| The use of vaccine is a very important way to control disease nowadays with the development of livestock cultivation, however failed immunoprotection or incomplete immunoprotection occurs invariably. The factors are numerous, such as the quality of vaccine, immune program, bad organism health status, animals infected with immunosuppressive disease, stress, chronic intoxication caused by drug abuse. which all could make livestock be in inferior health status, and could not make normal immune response to vaccine stimulation, resulting in failed immunoprotection or uncomplete immunoprotection. Meanwhile as the development of molecular biology technology gene vaccine exposed for the past few years, the immunogenicity of this vaccine is weak, the common immunoenhancers are hard to improve the immunogenicity. So the development of efficient, harmless neotype immunoenhancers is becoming a problem demanding prompt solution.As a kind of mitogen Phytohemagglutinin (PHA) has intense stimulation to immune system. The extraction technics of PHA was investigated in this article. According to factors affecting the aqueous solution diffusion of PHA, 3 methods were designed and adopted, meanwhile reverse micelle was taken to extract PHA, compare each other. The results showed that the extraction rate of reverse micelle was higher than aqueous solution diffusion. The single factor analysis was taken to detect factors (pH, ionic strength of aqueous phase, extraction time) affecting reverse micelle extraction. The optimization of reverse micelle to extract PHA is: ionic strength of aqueous phase was 0.15moL·L-1, pH of aqueous phase was 4～6, extraction time was 4min, pH of aqueous phase in the back extraction was 9～10, back etraction time was 8min. DEAE Sephadex A-50 column chromatography and Sephadex G150 column chromatography were taken to purify PHA, the purified product was proved electrophoretically to be pure by SDS-PAGE.The immunological enhancement of Chinese Herbal Polysaccharides(CHPs) has been testified by versatile experiments. Hot water diffusion, Sevag deproteinization, macroporous resin depigmenting were employed to extract and purify Astragalus Polysaccharides(APS), Poria Wolf Polysaccharides(PWP),Indigowoad Root Polysaccharides(IRP), Angelica Sinensis Polysaccharides (ASP). Saccharinity of four CHPs detected by Phenol-Sulphuric Acid method was 86%, 72%, 79%, 83%, respectively.Mice splenic lymphocyte proliferation experiment in vitro was taken to testify the stimulatory function of PHA to lymphocytic, the final density 6.5～200μg·mL-1 was considered to promote splenic lymphocyte proliferation significantly, 50μg·mL-1 promotion arrived the peak, afterward descended a little as the increase of density. Promotion effects of four CHPs used alone or together with PHA were observed, the results showed that four CHPs used alone could promote the proliferation of splenic lymphocyte in vitro, synergistic effect was found when used together with PHA. Among the total, OD570 was maximum when APS used alone or together with PHA.Immunoenhancement of APS together with PHA in mice was detecded. 60 Kunming mouse were divided into 6 groups, including the high dose group, middle dose group, low dose group, PHA control group, APS control group and blank control group. They were intraperitoneally igiven for 5 days, till the 7th day, the clearance rate of charcoal particles was detected to evaluate index of phagocytosis, and spleen index and thymus index were calculated. The results showed that high and middle does of APS together with PHA could increase spleen index of mice significantly, but contribution to thymus was quiet; high and middle does of APS together with PHA could increase index of phagocytosis of mice extreme significantly. It was proved that APS together with PHA could enhance specific immunity function and nonspecific immunity function of mice. In conclusion, reverse micelle was found to be a good method to extract PHA in this essay and PHA together with APS can promote specific immunity function and nonspecific immunity function of mice.
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