An mrna rabies vaccine
A technology for rabies virus and nucleic acid vaccines, applied in DNA/RNA vaccination, antisense single-stranded RNA viruses, vaccines, etc., can solve the problems of poor translation efficiency and low antigenic activity, and achieve the effect of reducing production costs
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[0090] The present invention provides a method of preparing the rabies viral nucleic acid vaccine, and the vaccine carrier and the optimized RVG mRNA provided by the present invention are mixed to obtain rabies virus nucleic acid vaccines. This method is simple and easy to operate, which greatly reduces production costs.
[0091] In a preferred embodiment, the vaccine carrier is a cationic lipid nanoparticle in the rabies virus nucleic acid vaccine according to the present invention, and the method of preparation of rabies virus nucleic acid vaccine includes:
[0092] (a) is dissolved in an organic solution in an organic solution in an organic solution according to the proportion of protonated cationic lipids, structural lipids, auxiliary lipid, and surfactant;
[0093] (b) Optimization of RVG mRNA is dissolved in PBS or citrate solution to obtain aqueous phase;
[0094] (c) The aqueous phase obtained by the organic phase and (b) obtained by (A) were mixed to give a mixed liquid, ...
Embodiment 1
[0106] Example 1 Method for preparing rabies virus nucleic acid vaccine
[0107] The mRNA of SEQ ID NO.2 was dissolved in the pH 4 citrate buffer, and the concentration was adjusted to 0.1 mg / mL to obtain aqueous phase.
[0108] The formulation ratio DLIN-MC3-DMA, cholesterol, DSPC and PEG-DMG were dissolved in anhydrous ethanol, and the total lipid concentration was adjusted to 6 mg / mL to obtain an organic phase.
[0109] The organic phase and the aqueous phase are mixed with a volume ratio of 1: 3 using a microfluidic device. The flow rate is 12.0ml / min when mixed with a microfluidic device.
[0110] The mixture immediately eliminated 50-100 times with pH 7.4 PBS solution, and the ethanol component in the solution was removed and concentrated to a mRNA concentration of about 100 μg / mL using tangential flow filtration (TFF) to obtain lipid nanoparticles wrap mRNA. It is a rabies virus nucleic acid vaccine.
Embodiment 2
[0111] Example 2 lyophilized preparation
[0112] The rabies virus nucleic acid vaccine in Example 1 was prepared into a lyophilized preparation, and the lyophilized protective agent sucrose and surfactant Tween 20, the process included pre-freeze, once lyophilized and secondary lyophilized: pre-freeze temperature is -50 ° C, temperature for 5 hours. The lyophilized temperature was -40 ° C for 24 hours, the secondary lyophilized temperature was 10 ° C for 17 hours, and the vacuum degree of lyophilization was 40 μBar.
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