Preparation of organic phosphonic acid functionalized magnetic nano material and application in protein enrichment
An organic phosphonic acid and magnetic nanotechnology, which is applied in the fields of organic material/organic magnetic material magnetism, material separation, material analysis, etc., can solve the problems of cumbersome steps and non-green, and achieve the ability to bind and enrich proteins. The effect is good, the synthesis method is simple
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Embodiment 1
[0028] Embodiment 1, Zr 4+ -Synthesis of NO2PA-MS and enrichment and detection of phosphorylated proteins
[0029] Weigh 2 mg of magnetic nanoparticles wrapped with aminosilane, and disperse them in 1 mL of secondary water under the action of ultrasound, then weigh 6 mg of NO2PA, EDC and NHS, dissolve them in 1 mL of LPBS buffer solution (pH5.6), add and mix, Sonicate for 30 minutes at room temperature. Then, under an external magnetic field, wash and separate, and wash three times with 1 mL of secondary water. Then add 1 mL of 50 mM ZrOCl 2 Hydrochloric acid solution (0.2M), stirred vigorously at room temperature for 20 minutes. Then wash three times with 0.2M hydrochloric acid solution and secondary water respectively.
[0030] Draw 50μL (1mg / mL) of the prepared Zr 4+ -NO2PA-MS particle suspension, add 50μL (10 -7 M or 10 -8 M) β-casein digestion solution, shake and mix at room temperature for 30 seconds. Then, under an external magnetic field, it was fully washed th...
Embodiment 2
[0032] Example 2, Zr 4+ -Synthesis of NC2P-MS and enrichment and detection of phosphorylated proteins
[0033] Weigh 1 mg of magnetic nanoparticles wrapped with aminosilane, and disperse them in 1 mL of secondary water under ultrasonication, then weigh 3 mg of NC2P and dissolve them in 1 mL of LPBS buffer solution (pH 5.6), weigh 3 mg of EDC and 3 mg of NHS, and add the mixture , ultrasonic reaction at room temperature for 30 minutes. Then, under an external magnetic field, wash three times with 1 mL of secondary water. Then add 500 μL of 50 mM ZrOCl 2 Hydrochloric acid solution (0.2M), stirred vigorously at room temperature for 20 minutes. Then wash three times with 0.2M hydrochloric acid solution and secondary water respectively.
[0034] Aspirate 50μL (1mg / mL) of the prepared Zr 4+-NC2P-MS suspension, add 50μL (10 -7 M or 10 -8 M) β-casein digestion solution, shake and mix at room temperature for 30 seconds. Then, under an external magnetic field, it was fully washe...
Embodiment 3
[0036] Example 3, Zr 4+ -Synthesis of CP-MS and its application in the enrichment and detection of phosphorylated proteins
[0037] Weigh 1mg of magnetic nanoparticles wrapped with aminosilane, disperse in 1mL of secondary water under the action of ultrasound, then add PBS buffer solution (pH 5.6) containing 3mg of CP, then weigh 3mg of EDC and 3mg of NHS, add the above mixture, Sonicate for 30 minutes at room temperature. Then, under an external magnetic field, wash three times with 1 mL of secondary water. Then add 500 μL of 50 mM ZrOCl 2 Hydrochloric acid solution (0.2M), stirred vigorously at room temperature for 20 minutes. Then wash three times with 0.2M hydrochloric acid solution and secondary water respectively.
[0038] Aspirate 50μL (1mg / mL) of the prepared Zr 4+ -CP-MS suspension, add 50μL (10 -7 M or 10 -8 M) β-casein digestion solution, shake and mix at room temperature for 30 seconds. Then, under an external magnetic field, it was fully washed three times...
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