Characteristic protein marker composition for screening thalassemia, mass spectrum model and application thereof

A technology for thalassemia and characteristic proteins, which is applied in the field of characteristic protein marker compositions for screening thalassemia, can solve problems such as interference with glycosylated hemoglobin detection, and achieve the effects of simple sample pretreatment, less sampling volume, and favorable application transformation

Active Publication Date: 2020-11-17
融智生物科技(青岛)有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In addition, the presence of abnormal hemoglobin often interfe

Method used

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  • Characteristic protein marker composition for screening thalassemia, mass spectrum model and application thereof
  • Characteristic protein marker composition for screening thalassemia, mass spectrum model and application thereof
  • Characteristic protein marker composition for screening thalassemia, mass spectrum model and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] Example 1 Establishment of MALDI-TOF mass spectrometry detection method for characteristic protein combinations of thalassemia and calculation of peak area ratios of different protein spectra

[0053] (1) Reagent preparation

[0054] Sample diluent: deionized water for dilution of blood samples.

[0055] Mass spectrometry matrix solution: 10 mg / mL sinapinic acid (SA) solution (acetonitrile: 0.1% trifluoroacetic acid = 4:6).

[0056] (2) Sample pretreatment

[0057] ① The sample is venous blood collected in an anticoagulant tube containing EDTA, and stored at -80°C within 48 hours. Use a pipette to draw 2 μL of the mixed blood sample into a 1.5 mL EP tube, add 998 μL of sample diluent to the tube, vortex and mix, and then centrifuge at 3000 (rpm) for 30 seconds.

[0058] ②Take the centrifuged upper layer liquid and 10mg / mL mass spectrometry matrix solution in a volume ratio of 1:9, mix by vortexing.

[0059] ③ Using the dry drop method, pipette 2.5 μL of the sample-m...

Embodiment 2

[0073] Example 2 Establishment of mass spectrometry model for thalassemia screening

[0074] (1) Using the detection method established in Example 1, analyze 100 cases of normal control samples, 52 cases of β-thalassemia samples (50 samples of β-thalassemia carriers and 2 samples of β-thalassemia patients), and α and β-thalassemia samples 35 cases of poor (α / β thalassemia) samples.

[0075] Identify the characteristic proteins in the sample by software analysis system (α globin=15127m / z, β globin=15868m / z, δ globin=15924m / z, γ globin=15995m / z, carbonic anhydrase I=28762m / z, the allowable mass-to-charge ratio error is ±0.1%) mass spectrum peak. Normal control samples such as figure 1 As shown, relative to normal control samples, β-thalassemia carries (as figure 2 Shown) The relative intensity of the δ-globin peak was significantly increased. β-thalassemia patients (such as image 3 Shown) the relative intensity of the β-globin peak decreased significantly, and the relati...

Embodiment 3

[0089] Repeatability evaluation of embodiment 3 methodology

[0090] Based on the detection method established in Example 1, 3 normal control samples and 3 α-thalassemia samples were selected. Under the same experimental conditions, the samples were repeatedly measured 8 times between batches to obtain the δ / β and CA I of the samples. / δ ratio. Calculate the average value (AVG) and relative standard deviation (RSD) of the results, evaluate the repeatability of the experimental process, as shown in table 4, the RSD of δ / β is between 4.66%-5.14%, the RSD of CA I / δ is in Between 3.15% and 6.53%, it shows that the repeatability of the δ / β and CA I / δ ratios determined by this method is good, and meets the requirements of clinical screening.

[0091] Table 4. Repeatability analysis of δ / β and CA I / δ ratios

[0092]

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Abstract

The invention discloses a characteristic protein marker composition for screening thalassemia, a mass spectrum model and application thereof. The invention firstly discloses a characteristic protein marker composition for screening thalassemia. The composition comprises alpha globin, beta globin, delta globin, gamma globin and/or carbonic anhydrase I, and the sequences of the alpha globin, the beta globin, the delta globin, the gamma globin and/or the carbonic anhydrase I are respectively shown as SEQ ID NO.1-5. The invention further discloses a mass spectrum model containing the marker composition and application of the mass spectrum model in preparation of products for screening thalassemia. Based on the MALDI-TOF mass spectrum technology, the method can screen beta thalassemia by analyzing the mass spectrum peak areas and ratios of different characteristic proteins, carries alpha and beta thalassemia, alpha thalassemia and abnormal hemoglobin samples, and has the characteristics ofsimple sample pretreatment, high detection flux, less reagent consumables, low cost, less sampling amount, high sensitivity and the like; and large-scale population screening can be carried out in regions with high occurrence rate of thalassemia.

Description

technical field [0001] The present invention relates to the field of biotechnology. More specifically, it relates to a characteristic protein marker composition, a mass spectrometry model and an application thereof for screening thalassemia. Background technique [0002] Thalassemia (referred to as thalassemia), also known as thalassemia, is a genetic hemolytic anemia disease, α-thalassemia and β-thalassemia are the most important types of diseases. Alpha-thalassemia is a hereditary hemolytic anemia caused by the deficiency or mutation of the alpha-globin gene, which leads to the disorder of alpha-globin chain synthesis. According to the type of gene defect, α-thalassaemia is divided into deletion type and non-deletion type. The most common missing α-thalassemia in China includes -- SEA , -α 3.7 and-alpha 4.2 , non-deletion α-thalassemia includes HbConstant Spring (HbCS), Hb Quong Sze (HbQS) and Hb Westmead (HbWS). According to the degree of gene defect, α-thalassemia ...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N27/62
CPCG01N33/6893G01N33/6851G01N27/62G01N2800/22
Inventor 孙德慧王玉玺李运涛周晓光
Owner 融智生物科技(青岛)有限公司
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