Atherosclerosis detection reagent and preparation method thereof
An atherosclerosis and detection reagent technology, applied in the fields of biotechnology and clinical diagnosis, can solve the problems of not well reflecting plaque vulnerability, elevated metabolic activity, false positives and false negatives, etc. Effects of high and low levels of inflammation
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Embodiment 1
[0052] Example 1. Synthesis of polypeptide derivative atherosclerosis detection reagent
[0053] Taking a specific detection reagent compound of the present invention as an example, its synthesis steps are as follows:
[0054] (1) Using solid-phase synthesis method to synthesize Fmoc-protected derivative peptide sequence on 2-chlorotrityl chloride resin (Fmoc-Pip-Lys(Boc)-Lys-(Boc)-Arg(Pbf)-Pro-Hyp (tBu)-Gly-Cpg-Ser(tBu)-D-Tic-Cpg). The resin was treated with 20% piperidine / DMF to remove the Na-Fmoc protecting group. Activate 4-(bromomethyl)phenylacetic acid (3 equivalents) with N,N'-diisopropylcarbodiimide (3 equivalents) in 5mL N-methyl-2-pyrrolidone, and then couple to N- End. Subsequently, 1,4,7-triazacyclononane-1,4-bis(tert-butyl acetate) (3 equivalents) was alkylated with a secondary amine in 5 mL of N-methyl-2-pyrrolidone Coupled to the polypeptide sequence. The peptide was cleaved from the resin with 95% TFA, and the synthesized peptide was purified by HPLC.
[0055] (...
Embodiment 2
[0057] Example two, stability analysis
[0058] Stability is an important indicator of in vivo diagnostic reagents. The detection reagents prepared in Example 1 were incubated in phosphate buffer and mouse serum at 37°C for 5, 15, 30, 60, and 120 minutes, and quenched with 70% acetonitrile at the end. It was quenched and centrifuged for 20 minutes, and the stability of the polypeptide was determined by radio HPLC. The stability of the polypeptide in phosphate buffer and mouse serum can maintain at least two half-lives, indicating that the polypeptide has good stability.
Embodiment 3
[0059] Example 3: Specific identification of atherosclerotic plaque
[0060] Feeding ApoE knockout mice with high-fat diet is a common method to construct atherosclerosis models. After ApoE- / - mice were fed high-fat diet for 40 weeks, conventional methods were used to identify high-risk atherosclerosis (easy to fall off, low calcification) individuals and low-risk atherosclerosis (not easy to fall off, high calcification) individuals. The detection reagent 7.4MBq prepared in Example 1 was injected through the tail vein, and PET / CT imaging was performed 20 minutes later. image 3 The results of CT and PET / CT imaging are given, indicating that the polypeptide has high radiation signals at the plaques that are easy to fall off, but there is almost no radiation signal at the calcified plaques visible on CT, which realizes the treatment of atherosclerosis and easy fall off plaques. The specific identification.
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