Fiddler M B, Hudson L D, White J G, Desnick R J
J Lab Clin Med. 1980 Aug;96(2):307-17.
Seven methods of erythrocyte entrapment--six by hypotonic exchange loading and one by chlorpromazine-induced endocytosis--were evaluted for (1) efficiency of incorporation of beta-glucuronidase, inulin, and glucose; (2) presence of beta-glucuronidase on the erythrocyte surface, as detected by hemagglutination or complement-dependent lysis in the prsence of antibody to beta-glucuronidase; (3) in vitro leakage of entrapped markers; and (4) morphologic alterations by scanning electron microscopy. Dependent on the method of entrapment, the incorporation of markers ranged from 0.7% to 6.2% for beta-glucuronidase, 3.4% to 31% for inulin, and 1.2% to 12.2% for glucose. Maximal incorporation by hypotonic exchange loading occurred when the initial concentration of sodium chloride (150 mM) was reduced to 50 or 75 mM. However, beta-glucuronidase was detected on the erythrocyte surface by hemagglutination for these methods as well as a dialysis method of loading. Essentially no leakage of entrapped enzyme was detected (< 1%) for all methods, although up to 11% of entrapped glucose was released during a 3 hr incubation at 37 degrees C in buffered whole blood. Finally, entrapment methods requiring the greatest reduction in salt concentration resulted in the formation of echinocytes, whereas stomatocytes were observed after entrapment by methods requiring lesser salt dilutions. These results demonstrate the efficiency of entrapment and relative integrity of erythrocytes following various loadng procedures and suggest that in vitro assessment may provide a useful predictor of the imunogenicity and in vivo fate of erythrocyte-entrapped enzymes or other therapeutic agents.
对七种红细胞包封方法进行了评估，其中六种是通过低渗交换加载，一种是通过氯丙嗪诱导的内吞作用，评估内容包括：(1)β-葡萄糖醛酸酶、菊粉和葡萄糖的包封效率；(2)通过血细胞凝集或在存在抗β-葡萄糖醛酸酶抗体时的补体依赖性裂解检测红细胞表面β-葡萄糖醛酸酶的存在；(3)包封标记物的体外渗漏；(4)通过扫描电子显微镜观察形态学改变。根据包封方法的不同，β-葡萄糖醛酸酶的标记物包封率在0.7%至6.2%之间，菊粉在3.4%至31%之间，葡萄糖在1.2%至12.2%之间。当氯化钠初始浓度(150 mM)降至50或75 mM时，通过低渗交换加载实现最大包封率。然而，对于这些方法以及透析加载方法，通过血细胞凝集在红细胞表面检测到了β-葡萄糖醛酸酶。对于所有方法，基本上未检测到包封酶的渗漏(<1%)，尽管在37℃的缓冲全血中孵育3小时期间，高达11%的包封葡萄糖被释放。最后，需要最大程度降低盐浓度的包封方法导致棘红细胞形成，而在需要较小盐稀释度的方法包封后观察到口形红细胞。这些结果证明了各种加载程序后红细胞包封的效率和相对完整性，并表明体外评估可能为红细胞包封酶或其他治疗剂的免疫原性和体内命运提供有用的预测指标。
