Gangopadhyay A, Saravis C A, Kassis A I
Department of Radiology (Division of Nuclear Medicine) Harvard Medical School, Boston, Massachusetts 02115.
J Nucl Biol Med (1991). 1994 Mar;38(1):61-7.
Diethylenetriaminepentaacetic acid (DTPA)-conjugated purified rabbit anti-human serum albumin antibody was subjected to free-flow, semipreparative isoelectrophoresis using an apparatus that fractionates protein molecules based on isoelectric point (pI). The results indicate that (1) the apparatus was capable of developing linear pH gradients and separating proteins, (2) > or = 86% of the protein was recovered following fractionation, and (3) the protein concentration of the individual fractions varied. Focusing the fractions on an isoelectric slab gel revealed that the pIs of the isoforms were modified. The isoforms were radiolabeled with indium-111, their DTPA to antibody molar ratios determined, and their immunoreactivities evaluated by a solid-phase radioimmunoassay. The results demonstrate that (1) the molar ratio of DTPA to antibody varied among the fractions, and (2) the immunoreactivity of the majority of fractions was higher than that of unfractionated DTPA-antibody conjugate. Semipreparative isoelectric focusing may therefore improve the potential of radioimmunoconjugates in the radio-diagnosis and therapy of disease.
将二乙烯三胺五乙酸（DTPA）偶联的纯化兔抗人血清白蛋白抗体，使用基于等电点（pI）对蛋白质分子进行分离的仪器，进行自由流半制备等电聚焦电泳。结果表明：（1）该仪器能够形成线性pH梯度并分离蛋白质；（2）分级分离后蛋白质回收率≥86%；（3）各分级部分的蛋白质浓度有所不同。将各分级部分在等电平板凝胶上聚焦显示，同工型的pI发生了改变。用铟-111对同工型进行放射性标记，测定其DTPA与抗体的摩尔比，并通过固相放射免疫测定法评估其免疫反应性。结果表明：（1）各分级部分中DTPA与抗体的摩尔比不同；（2）大多数分级部分的免疫反应性高于未分级的DTPA-抗体偶联物。因此，半制备等电聚焦可能会提高放射免疫偶联物在疾病放射性诊断和治疗中的潜力。
