Isolation of high-affinity antibodies from hyperimmune serum

A group of high-affinity antibodies has been isolated from horse hyperimmune antiplague serum with the use of antigenic immunosorbent. High-affinity antibodies have proved to be homogeneous, which is indicated by the rectilinear character of the graph of the inverse values of the free antigen concentration.     

Use of hyperimmune serum in treatment of endometritis in mares

Endometritis was induced in four progesterone-treated ovariectomized mares and in two seasonally anestrous mares by intrauterine infusion of Streptococcus zooepidemicus. The bacteria were suspended in phosphate buffered saline (PBS), or in nonimmune or hyperimmune serum. Uterine lavage was performed after 24 h. Significantly fewer (P < 0.05) bacteria were recovered from mares which received hyperimmune serum than from mares which received nonimmune serum or PBS. There was no significant difference between treatment with nonimmune serum or PBS. It is therefore suggested that increasing the availability of specific antibody reduced the severity of intrauterine infection.     

Production and preparation of hyperimmune bovine colostrum for passive immunotherapy of cryptosporidiosis.

Pregnant cows were immunized to produce hyperimmune bovine colostrum (HBC) by intramuscular injection or intramammary infusion (TI) followed by 3 successive TI boosters with Cryptosporidium parvum (Cp) oocyst antigen mixed with Freund's (F) or Ribi (R) adjuvant. Control cows received no Cp. Colostrum from all cows was skimmed of butterfat and tested for specific anti-Cp immunoglobulin isotypes by ELISA. The HBC from Cp-F and Cp-R immunized cows had IgG1 titers exceeding 1:400,000 and 1:800,000, respectively. Some HBC from Cp-F immunized cows was freeze-dried to facilitate storage and some were irradiated at 42.5 kGy to kill potentially contaminating pathogens. Freeze-drying, but not irradiation, reduced IgG1 titers by only one dilution. Neither treatment affected Western blot banding patterns.     

Passive protection across subgroups of alphaviruses by hyperimmune non-cross-neutralizing anti-Sindbis serum

Extended hyperimmunization of rabbits with Sindbis (SIN) or Semliki Forest (SF) viruses causes the production of antisera that are cross-reactive with virus-infected cells in antibody-dependent, complement-mediated cytotoxicity assays but that do not cross-neutralize viruses in vitro. C3H/HeJ mice given gamma globulin fractionated from the extended hyperimmune antiserum against SIN, but not control sera, were protected from challenge by 100 LD50 of SF, a virus which is in a different subgroup than SIN. All mice survived if the gamma globulin was given 24 hr before challenge virus and partial protection occurred if the globulin was given 24 hr after the virus. Cobra venom factor treatment of normal C3H mice challenged with SF did not reduce the protection, suggesting that complement was not involved. Methyl palmitate (40 mg/mouse) given before gamma globulin and virus challenge suppressed macrophage activity and reduced the level of protection 23% in females and 70% in males. Silica treatment (3 mg/mouse) reduced the protection equally in both males and females by 92%. In vitro experiments were done to test if it were possible that cross-antibody-dependent cellular cytotoxicity (ADCC) could account for the passive cross-protection observed in this system. Cross-ADCC could be demonstrated in vitro at high dilutions of antiserum (1:25,600). On the basis of the in vitro and in vivo results presented, we suggest that cross-ADCC against SF-infected target cells is one of the likely mechanisms to explain the passive cross-protection observed.     

Determination of total protein in hyperimmune serum samples by near-infrared spectrometry and multivariate calibration

In the present work, the determination of the total protein concentration in hyperimmune serum samples was performed through a partial least-squares near-infrared (NIR-PLS) method. The method was based on the chemometric treatment of the NIR spectra of samples. The influences of spectra preprocessing and spectral window utilized in the construction of PLS model were studied. Models were built using reference data of 19 samples selected through the use of hierarchical cluster analysis (HCA) of NIR spectra of samples and another 24 samples were employed for external validation of the method. A model with better prediction capacity was obtained after whole spectra preprocessing by multiplicative scattering correction (MSC) algorithm and using data in the spectral range of 2158-2209 nm. Under optimized conditions a RMSEP of 0.21 g dl⁻¹ and a quality coefficient value (QC) of only 5.8% were obtained for the prediction of total protein content in the samples used for external validation. Also, a determination coefficient, r², of 0.97 was obtained in the correlation of predicted and reference data of samples situated in the validation set.     

尼帕病毒融合蛋白、受体结合蛋白的表达及特异性高免血清制备

尼帕病毒膜融合蛋白F和受体结合蛋白G在病毒感染和诱导机体产生保护性免疫中起重要的作用。通过PCR扩增获得尼帕病毒F1和G基因片段(均去掉信号肽和跨膜区),克隆至原核表达载体,IPTG诱导大肠杆菌表达目的蛋白,Western blot表明重组F1、G蛋白与兔抗尼帕病毒血清具有良好的反应原性;同时将F1和G基因克隆至经改造过的杆状病毒表达载体,获得了含有目的基因的重组杆状病毒,接种sf9单层细胞,间接免疫荧光检测表明F1、G蛋白在杆状病毒中正确表达,并与抗尼帕病毒血清具有良好的反应原性。以纯化原核表达的F1、G蛋白免疫兔获得了抗F1和抗G重组蛋白的特异血清,Western blot和间接免疫荧光检测表明所制备的血清具有特异性。试验所表达的抗原和制备的特异血清可用于尼帕病的诊断。