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Liquid phase chip kit for detecting refractory epilepsy
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A liquid phase chip and kit technology, applied in the field of medicine and biology, can solve the problems of lack of high-level research on diagnostic markers, and achieve rapid detection, high-throughput detection, and good stability
Inactive Publication Date: 2019-04-26
WEST CHINA HOSPITAL SICHUAN UNIV
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Problems solved by technology
However, high-level research on MMPs as diagnostic markers at home and abroad is still very scarce.
Method used
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[0029] (1) Take the microsphere suspension (magnetic microspheres, bio-rad), vortex and sonicate for about 20 seconds, take 250 μL (about 5×10 6 microspheres) in a clean centrifuge tube (USA, Axygen), marked with the microsphere model and protein name; then put it into a centrifuge, separate at 14000g for 4min at room temperature, and then suck off the supernatant;
[0030] (2) Add 100 μL ddH to the centrifuge tube 2 O, vortex and sonicate for about 20s in turn, then separate at 14000g for 4min at room temperature, and suck off the supernatant;
[0031] (3) Add 160 μL of NaH with a pH of 6.2 and a concentration of 0.1M to the centrifuge tube 2 PO 4 , vortex and sonicate for about 20s, then add 20 μL N-hydroxysulfosuccinimide (S-NHS), vortex to mix, then add 20 μL 1-ethyl-(3-dimethylaminopropyl) Carbodiimidehydrochloride (EDC), vortex and sonicate for about 20s in tu...
Embodiment 2
[0039] Embodiment 2. The kit is used to detect refractory epilepsy
[0040] 1. Detection method
[0041] (1) Take out all the reagents from the kit before use and let it cool to room temperature;
[0042] (2) Take out the hippocampus tissues of rhesus monkeys with epilepsy and healthy rhesus monkeys, place them on ice to melt, vortex and mix well, and dilute the hippocampus tissues 20 times in a 96-well plate;
[0043](3) Prepare TIMP-1, MMP2, MMP3, and MMP9 standard proteins, take 7 EP tubes and mark them with S1, S2, S3, S4, S5, S6, and S7 respectively, and perform 2-fold serial dilutions, each standard proteindilution Afterwards, vortex to mix and replace the pipette tip;
[0044] (4) Take a 96-well plate, turn on the plate washer--Prime--rinse(channel2)--Prime--place the plate--Run5: MAGX3;
[0045] (5) Take the prepared capture antibody-coated microspheres, vortex and mix for 20s, sonicate for 20s, add 2500 microspheres per well into 1% PBS, vortex and mix for 20s, so...
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Abstract
The invention discloses a liquid phasechip kit for detecting the refractoryepilepsy. The kit comprises coated microspheres, biotin labeled detection antibodies, streptomycin R phycoerythrin, a reaction buffer and a reaction dilution solution; the coated microspheres comprise microspheres coated with TIMP-1 capture antibodies, microspheres coated with MMP2 capture antibodies, microspheres coatedwith MMP3 capture antibodies and microspheres coated with MMP9 capture antibodies, and the microspheres coated with different capture antibodies have different color codes; the detection antibodies include a TIMP-1 detection antibody, an MMP 2 detection antibody, an MMP 3 detection antibody and an MMP 9 detection antibody; the reaction buffer is a weakly alkaline PBS buffer; a dilution buffer is aPBS buffer containing 0.05wt%-0.15wt% BSA and has a pH value of 7.0-7.5. By using the kit, the purpose of timely discovery and accurate diagnosis of the epilepsy can be achieved.
Description
technical field [0001] The invention belongs to the technical field of medicine and biology, and in particular relates to a liquid phasechip kit for detecting refractoryepilepsy. Background technique [0002] Epilepsy is a chronic recurrent short-term brain dysfunction syndrome characterized by recurrent epileptic seizures caused by highly synchronized abnormal discharges of brain neurons. Epilepsy is one of the common diseases of the nervous system, second only to stroke in prevalence. The incidence of epilepsy is related to age. It is generally believed that the prevalence rate is the highest within 1 year old, and gradually decreases after 1 to 10 years old. The male to female ratio of epilepsy patients in my country is 1.15-1.7:1. [0003] Refractory epilepsy, also known as intractable epilepsy, usually refers to no progressive disease or space-occupying lesions of the central nervous system, but clinical protraction, after more than 2 years of regular antiepileptic ...
Claims
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