小鼠组蛋白去乙酰化酶2多肽抗血清的制备

目的利用人工合成小鼠组蛋白去乙酰化酶2（histone deacetylase 2,HDAC2）多肽制备特异性抗HDAC2抗血清,用于相关疾病的体内外诊断。方法根据HDAC2基因编码的氨基酸序列合成多肽,与载体偶联后免疫动物,所制备的抗血清用ELISA、Western blot及免疫组织化学方法鉴定。结果 ELISA检测表明所制备的抗血清可同多肽抗原发生阳性反应,效价1∶4 000;Western blot结果显示抗血清可与多肽抗原及APP/PS1转基因小鼠的脑组织发生反应;免疫血清1∶100,1∶200,1∶400,1∶800四个稀释度均能与小鼠脑组织中的HDAC2反应。结论所制备的多肽抗血清可识别组织及血清中的HDAC2,可应用于相关领域的体内外研究。

Preparation and application of peptide antibody of histone deacetylase 2

Objective To prepare polyclonal antibody against mouse HDAC2 and identify its reactivity and specificity. Methods An anti-peptide antibody was developed by immunizing rabbits with a peptide synthesized according to the amino acid sequences of mouse HDAC2 protein. The peptide was coupled with keyhole limpet hemocyanin（KLH） and subcutaneously injected into rabbits. Sera were collected and the antibody activity was evaluated with ELISA, Western blot and immunohistochemistry. Results The titer of the antibody in ELISA was estimated as high as 1 ： 4 000. Western b]ot assay of the brain extracts from the APP/PSlmice and HDAC2 peptide showed a positive band at 80 kD when incubated with the antiserum. Immunohistochemistry staining showed that the antiserum got positive result with APP/PSlmice brain at titers of 1 ： 100, 1 ： 200, 1 ： 400, and 1 ： 800. Conclusions The prepared antisera in this study have specific reactivity with HDAC2 in serum and tissues, and may be applied in relevant in vivo and in vitro studies on related diseases.