微量抗原脾内包理免疫法制多CC－1的第二代单克隆抗体

PRODUCTION CHARACTERIZATON OF CC-1 SECONDGENERATION MONOCLONAL ANTIBODIES BY INTRASPLENICDEPOSITED IMMUNlZATION WITH MINUTE AMOUNTS OFANTIGEN

采用革克隆抗体ＣＣ－１（ＩｇＭ）免疫组化阳性的新鲜大肠癌组织制备粗抗原，经ＳＤＳ－ＰＡＧＥ分离，Ｗｅｓｔｅｒｎ－ｂｌｏｔ后进行免疫染色，检测刮ＣＡ－ＣＣ－１带。用附载ＣＡ－ＣＣ－１的硝酸纤维膜（ＮＣ）进行二次小鼠脾内包埋免疫和一次腹腔粗抗原加强免疫，成功地制备了两株ＣＣ－１的第二代单抗Ｗ１３５（ＩｇＭ）与Ａ１２－２（ＩｇＧｌ）。亲和力均比原代高；抗原决定族为糖链结构；Ｗｅｓｔｅｒｎ－ｂｌｏｔ免疫染色的比较发现：Ｗ１３５与Ａ１２－２除能识别ＣＡ－ＣＣ－１带外，尚能分别识别分子量为９４ＫＤ左右和３３ＫＤ的抗原带。免疫组化比较表明：对于大肠癌阳性反应率，Ａ１２－２基本与ＣＣ－１相同，而Ｗ１３５明显优于ＣＣ－１。以上研究表明：脾内包埋免疫法为一行之有效的免疫方式；第二代单抗可能较原代ＣＣ－１具有更大的临床应用价值。

he crude materials, taking from human coloilic carcinoma biopsy which reacted positivelywith monocloial antibody CC-1(IgM),were resolved by SDS-PAGE and transboltted to nitro-cellulose paper.The CA-CC -1 binding zone was found beyoiid the 94KD by immunostaining.Twice intrasplenic immunization with CA-CC-1-NC paper followed by an intraperitoneal boost with crude antigen raised two 2nd generation McAbs,i.e,W135(IgM)aidA12-2(IgGl).Bothof them had higher affinity than CC-1.Western-blot analysis showed that W135 and A12-2could not only recognize the CA-CC-1, but they also recognize other two glycoprotein zones,m. w. about 94KD and 33KD respectively.In test for coloilic carciiiomas with immunohis-to chemistry,the reactivities of W135 was superior to CC-1(42%)and A12-2(46%).This studythus provides evidence that the intraplenic immunization is a effective immunization route and these second generation McAbs may be more efficient than CC-1 in clinical diagnosis and thera-py of colorectal carcinoma.