A kit for magnetic particle chemiluminescence immunodetection of CA125 surface TN antigen and its preparation method
A chemiluminescence immunity, CA125 technology, applied in chemiluminescence/bioluminescence, analysis and fermentation through chemical reaction of materials, can solve problems such as long reaction time, affecting detection sensitivity and linear range, and Fab segment influence, etc. Achieve the effect of avoiding non-specific adsorption, avoiding false positive phenomenon, and improving the linear range
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Embodiment 1
[0076] The kit for magnetic particle chemiluminescence immunodetection of CA125 surface Tn antigen in this example includes magnetic separation reagent coupled with Fab fragment of Ov185 monoclonal antibody, Biotin-VVA reagent containing sialidase, SA-Biotin-ALP reagent, chemical Luminescence substrate solution, washing solution and calibrator working solution.
[0077] The preparation method of the kit for magnetic particle chemiluminescence immunodetection of Tn antigen on the surface of CA125 in the present embodiment includes the following operation steps:
[0078] 1. Preparation of magnetic separation reagent coupled with Fab fragment of Ov185 monoclonal antibody:
[0079] 1. Preparation of immobilized papain:
[0080] Ⅰ. Accurately measure 50mg Fe 3 O 4 Nanomagnetic particles were placed in a 10ml polystyrene plastic test tube, placed on a magnetic rack, and the supernatant was removed after precipitation, and washed twice with 5ml of pure water. Na 2 HPO 4 , 2mM K...
Embodiment 2
[0146] Unlike Example 1,
[0147] Step Ⅰ is to accurately measure 50mg Fe 3 O 4 Nanomagnetic particles were placed in a 10ml polystyrene plastic test tube, placed on a magnetic rack, and the supernatant was removed after precipitation, washed twice with 10ml pure water, and after washing was completed, 10ml of 0.01M PBS buffer (0.14MNaCl, 3mM KCl, 10mM Na 2 HPO 4 , 2mM KH 2 PO 4, pH=7.2±0.05), then add papain with 0.05 times the mass of magnetic beads, and mix at room temperature for 1.5 hours;
[0148] Step II is to place the magnetic beads on a magnetic rack after mixing, remove the supernatant after precipitation, add 7.5 ml of 0.2 mM hydrochloric acid buffer, and mix at room temperature for 10 minutes;
[0149] Step III is to add 0.3mg 0.5M HEPES buffer (pH=7.4±0.1) after mixing, adjust pH to 7.3±0.1, and mix at 37°C for 2.5 hours;
[0150] Step IV is adding 225 mg of glycine after mixing, and mixing at 37°C for 1 hour;
[0151] Step 5: After mixing, place the magn...
Embodiment 3
[0167] Unlike Example 1,
[0168] Step Ⅰ is to accurately measure 50mg Fe 3 O 4 Nanomagnetic particles were placed in a 10ml polystyrene plastic test tube, placed on a magnetic rack, and the supernatant was removed after precipitation, washed twice with 25ml of pure water, and 25ml of 0.01M PBS buffer (0.14MNaCl, 3mM KCl, 10mM PBS) was added after washing. Na 2 HPO 4 , 2mM KH 2 PO 4 , pH=7.2±0.05), then add papain with 0.04 times the mass of magnetic beads, and mix at room temperature for 1 hour;
[0169] Step II is to place the magnetic beads on a magnetic rack after mixing, remove the supernatant after precipitation, add 15 ml of 0.2 mM hydrochloric acid buffer, and mix at room temperature for 10 minutes;
[0170] Step III is to add 0.125mg 0.5M HEPES buffer (pH=7.4±0.1) after mixing, adjust pH to 7.3±0.1, and mix at 37°C for 2.5 hours;
[0171] Step IV is adding 75 mg of glycine after mixing, and mixing at 37°C for 1 hour;
[0172] Step V: After mixing, place the ma...
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