| Classical swine fever(CSF) is one of the major animal diseases in China,which causes great harm and economic loss for trade.Dignostic technique,reagent and vaccine for CSF are very important for its control.There are many diagnostic methods for classical swine fever which have their own advantages and disadvantages,while diagnostic reagents with a large variety of uneven quality.A standard to evaluating the diagnostic methods,monitoring the production process of diagnostic reagents and vaccines to control the product quality,ensuring the consistency and comparability of diagnosis is lack. At present,the establishment of veterinary biological reference standards has no preparation procedures, and the process is not clear.It is difficult to display the character and the purpose of reference standards because of the very few parameter values.So,it is necessary to develop and establish national reference standard serum for classical swine fever to solve these problems.This paper describes the study and preparation of national reference standard serum for classical swine fever,including raw materials selection,preparation and selection of candidates,research for the stability and collaborative study.Based on the experiments,the principle of the selection of raw materials is determined and the quality standard is formed too.Three batchs of classical swine fever-negative serum were prepared,and nine batchs of swine fever positive serum were also prepared by the basis of immunization and booster.After a series of analysis like physical character,sterility and specificity,et al,one batch of negative serum,weak positive serum,moderate and strong positive serum, coded reference standard A,B,C and D,are all distributed into containers(1 ml/ampoule),freeze-dried, vacuum sealed,which are chossed by the neutralization test in rabbits.Then we get 1580,1576,1581 and 1478 sealed glass ampoules for each batch of serum that physical character examination and purity test are perfect.Precision of filling for each batch serum are determined as 0.1641g with CV of 4.1%,0.1659g with CV of 3.6%,0.1672g with CV of 3.4%and 0.1644g with CV of 2.6%which can not comply with the requiers CV of 1%.Therefore further homogeneity studies will be required if high CV for precision of filling has any impact on the homogeneity.Residual moisture content is determined using Karl Fischer method,and the calculated value for each batch serum represents 2.35%,1.59%,1.68%and 1.82%mositure of total dry weight which complies with the requiers residual moisture content of 4%.The test conditions of the neutralization test in cells are optimized,and the homogeneity and stability of freeze-dried serum are tested by this method.No impact on homogeneity is found for high CV for precision of filling.Ampoules stored at elevated temperature 37℃for 7 months were tested in our laboratory which shows high stability.Last,collaborative study is carried out to demonstrate that the candidate national reference standard serum for CSF is suitable for its intended use and to assign potency by the neutralization test in cells and the neutralization test in rabbits after valuating in our laboratory.Then the raw datas from every participant are analyzed by statistical analysis,and the uncertainty of assay is introduced into the assignment of national reference standard serum for CSF to get more precise result with an fiducial range.National reference standard serum for CSF including reference standard A,B,C and D was assigned values in biological potency using the neutralization test in rabbits:reference standard A,0U/ml;reference standard B,6.3±1.8 U / ml; reference standard C,552±81 U / ml and reference standard D,1899±265 U / ml,and also was assigned values in biological potency using the neutralization test in cells:reference standard A,0U/ml; reference standard B,1.48±0.1 U / ml;reference standard C,502.2±62.8 U / ml and reference standard D,270.6±26.2 U/ml. |
PDF Full Text Request