Urine protein marker for diagnosing myeloblastoma and application thereof
A medulloblastoma, urine protein technology, applied in the field of clinical diagnosis, can solve the problem of no urine proteome or biomarker report
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Embodiment 1
[0050] Example 1. Collection and preparation of urine samples for mass spectrometry detection
[0051] 1. Enrolled patients and sample collection
[0052] In the biomarker discovery phase, urine was collected from nine children with medulloblastoma before and after tumor resection. These patients were all diagnosed at Beijing Tiantan Hospital Affiliated to Capital Medical University and underwent surgery there. The inclusion criteria for children were as follows: age ≤ 16 years old, postoperative pathologically confirmed medulloblastoma, and did not receive any treatment (radiotherapy and chemotherapy) before operation and during sample collection. The preoperative urine was collected before the operation, and the postoperative urine was collected 1-2 weeks after the operation. At this stage, the urine of 9 healthy children was also collected as a control group. In order to verify the results, the urine of 29 children with medulloblastoma before and after surgery, the urine...
Embodiment 2
[0070] Example 2. Urine protein profile analysis
[0071] 1. TMT (Tandem mass tag, tandem mass spectrometry tag) labeled quantitative proteomics analysis
[0072] 1.1 TMT labeling and offline liquid phase separation
[0073] 1) TMT quantitative labeling
[0074] 3 preoperative and corresponding postoperative samples, 3 healthy control samples, a total of 9 peptide samples were randomly labeled as 127N, 127C, 128N, 128C, 129N, 129C, 130N, 130C, 131. 10-plex Tandem Mass Tags (TMT) reagents were used according to the manufacturer's instructions (Thermo Fisher Scientific). The mixed samples of preoperative, postoperative and healthy control groups were labeled with 126TMT.
[0075] 2) Off-line high pH reversed-phase high-performance liquid chromatography for separation of TMT-labeled peptides
[0076] TMT-labeled samples were mixed and fractionated using a reversed-phase HPLC column (Waters, 4.6mm×250mm, Xbridge C18, 3μm). Put the sample on the chromatographic column and put ...
Embodiment 3
[0085] Example 3. Pathway analysis of differential proteins (Ingenuity Pathway Analysis, IPA)
[0086] All differentially expressed proteins were analyzed using IPA software (Ingenuity Systems, Mountain View, CA). These proteins are classified into disease-associated and function-associated categories and are profiled in biological pathways in Ingenuity and other databases. Sort by z-score and p-value, respectively.
[0087] Functional analysis of differentially expressed proteins
[0088] These 114 differential proteins were analyzed by IPA, and these differential proteins can be divided into two categories related to disease and function, such as tumor cell migration, apoptosis, central nervous system metastasis, stage III tumors, recurrent tumors, head Neck tumors and metastases suggest that these proteins may be associated with nervous system tumors. Proteins enriched on biological pathways associated with: clathrin-mediated endocytic signaling, glycolysis I, role of ti...
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