Pajuaba Ana C A M, Silva Deise A O, Mineo José R
Laboratory of Immunoparasitology, Institute of Biomedical Sciences, Federal University of Uberlândia, 38400-902 Uberlândia, MG, Brazil
Clin Vaccine Immunol. 2010 Apr;17(4):588-95. doi: 10.1128/CVI.00444-09. Epub 2010 Feb 10.
This study aimed to evaluate the use of protein A-peroxidase (horseradish peroxidase [HRPO]) in indirect enzyme-linked immunosorbent assays (iELISAs) and IgG avidity assays for serological distinction between Brucella abortus S19-vaccinated and -infected cows. Four groups were analyzed: GI, 41 nonvaccinated seropositive cows; GII, 79 S19-vaccinated heifers analyzed at 3 months postvaccination; GIII, 105 S19-vaccinated cows analyzed after 24 months of age; and GIV, 278 nonvaccinated seronegative cows. IgG levels and avidity to B. abortus smooth lipopolysaccharide (S-LPS) were determined using anti-bovine IgG-HRPO or protein A-HRPO conjugates. Similar levels of IgG anti-S-LPS were found with GI using both conjugates. Lower IgG levels were detected with GII, GIII, and GIV using protein A-HRPO. Both conjugates showed high performance in discriminating GI from GIII, with high sensitivity (Se; 97.6%) and specificity (Sp; 97.1%). Protein A-HRPO was better in distinguishing GI from GIV (Se, 97.6%; Sp, 94.6%) and GI from GII (Se, 80.5%; Sp, 94.9%). Protein A-HRPO excluded a higher number of positive samples with GII and GIV. IgG avidity showed that protein A-HRPO, but not anti-IgG-HRPO, was able to distinguish nonvaccinated from vaccinated cattle, showing a higher avidity index (AI) with GI than with GII, with 78% of serum samples in GII showing an AI of <50%. Therefore, the iELISA using B. abortus S-LPS antigen and protein A-HRPO conjugate for preferential detection of the IgG2 subclass was shown to be suitable for serological distinction between S19-vaccinated and -infected cows. Also, antibodies generated after vaccination showed lower avidity, suggesting a role for the IgG2 subclass as an antibody of higher-affinity maturation after infection, constituting an additional tool for differentiating vaccinated from infected cattle.
本研究旨在评估蛋白A-过氧化物酶（辣根过氧化物酶[HRPO]）在间接酶联免疫吸附测定（iELISA）和IgG亲和力测定中用于区分接种布鲁氏菌流产疫苗S19和感染布鲁氏菌流产疫苗S19的奶牛的血清学差异。分析了四组：GI组，41头未接种疫苗的血清阳性奶牛；GII组，79头在接种疫苗后3个月进行分析的接种S19疫苗的小母牛；GIII组，105头在24月龄后进行分析的接种S19疫苗的奶牛；GIV组，278头未接种疫苗的血清阴性奶牛。使用抗牛IgG-HRPO或蛋白A-HRPO偶联物测定针对布鲁氏菌流产光滑脂多糖（S-LPS）的IgG水平和亲和力。使用两种偶联物在GI组中发现了相似水平的抗S-LPS IgG。使用蛋白A-HRPO在GII、GIII和GIV组中检测到较低的IgG水平。两种偶联物在区分GI组和GIII组时均表现出高性能，具有高灵敏度（Se；97.6%）和高特异性（Sp；97.1%）。蛋白A-HRPO在区分GI组和GIV组（Se，97.6%；Sp，94.6%）以及GI组和GII组（Se，80.5%；Sp，94.9%）方面表现更好。蛋白A-HRPO排除了GII组和GIV组中更多的阳性样本。IgG亲和力表明，蛋白A-HRPO而非抗IgG-HRPO能够区分未接种疫苗和接种疫苗的牛，GI组的亲和力指数（AI）高于GII组，GII组中78%的血清样本AI<50%。因此，使用布鲁氏菌流产S-LPS抗原和蛋白A-HRPO偶联物进行iELISA以优先检测IgG2亚类，被证明适用于区分接种S19疫苗和感染S19疫苗的奶牛的血清学差异。此外，接种疫苗后产生的抗体显示出较低的亲和力，这表明IgG2亚类作为感染后高亲和力成熟的抗体发挥作用，构成了区分接种疫苗和感染牛的另一种工具。
