Ovarian-cancer marker detection kit based on PRM (Parallel Reaction Monitoring) detection and detection method
A detection kit and detection method technology, applied in the field of ovarian cancer marker detection, can solve the problems of time-consuming, poor sensitivity and specificity, complicated operation, etc., and achieve the effect of high safety, strong specificity, and simple operation
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Embodiment 1
[0031] Embodiment one: chromatographic analysis column.
[0032] Such as figure 1 A chromatographic analysis column suitable for the detection of ovarian cancer PRM is shown, which includes a column tube (1), a filler (2) and a tip (3), the filler (2) is filled in the column tube (1), and the tip (3) Cover one end of the column tube (1); the length of the column tube (1) is 120mm, the outer diameter is 360um, and the inner diameter is 75um; the filler (2) is a C18 reverse phase filler with a particle size of 3um .
Embodiment 2
[0033] Example 2: Ovarian Cancer Marker Detection Kit
[0034] A detection kit for ovarian cancer markers based on PRM detection, including reagents for conventional mass spectrometry detection, heavy isotope-labeled standard peptide test tubes, the above-mentioned chromatographic analysis column, and an aqueous solution containing 0.1% formic acid; the heavy isotope-labeled standard peptide The test tube contains ten unique sequences among thyroxine, apolipoprotein A–1, β2-microglobulin (Beta2M), transferrin, and cancer antigen 125, where each protein has two peptide sequences, namely VEHSDLFSK, For VNHVTLSQPK, EFQLFSSPHGK, HQTVPQNTGGK, GSPAINVAVHVFR, AADDTWEPFASGK, DLATVYVDVLK, DYVSQFEGSALGK, LGIWDDFIPK, and ELPLLSPPQDK, the heavy isotope-labeled amino acids were located on arginine and lysine at the C-terminal of the peptide sequence, respectively.
[0035] As a preferred embodiment, the concentration of each standard peptide in the heavy isotope-labeled standard peptide te...
Embodiment 3
[0036] Example 3, detection of ovarian cancer markers.
[0037] Step S10, take a heavy isotope-labeled standard peptide test tube, add 90-110uL of dissolving buffer into it, shake and mix, use a centrifuge to centrifuge at a speed of 13000g for 60s, and take the supernatant for later use;
[0038] Step S20, taking 0.1 mg of serum proteolytic peptides and adding 100 uL of dissolving buffer, shaking and mixing to make a serum peptide solution for later use;
[0039] Step S30, taking 2uL of the serum peptide solution prepared in step S20, and adding 10uL of the heavy isotope-labeled standard peptide solution prepared in step S10, mixing evenly, taking 4uL for chromatographic injection;
[0040] The chromatographic separation gradient was set as follows:
[0041] time (min)
[0042] The PRM detection ion table is set as follows:
[0043] M / Z
[0044] The mass spectrometer was set up as follows:
[0045]First-level mass spectrometry, scanning range: 500-800m / ...
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