Roh Hang Sik, Song Hye Min, Yun Bo Reum, Kang Hyun Kyung, Choi Keum Suk, Park Yun Ju, Kim Dong Sub, Kim Seung Hee, Mo In Pil, An Beum-Soo, Ahn Chi Young
Biologics Research Division, National Institute of Food and Drug Safety Evaluation, Korea Food and Drug Administration, Cheongwon‑gun, Chungcheongbuk‑do 363‑951, Republic of Korea.
College of Veterinary Medicine, Chungbuk National University, Cheongju‑si, Chungcheongbuk‑do 361‑763, Republic of Korea.
Mol Med Rep. 2015 Apr;11(4):2819-24. doi: 10.3892/mmr.2014.3049. Epub 2014 Dec 4.
Single radial immunodiffusion (SRID) assay requires a reference antigen and an antibody to the hemagglutinin (HA) of an influenza vaccine. As it takes 2‑3 months to develop the reference antigen, vaccine development is delayed in cases of an influenza pandemic. In the present study, the measurement of the HA content of influenza vaccines was assessed using size exclusion high performance liquid chromatography (SE‑HPLC) for the rapid development of a pandemic vaccine. When the 2009 H1N1 reference antigen, pandemic 2009 H1N1 vaccine and 2010 seasonal influenza vaccines were analyzed by SE‑HPLC, the HA of the reference antigen and vaccines was specifically separated. The presence and specificity of HA were evidenced with immunoprecipitation and ELISA assays. For the influenza vaccines, the chromatogram pattern and retention time of HA were similar among the antigen types (2009 H1N1, 2010 H3N2 and 2010 B). In addition, when SE‑HPLC was applied, the ratio of HA chromatogram to peak area revealed a significant correlation with HA concentration for the reference antigen and vaccine. The result of the HA content calculation based on SE‑HPLC exhibited 99.91‑100% similarity, compared with that of SRID. These findings suggest that the measurement of peak area ratio/HA content using SE‑HPLC may be a substitute for SRID and rapidly measure HA content to enable faster development of a vaccine during an influenza pandemic.
单向辐射免疫扩散（SRID）测定需要一种参考抗原和针对流感疫苗血凝素（HA）的抗体。由于开发参考抗原需要2至3个月的时间，在流感大流行的情况下，疫苗开发会延迟。在本研究中，为了快速开发大流行疫苗，使用尺寸排阻高效液相色谱法（SE-HPLC）评估流感疫苗的HA含量。当通过SE-HPLC分析2009年H1N1参考抗原、2009年大流行H1N1疫苗和2010年季节性流感疫苗时，参考抗原和疫苗的HA被特异性分离。通过免疫沉淀和ELISA测定证明了HA的存在和特异性。对于流感疫苗，不同抗原类型（2009年H1N1、2010年H3N2和2010年B）的HA色谱图模式和保留时间相似。此外，当应用SE-HPLC时，HA色谱图与峰面积的比值与参考抗原和疫苗的HA浓度显示出显著相关性。基于SE-HPLC的HA含量计算结果与SRID相比显示出99.91%-100%的相似性。这些发现表明，使用SE-HPLC测量峰面积比/HA含量可能替代SRID，并快速测量HA含量，以便在流感大流行期间更快地开发疫苗。
