Kayano T, Motoda R, Usui M, Ando S, Matuhasi T, Kurimoto M
Hayashibara Biochemical Laboratories, Inc., Okayama, Japan.
J Immunol Methods. 1990 Jun 12;130(1):25-31. doi: 10.1016/0022-1759(90)90295-7.
Rat-mouse hybridoma cells producing anti-mouse IgE antibodies were intraperitoneally or subcutaneously inoculated into newborn or suckling hamsters receiving rabbit anti-hamster thymocyte globulin from the day of birth twice a week for at least 3 weeks. The hybridoma cells were found to grow in the abdominal cavity of the hamsters as ascites tumor or in subcutaneous tissue as solid tumor without loss of antibody-secreting activities. For the production of ascites, 2-week-old hamsters were preferable to newborn hamsters. In 3-week-old hamsters, the hybridoma cells could scarcely survive. The antibody titers of the ascites were determined to be 10(5)-10(6) in the ELISA and in the ability to neutralize the skin-sensitizing capacity of mouse IgE antibodies. The rat monoclonal antibodies were easily separated from ascites, serum or cell culture supernatant with affinity chromatography using Affigel protein A-Sepharose and anti-hamster IgG-Sepharose columns. The described method could be efficiently applicable for the proliferation of other hybridomas, such as human-human, human-mouse or hamster-mouse, etc.
将产生抗小鼠IgE抗体的大鼠 - 小鼠杂交瘤细胞腹腔内或皮下接种到新生或哺乳的仓鼠中，从出生当天起每周两次给予兔抗仓鼠胸腺细胞球蛋白，持续至少3周。发现杂交瘤细胞在仓鼠腹腔内以腹水瘤形式生长，或在皮下组织中以实体瘤形式生长，且不丧失抗体分泌活性。对于腹水的产生，2周龄的仓鼠比新生仓鼠更合适。在3周龄的仓鼠中，杂交瘤细胞几乎无法存活。腹水的抗体效价在ELISA中测定为10(5)-10(6)，并且具有中和小鼠IgE抗体皮肤致敏能力。使用Affigel蛋白A - 琼脂糖和抗仓鼠IgG - 琼脂糖柱通过亲和色谱法可轻松从腹水、血清或细胞培养上清液中分离大鼠单克隆抗体。所描述的方法可有效应用于其他杂交瘤的增殖，如人 - 人、人 - 鼠或仓鼠 - 鼠等杂交瘤。
