dissolved in CH 3 CN: 0.1% TFA aqueous solution (v:v) = 1:1 mixed solvent, prepared as a matrix solution with a mass concentration of 20.00 mg / mL.

[0105] Pipette 0.5 μL each of the matrix DHB solution, reducing sugar chain solution, and Girard reagent T solution, and place them on the same point on the target plate ...

Embodiment 2

[0123] Example 2: Taking lactose as an example to optimize the derivatization conditions of the target plate

[0124] The target plate derivatization and MALDI-TOF-MS analysis of lactose were carried out according to the experimental procedure of Example 1, and the relationship between the amount of Girard reagent T and the amount of sugar chains was observed.

[0125] Weigh lactose, use ultrapure water as a solvent, and prepare a lactose solution with a mass concentration of 3.42 mg / mL; weigh Girard reagent T, dissolve it in a mixed solvent of methanol, acetic acid and water to obtain an acidic Girard reagent T solution, in a mixed solvent, methanol: acetic acid: water (v:v:v) = 1:3:6, prepared to have a mass concentration of 1.67mg / mL, 3.34mg / mL, 5.01mg / mL, 6.68mg / mL , 8.35mg / mL, 10.02mg / mL, 11.69mg / mL, 13.36mg / mL, 15.03mg / mL, 16.7mg / mL. Converted to molar concentration, that is, the molar concentration ratio of Girard reagent T solution and reducing sugar chain solution ...

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Abstract

The invention belongs to the technical field of glycobiology, and particularly relates a method of the target board derivatization of a reducible carbohydrate chain. The method comprises the following steps: preparing a reducible carbohydrate solution, a Girard reagent T solution and a matrix DHB solution separately, and applying the matrix DHB solution, the reducible carbohydrate solution and the Girard reagent T solution at the same point of an MALDI-TOF-MS instrument target board separately; placing the target board subjected to sample application at room temperature till a sample point is completely dried. The invention further provides an analysis method of the MALDI-TOF-MS of a sample subjected to derivatization. The method is high in derivatization efficiency, can effectively prevent the degradation of acid carbohydrate chains, can detect both neutral carbohydrate chains and acid carbohydrate chains in a positive ion mode without switching, and is high in universality and simple and more efficient.

Description

technical field [0001] The invention belongs to the technical field of glycobiology, and in particular relates to a target plate derivatization and MALDI-TOF-MS analysis method of reducing sugar chains. Background technique [0002] As the third type of biomacromolecules that build living organisms, carbohydrates have very important biological functions, such as providing energy substances for organisms, and are the main components of plant cell walls. Intercellular signal transduction, recognition, immune response, etc. play an important role. Glycosylation is one of the most important post-translational modifications of proteins, and plays a very important role in protein translation regulation and protein degradation. More than 50% of proteins exist in the form of glycoproteins, and more and more studies have shown that abnormal glycosylation can lead to human diseases. In the course of clinical treatment, the antibody drugs used are all glycosylated. [0003] At prese...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/06G01N30/72
CPCG01N30/06G01N30/72
Inventor 王仲孚王波张英
Owner 西安泰乐星生物科技有限公司
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