Medical Affairs and Innovation, Héma-Québec, Québec, Quebec, Canada.
Center for Innovation, Canadian Blood Services, Toronto, Ontario, Canada.
Transfusion. 2020 Sep;60(9):2090-2096. doi: 10.1111/trf.15932. Epub 2020 Jul 6.
Treatment of red blood cells with dithiothreitol (DTT) or trypsin effectively denatures CD38; however, this treatment damages other antigens, some of which are of clinical importance. Thus, other avenues to deplete daratumumab (DARA) from plasma samples should be explored.
The Daudi B-cell line was found to express high levels of CD38 and was sonicated in a sonication buffer to achieve complete cell lysis. The resulting stroma preparation was centrifuged at 20 000g for 20 minutes and then mixed with 250 μL of DARA-plasma and incubated for 10 minutes at 37°C. The stroma-DARA-plasma mixture was centrifuged again, and the supernatant was collected and subjected to four additional rounds of adsorption with fresh stroma. DARA-depleted plasma was tested by gel indirect antiglobulin test (IAT).
CD38 expression on Daudi cells was confirmed by flow cytometry. Gel IAT analysis showed that the incubation of plasma from DARA-treated patients with Daudi cells stroma resulted in a significant depletion of DARA but allowing detection of other alloantibodies of interest such as anti-K, anti-Yt , and anti-Gy .
Daudi cell stroma is inexpensive, easy to prepare in large batches, and can be used as an off-the-shelf reagent. Incubation of plasma from DARA-treated patients with Daudi cell stroma can efficiently overcome DARA interference in serologic testing without affecting DTT- or trypsin-sensitive antigens.
用二硫苏糖醇（DTT）或胰蛋白酶处理红细胞可有效使 CD38 变性；然而，这种处理会损害其他抗原，其中一些具有临床意义。因此，应该探索其他去除达雷木单抗（DARA）的方法。
发现 Daudi B 细胞系表达高水平的 CD38，并在超声缓冲液中进行超声处理以实现完全细胞裂解。将得到的基质制备物在 20,000g 下离心 20 分钟，然后与 250μL 的 DARA-血浆混合，并在 37°C 下孵育 10 分钟。再次离心基质-DARA-血浆混合物，收集上清液并通过新鲜基质进行另外四轮吸附。通过凝胶间接抗球蛋白试验（IAT）测试 DARA 耗尽的血浆。
通过流式细胞术证实了 Daudi 细胞上的 CD38 表达。凝胶 IAT 分析表明，用 Daudi 细胞基质孵育来自 DARA 治疗患者的血浆会导致 DARA 的显著耗尽，但仍能检测到其他感兴趣的同种异体抗体，如抗-K、抗-Yt 和抗-Gy。
Daudi 细胞基质价格低廉，易于大量制备，并可作为即用型试剂。用 Daudi 细胞基质孵育来自 DARA 治疗患者的血浆可以有效地克服 DARA 在血清学检测中的干扰，而不影响 DTT 或胰蛋白酶敏感抗原。
