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N-Glycan聚糖分子标签／N-Glycan Label Reagent

N-Glycan 标签.png
N-Glycan 聚糖标签: N_Glycan Label Reagent

 

用于N-linked糖蛋白（包括抗体)经过PNGase F糖苷酶酶切后溶液中聚糖的超快速标记。 标记后的聚糖具有灵敏的荧光和质谱离子化特性。使用LC-FLR or LC-MS的定量分析检测。该试剂独特的化学属性提供了更高的荧光定量和最高的质谱响应。

 

产品用途：

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  • 生物治疗蛋白N-聚糖表征分析

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产品特点：

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  • 质谱级干粉试剂

  • 标记效率高：3min

  • 热稳定性好：24小时

 

超快速30min标记流程：

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  1. 取 10ul. 1.5 ug/ul mAb 脱盐样品

  2. 加 10ul   3% BT  Surfactant w/5mM TCEP 变性还原缓冲液 

  3. 干浴 95°C 加热3min； 冷却至室温

  4. 加 10µl   1.5 unit/ul  rPNGase F糖苷酶， 干浴50°C 孵育15min

  5. 加 10ul    82 ug/ul N-Glycan Label标签试剂， 室温反应5min

  6. 加 15ul 无水DMF； 加45ul 乙腈

  7. LC-MS分析

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Part No.                     Name                              Size

HLS GLAB01W        N-glycan label-W        4.5 mg

HLS DMF001           Anhydrous DMF           200 μL

 

Description: N-glycan label is used in ultra-fast labeling of glycans of PNGase F glycosidase-digested N-linked glycoproteins (including antibodies). Labeled glycan has highly sensitive fluorescence characteristics and ionization properties by mass spectrometry, allowing for quantitative analysis using LC-FLR and LC-MS. The unique chemical properties of this label provide higher fluorescence quantification and mass spectrometry response.

Physical Appearance: Lyophilized powder

Molecular Weight: 427.1850 Da

Molecular Formula: C21H25N5O5    

Mass Shift: + C17H20N4O2 = 312.3663 Da 

Labeled N-Glycan (Da) = Released N-Glycan + Glycan Label Reagent - Reaction by-product ( C4H5NO3, MW: 115.0268) - H

Resuspension: Dissolve 4.5 mg N-Glycan label in 55 μL freshly prepared DMF solution to reach 82 μg/μL.

Storage Conditions: -20 °C refrigerator as lyophilized powder.

Shelf Life: 24 months at -20 °C.

Application: 4.5 mg N-Glycan labeling reagent can label five digested glycan samples.

Reference Procedure: 

  1. Add 10 μL desalted 1.5 μg/μL mAb samples into 0.6 mL low protein binding collection tube.

  2. Add 10 μL 3% BT Surfactant with 5 mM TCEP and 50 mM HEPES (pH 7.9) to the tube and mix well. 

  3. Heat at 95 °C for 3 min, then cool to the room temperature.

  4. Add 10 μL 1.5 U/μL rPNGase F and mix well.

  5. Incubated at 50 °C for 15 - 30 min, then cool to room temperature.

  6. Add 10 μL 82 μg/μL Glycan Label, incubate at room temperature for 5 min.

  7. Add 15 μL anhydrous DMF and 45 μL acetonitrile. Mix well.

Labeled N-glycan samples can be used for LC-FLR and LC-MS analysis.

 

| Quality Control

Product Composition: The ratio of N-Glycan Label: NHS compound is 3:2 analyzed by LC-QE HF.

Labeling Efficiency: 99.5% when 0.37 μg/μL mAb and 20 μg/μL N-Glycan tags are used to label 15 μg N-glycans released by glycoproteins.

Thermal stability: N-Glycan labeling reagent is stable in anhydrous DMF at room temperature for 8 h; Labeled glycoamines are stable at room temperature for 24 h, and can withstand 320 °C of mass spectrometry ionization temperature.

Chemical Instability: Unstable when concentrations of affinity reagents, SDS, Tris, DTT, amine, and mercaptans greater than 0.1 mM.

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